CN116745418A - 用于等位基因的rna编码的dna替换的组合物和方法 - Google Patents

用于等位基因的rna编码的dna替换的组合物和方法 Download PDF

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CN116745418A
CN116745418A CN202180088340.2A CN202180088340A CN116745418A CN 116745418 A CN116745418 A CN 116745418A CN 202180088340 A CN202180088340 A CN 202180088340A CN 116745418 A CN116745418 A CN 116745418A
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crispr
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nucleic acid
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cas
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Y·金
J·M·瓦特斯
A·哈默尔
S·J·拉维特
D·施瓦克
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Pairing Plant Service Co ltd
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Abstract

本发明涉及包含CRISPR‑Cas效应蛋白、逆转录酶和延伸型指导核酸的重组核酸构建体，以及使用其来在植物中修饰核酸的方法。

Description

用于等位基因的RNA编码的DNA替换的组合物和方法
关于电子提交序列表的声明
代替纸质副本，提供了以ASCII文本格式的序列表，其按照37C.F.R.§1.821递交，名称为1499.57.WO_ST25.txt，大小为1,043,362个字节，于2021年11月5日生成，并且经由EFS-Web提交。该序列表通过关于其公开内容在说明书中提及而特此合并入本文。
优先权声明
本申请按照35U.S.C.§119(e)要求于2020年11月6日提交的美国临时申请号63/110,386(其整个内容通过提及而合并入本文)的权益。
发明领域
本发明涉及包含CRISPR-Cas效应蛋白、逆转录酶和延伸型指导核酸的重组核酸构建体，以及使用其来在植物中修饰核酸的方法。
发明背景
碱基编辑已显示出是分别将胞嘧啶和腺嘌呤残基改变为胸腺嘧啶和鸟嘌呤的有效方式。这些工具虽然强大，但确实有一些限制，例如旁观者碱基，小的碱基编辑窗口(其给出有限的对于性状相关靶标的可及性，除非具有高PAM密度的酶可用于进行补偿)，有限的分别将胞嘧啶和腺嘌呤转变为除了胸腺嘧啶和鸟嘌呤之外的其他残基的能力，以及没有编辑胸腺嘧啶或鸟嘌呤残基的能力。因此，目前的可用于碱基编辑的工具是有限的。因此，为了通过对于更多数目的生物增加可能编辑的范围来使得核酸编辑更有用，需要新的编辑工具。
发明概述
在第一个方面，提供了修饰靶核酸的方法，所述方法包括：使所述靶核酸与下列组分相接触：(a)V型CRISPR-Cas效应蛋白或II型CRISPR-Cas效应蛋白；(b)逆转录酶，和(c)延伸型指导核酸(extended guide nucleic acid)(例如，延伸型II型或V型CRISPR RNA、延伸型II型或V型CRISPR DNA、延伸型II型或V型crRNA、延伸型II型或V型crDNA)，由此修饰所述靶核酸。
在第二个方面，提供了修饰靶核酸的方法，所述方法包括：使所述靶核酸在第一位点处与下列组分相接触：(a)(i)第一CRISPR-Cas效应蛋白；和(ii)第一延伸型指导核酸(例如，延伸型CRISPR RNA、延伸型CRISPR DNA、延伸型crRNA、延伸型crDNA)；和(b)(i)第二CRISPR-Cas效应蛋白，(ii)第一逆转录酶；和(ii)第一指导核酸，由此修饰所述靶核酸。
在第三个方面，提供了在植物或植物细胞中修饰靶核酸的方法，其包括将本发明的表达盒引入到所述植物或植物细胞中，由此在所述植物或植物细胞中修饰所述靶核酸，并且产生包含所述经修饰的靶核酸的植物或植物细胞。
在第四个方面，提供了复合物，其包含：(a)V型CRISPR-Cas效应蛋白或II型CRISPR-Cas效应蛋白；(b)逆转录酶；和(c)延伸型指导核酸(例如，延伸型CRISPR RNA、延伸型CRISPR DNA、延伸型crRNA、延伸型crDNA，例如所靶向的等位基因指导(tag)核酸(即，tagDNA、tagRNA))。
在第五个方面，提供了经密码子优化以用于在生物中进行表达的表达盒，所述表达盒以5'至3'包含：(a)编码植物特异性启动子序列(例如，ZmUbi1、MtUb2、RNA聚合酶II(Pol II))的多核苷酸，(b)经植物密码子优化的编码V型CRISPR-Cas核酸酶(例如，Cpf1(Cas12a)、dCas12a等)的多核苷酸；(c)连接体序列；和(d)经植物密码子优化的编码逆转录酶的多核苷酸。
在第六个方面，提供了经密码子优化以用于在生物中进行表达的表达盒，所述表达盒包含：(a)编码启动子序列的多核苷酸，和(b)延伸型RNA指导序列，其中所述延伸型指导核酸包含延伸部分，所述延伸部分在其3'末端处包含引物结合位点和待被掺入到所述靶核酸中的编辑(例如，逆转录酶模板)，任选地其中所述延伸型指导核酸被包含在表达盒中，任选地其中所述延伸型指导核酸可操作地连接至Pol II启动子。
本发明进一步提供了包含通过本发明的方法进行修饰的靶核酸的细胞，包括植物细胞、细菌细胞、古细菌细胞、真菌细胞、动物细胞，以及包含所述细胞的生物，包括植物、细菌、古细菌、真菌和动物。另外，本发明提供了包含本发明的多核苷酸、多肽和表达盒的试剂盒。
本发明的这些和其他方面在下面的发明描述中更详细地进行阐述。
附图简述
图1提供了示意图，其显示了从crRNA的逆转录生成DNA序列并且随后整合到切口位点中。延伸型指导crRNA(tagRNA)与Cpf1切口酶(cas12a切口酶)(nCpf1，左上)相结合。备选地，编码编辑模板的延伸可以位于crRNA的5'。crRNA的3'末端与DNA在所述切口位点处互补(非粗体配对线，左上)。nCpf1可以共价连接至逆转录酶(RT)，或者RT可以被募集至nCpf1，在这种情况下多个逆转录酶蛋白可以被募集至nCpf1。RT在第二链上从DNA切口的3'末端开始对DNA进行聚合，从而生成与crRNA互补的DNA序列，具有与基因组不互补的核苷酸(加粗的配对线，大括号，右上)，随后为互补的核苷酸(非粗体配对线，右上)。解离后，所得的DNA具有延伸型ssDNA，其具有3'突出端，其是与原始DNA(非加粗的配对线，右下)在很大程度上相同的序列，但是具有一些非天然核苷酸(加粗的配对线，大括号，右下)。该翼片(flap)与具有5'突出端的结构(左下)处于平衡状态，其中存在掺入到DNA中的错配的核苷酸。该平衡可以通过减少错配修复、在修复和复制期间去除5'翼片和还有对第一链产生切口(如本文中所描述的)而被驱动朝向左边的结构。
图2提供了示意图，其显示了用于减少错配修复的方法。为了驱动所述平衡更有利于形成具有经修饰的核苷酸(加粗的，大括号)的最终产物，引导切口酶(经由指导核酸)在RT编辑区域之外的区域(闪电箭头)中-离在第二链(例如，靶标链或上部链)中的切口一定距离-切割靶核酸的第一链(例如，靶标链或下部链)。nCpf1:crRNA分子可以在编辑泡的任何一侧或两侧。对第一链(虚线)产生切口向细胞表明，在错配修复和复制期间，新掺入的核苷酸是正确的核苷酸，因此有利于具有新核苷酸的最终产物。驱动所述平衡朝向所希望的产物的其他可能方式可以包括去除5'翼片。
图3显示了备选的通过使用本发明的组合物来修饰核酸的方法，其中将两个切口引入到第二链中，并且由RT所引入的序列取代带双切口的WT序列并且由此更有效地被掺入到基因组中。
图4.LbCas12a_R1138A为切口酶，如在体外所证实的，在1％TAE-琼脂糖凝胶上进行解析。超螺旋的2.8kB质粒以2.0kB的表观大小进行走样(泳道2)，直至由野生型LbCas12a生成双链断裂(泳道3)。
图5显示了在大肠杆菌(E.coli)中测试的REDRAW编辑器的构造(参见实施例1)。
图6显示了在第一文库中测试的tagRNA的构造。
图7显示了用于在REDRAW编辑中使用的实例性的所设计的发夹序列(SEQ ID NO:203)的结构。
图8显示了桑格(Sanger)测序结果，其证实了在去功能的aadA基因中的TGA>CTG编辑，从而恢复了抗生素抗性(SEQ ID NO:204-208)。从在选择10中的菌落观察到所述编辑，其中具有蛋白质构造SV40-MMLV-RT-XTEN-nLbCas12a-SV40(SEQ ID NO:71)。
图9显示了桑格测序结果，其证实了在大肠杆菌基因组中的rpsL基因中的AAA>CGT编辑，从而赋予对于抗生素链霉素的抗性(SEQ ID NO:209-211)。从在选择2.5中的菌落观察到所述编辑，其中具有蛋白质构造SV40-MMLV-RT-XTEN-nRVRLbCas12a(H759A)-SV40(SEQID NO:79)。
图10显示了桑格测序结果，其证实了在去功能的aadA基因中的TGA>GAT编辑，从而恢复了抗生素抗性(SEQ ID NO:212-215)。从在选择2.25中的菌落观察到所述编辑，其中具有蛋白质构造SV40-nLbCas12a-XTEN-MMLV-RT-SV40(SEQ ID NO:73)。
图11显示了桑格测序结果，其证实了在去功能的aadA基因中的TGA>GAT编辑，从而恢复了抗生素抗性(SEQ ID NO:212-215)。从在选择2.31中的菌落观察到所述编辑，其中具有蛋白质构造SV40-MMLV-RT-XTEN-nLbCas12a(H759A)-SV40(SEQ ID NO:83)。
图12显示了在人细胞中进行的实例性的编辑方法(参见实施例2)。图版A显示了双链靶核酸。Cas12a复合物(复合物包括延伸型指导核酸，其未被显示)被募集至第一链(靶标链，下部链)，其中在第二链(上部链，非靶标链)中的5'翼片任选地用5'-3'核酸外切酶来去除(图版B)。图版C显示了逆转录酶MMuLV-RT(5M)(SEQ ID NO:53)在靶核酸上从引发位点或引物(与引物结合位点互补)开始进行延伸(虚线＝延伸)。图版D和E显示了经由错配修复和DNA连接的DNA中间体的分解以及新的经编辑的DNA链的生成。
图13显示了在HEK293T细胞中在FANCF1位点处使用各种指导构象的精确编辑。构建体名称为Cas12a(H759A)+RT(5M)+RecE FANCF1。
图14显示了在HEK293T细胞中在DMNT1位点处使用各种指导构象的精确编辑。构建体名称为Cas12a(H759A)+RT(5M)。
图15显示了核酸外切酶转染对于在DMNT1位点处的精确编辑活性(标准化至无核酸外切酶处理；pUC19＝1)的效应。
图16显示了各种形式的REDRAW建构(即，本发明的构建体)以及每一个的精确编辑百分比。左图版显示了以反式提供的逆转录酶(RT)(无募集)。中间图版显示了RT的募集，其中作为例子，使用融合至LbCpf1(LBCas12a)的C-末端的SunTag(例如，GCN4,例如SEQ IDNO:23)(LBCpf1-SunTag)，其可以募集融合至RT(5M)的N-末端的抗体(scFv-RT(5M))(例如，scFv，SEQ ID NO:25)。右图版显示了RT和lbCpf1融合蛋白。右图版的左侧显示了用融合至LbCpf1的C-末端的RT的结果，和右图版的右侧显示了用融合至LbCpf1的N-末端的RT的结果。
图17提供了使用5'-3'核酸外切酶来在REDRAW过程期间生成的双链断裂的两个末端处降解DNA的示意图。
图18显示了使用融合至Cas多肽(LbCpf1)的C-末端的5'-3'核酸外切酶(RecE(SEQID NO:129)、RecJ(SEQ ID NO:130)、T5_Exo(SEQ ID NO:131)、T7_Exo(SEQ ID NO:132))的REDRAW的精确编辑百分比。在该构造中，RT(5M)(SEQ ID NO:53)以反式进行表达(无募集)。
图19显示了使用5'-3'核酸外切酶sbcB(SEQ ID NO:134)或5'-3'核酸外切酶Exo(SEQ ID NO:135)(各自融合至Cas多肽(LbCpf1)的C-末端)的REDRAW的精确编辑百分比。RT(5M)以反式进行表达(无募集)。
图20显示了使用核酸外切酶的反式表达的REDRAW的精确编辑百分比。LbCpf1和RT以融合蛋白来提供。图20的右侧显示了用融合至LbCpf1的N-末端的RT(RT(5M)-LbCpf1(H759A))的结果，和该图的左侧显示了使用融合至LbCpf1的C-末端的RT(LbCpf1(H759A)-RT(5M))的结果。
图21显示了在REDRAW过程中在Cas12a(LbCpf1)中的实例性的突变对于REDRAW的精确编辑百分比的效应。所测试的实例性的突变包括K167A、K272A、K349A、K167A+K272A、K167A+K349A、K272A+K349A和K167A+K272A+K349A(相对于LbCas12a(H759A)SEQ ID NO:148而言的位置)。
图22显示了在单链DNA结合蛋白(ssDNA BP)存在下REDRAW的精确编辑百分比。ssDNA BP在CRISPR-Cas效应多肽(例如，LbCpf1(H759A))、RT(5M)和tagRNA1存在下以反式进行表达。在该实例中，RT和LbCpf1(H759A)也以反式进行表达。所测试的ssDNA BP为hRad51_s208E_A209D、hRad52、BsRecA、EcRecA和T4SSB。模拟物为无ssDNA BP。
图23显示了当融合至CRISPR-Cas效应多肽(例如，LbCas12a H759A)时在单链DNA结合蛋白(ssDNA BP)存在下REDRAW的精确编辑百分比。将ssDNA结合蛋白(hRad51、hRad52、BsRecA、EcRecA、T4SSB和Brex27)融合至LbCpf1(H759A)的N-末端或C-末端。RT(5M)和tagRNA以反式进行表达。
图24显示了对于当在阻止NHEJ的多肽存在下进行REDRAW时所产生的插入/缺失(indel)的百分比的效应。在该实例中，所述阻止NHEJ的多肽为Gam蛋白(埃希氏菌(Escherichia)噬菌体Mu Gam蛋白)(SEQ ID NO:147)，并且所述逆转录酶以反式进行表达，要么作为天然序列(例如，RT(5M))要么与融合至RT的N-末端的Gam(例如，Gam-RT(5M))一起。将这些构建体与LbCas12a(H759A)或者与具有融合至其N-末端的Gam蛋白的LbCas12a(H759A)(例如，Gam-LbCas12a H759A)同时地进行表达。
图25显示了在Gam蛋白存在下REDRAW的精确编辑百分比。Gam蛋白以反式来提供，作为与逆转录酶的融合蛋白(N-末端融合；Gam-RT(5M))和/或作为与CRISPR-Cas效应多肽的融合蛋白(例如，Gam-LbCas12a H759A)。
图26显示了使用不同长度引物结合位点(PBS)和逆转录酶模板(RTT)的REDRAW的精确编辑百分比。上图版和下图版显示了使用两种不同的间隔子的结果(上图版：pwsp143(GCTCAGCAGGCACCTGCCTCAGC)(SEQ ID NO:136)，下图版：pwsp139(CTGATGGTCCATGTCTGTTACTC)(SEQ ID NO:137))。
图27显示了在两种不同的逆转录酶模板(RTT)中取决于编辑的位置的编辑百分比。在每个RTT中将所述编辑放置在从位置-1至位置19(编号是相对于在靶核酸中的原间隔子邻近基序(protospacer adjacent motif)编号而言的)变动的位置处(编辑以粗体)。上图版中的RTT：TTTGGCTCACTCCTGCTCGGTGAATTT SEQ ID NO:187；下图版中的RTT：TTTCGCGCTTGTTCCAATCAGTACGCA SEQ ID NO:188。
图28显示了使用两种形式的Cas9即核酸酶(Cas9)和切口酶(nCas9(D10A突变体))的REDRAW的精确编辑百分比。Cas9和nCas9均通过使用具有附接至指导RNA的3'末端或5'末端的延伸(称为3'延伸或5'延伸)的tagRNA来进行测试。所述tagRNA延伸的RTT和PBS的长度是变化的，并且所述间隔子靶向四个不同的位点(pwsp10：GAGTCCGAGCAGAAGAAGAA(SEQ IDNO:140)；pwsp621：GCATTTTCAGGAGGAAGCGA(SEQ ID NO:141)；pwsp15：GTCATCTTAGTCATTACCTG(SEQ ID NO:142)；pwsp11：GGAATCCCTTCTGCAGCACC(SEQ ID NO:143))。
图29显示了使用BhCas12b的REDRAW的精确编辑百分比。BhCas12b通过使用具有附接至指导RNA的3'末端或5'末端的延伸(称为3'或5')的tagRNA来进行测试。所述tagRNA延伸的RTT和PBS的长度是变化的，并且所述间隔子靶向三个不同的位点(PWsp1099：ACGTACTGATGTTAACAGCTGA(SEQ ID NO:144)；PWsp1098：GGTCAGCTGTTAACATCAGTAC(SEQ IDNO:145)；PWsp1094：TCCAGCCCGCTGGCCCTGTAAA)(SEQ ID NO:146))。
图30显示了使用EnAsCpf1(H800A)(SEQ ID NO:149)的REDRAW的精确编辑百分比。左图版显示了没有RT(5M)的编辑，中间图版显示了用具有C-末端融合的RT(5M)的EnAsCpf1(H800A)(EnAsCpf1(H800A)-RT(5M))的编辑，和右图版显示了用具有N-末端融合的RT(5M)的EnAsCpf1(H800A)(RT(5M)-EnAsCpf1(H800A))的编辑。在该实例中，单一位点用具有CCTCACTCCTGCTCGGTGAATTT(SEQ ID NO:171)的序列的间隔子来进行靶向。
图31显示了通过使用本发明的方法(REDRAW)对于在酵母中的URA3-1靶基因的编辑结果。上版图显示了使用具有融合至其C-末端的逆转录酶(RT)的LbCas12a的编辑结果(在通过编辑而修复腺嘌呤营养缺陷后的菌落形成)。下图版显示了使用具有融合至其N-末端的RT的LbCas12a的编辑结果(在通过编辑而修复腺嘌呤营养缺陷后的菌落形成)。用于在图31中所显示的编辑的延伸型指导不具有假结(pseudoknot)或者在其3'末端处包含假结。所述假结称为诱饵发夹(SEQ ID NO:95；SEQ ID NO:203)、tEvoPreQ1(SEQ ID NO:158)或EvoPreQ1(SEQ ID NO:191)。所述延伸型指导进一步包含具有47、55或63个核苷酸的长度的RTT和具有48个核苷酸的长度的PBS。
图32显示了通过使用本发明的方法(REDRAW)对于在酵母中的ADE2靶基因的编辑结果。上版图显示了使用具有融合至其C-末端的RT的LbCas12a的编辑结果(在通过编辑而修复尿嘧啶营养缺陷后的菌落形成)。下图版显示了使用具有融合至其N-末端的RT的LbCas12a的编辑结果(在通过编辑而修复尿嘧啶营养缺陷后的菌落形成)。用于在图32中所显示的编辑的延伸型指导不具有假结(pseudoknot)或者在其3'末端处包含假结。所使用的假结称为诱饵发夹(SEQ ID NO:95，SEQ ID NO:203)、tEvoPreQ1(SEQ ID NO:158)或EvoPreQ1(SEQ ID NO:191)。所述延伸型指导进一步包含具有40、50或72个核苷酸的长度的RTT和具有48个核苷酸的长度的PBS。通常，所述延伸型指导核酸以5'-3'包含RTT、PBS和(当存在时)3'假结。在关于诱饵发夹的第一列数据中，在上图版和下图版中，具有40-bp RTT和诱饵发夹的tagRNA均未能合成并且所述条件未进行测试。
图33显示了当使用各自融合至RT-LbCas12融合蛋白(例如，RT-LbCas12a)的N-末端的ssRNA结合蛋白(防卫素(SEQ ID NO:152)和ORF5(SEQ ID NO:153))时的精确编辑百分比结果，相比于不包含在其N-末端处融合的ssRNA结合蛋白的相同的RT-Cas12a融合蛋白而言。
图34显示了当使用在其N-末端处融合至具有不同突变的逆转录酶(RT)结构域的LbCas12a(H759A)时的精确编辑百分比结果。所述RT包括：RT(L139P,D200N,W388R,E607K)、RT(L139P,D200N,T306K,W313F,W388R,E607K)、RT(5M,F155Y,H638G)、RT(5M,Q221R,V223M)和RT(5M,D524N)。
图35显示了使用四种不同的tagRNA(其在每一个tagRNA的3’末端处包含结构化RNA)的精确编辑百分比结果。所述结构化RNA的核酸序列提供在表16中。
图36显示了使用以各种融合方向融合至本发明的构建体的染色质调节肽的精确编辑百分比结果。所测试的染色质调节肽包括HN1、HB1、H1G和CHD1。
图37显示了使用MS2/MCP系统的关于融合的精确编辑百分比结果。瞬时表达具有RT(5M)的LbCas12a H759A，没有MCP(在反式对照中)或具有MCP-RT(5M)(融合构建体)。测试了两种tagRNA，即tagRNA5和tagRNA6。所测试的不同tagRNA版本包括在其3’末端处用MS2序列进行修饰的tagRNA。
序列简述
SEQ ID NO:1-20和148-150为实例性的Cas12a氨基酸序列。
SEQ ID NO:21和SEQ ID NO:22为示例性的编码启动子和内含子的调控序列。
SEQ ID NO:23-25提供了实例性的肽标签和亲和多肽。
SEQ ID NO:26-36提供了实例性的RNA募集基序和相应的亲和多肽。
SEQ ID NO:37-52提供了实例性的单链RNA结合结构域(RBD)。
SEQ ID NO:53、97和172分别提供了实例性的逆转录酶多肽序列：莫洛尼鼠类白血病病毒(M-MuLV)5(M)，侧翼为NLS的5(M)，和M-MuLV。
SEQ ID NO:54-56提供了关于V型CRISPR-Cas12a核酸酶的原间隔子邻近基序位置的实例。
SEQ ID NO:57和SEQ ID NO:58提供了实例性的本发明的构建体。
SEQ ID NO:59和SEQ ID NO:60提供了实例性的CRISPR RNA和实例性的原间隔子。
SEQ ID NO:61和SEQ ID NO:62提供了实例性的内含子。
SEQ ID NO:63-86和SEQ ID NO:154-157提供了实例性的REDRAW编辑器构建体。
SEQ ID NO:87提供了具有11碱基对(bp)引物结合序列和96bp逆转录酶模板的tagRNA的实例。
SEQ ID NO:88-91提供了实例性的质粒的序列。
SEQ ID NO:92-94分别提供了与在图9-11中所显示的编辑相关联的tagRNA的序列。
SEQ ID NO:95、SEQ ID NO:158、SEQ ID NO:191和SEQ ID NO:203提供了实例性的假结序列。
SEQ ID NO:96提供了实例性的具有H759A的突变和在两侧均侧翼有NLS的LbCas12a。
SEQ ID NO:98-101提供了实例性的5'-3'核酸外切酶多肽。
SEQ ID NO:102和SEQ ID NO:103提供了实例性的DMNT1靶位点和靶间隔子。
SEQ ID NO:104和SEQ ID NO:105提供了实例性的FANCF1靶位点和靶间隔子。
SEQ ID NO:106和SEQ ID NO:107提供了实例性的Cas9多肽。
SEQ ID NO:108-122提供了实例性的Cas9多核苷酸。
SEQ ID NO:123-128提供了实例性的单链DNA结合蛋白。
SEQ ID NO:129-135提供了实例性的5'-3'核酸外切酶。
SEQ ID NO:136、137、140-146、159-161和171为实例性的间隔子。
SEQ ID NO:138、139和164-169提供了实例性的逆转录酶模板。
SEQ ID NO:140提供了实例性的Gam蛋白。
SEQ ID NO:151提供了实例性的Cas12b多肽。
SEQ ID NO:152和SEQ ID NO:153分别提供了实例性的单链RNA结合蛋白，防卫素和ORF5。
SEQ ID NO:162和SEQ ID NO:163提供了实例性的引物结合位点(PBS)序列。
SEQ ID NO:170提供了实例性的LbCas12a crRNA支架。
SEQ ID NO:173-186提供了实例性的tagRNA(分别为tagRNA 1、tagRNA 2、tagRNA3、tagRNA 4、tagRNA 5、tagRNA 6、tagRNA 7、tagRNA 8、tagRNA 9、tagRNA 10、tagRNA 11、tagRNA 12、tagRNA 13和tagRNA 14)。
SEQ ID NO:187和SEQ ID NO:188为在图27中所显示的逆转录酶模板。
SEQ ID NO:95、189-198和203为实例性的RNA结构。
SEQ ID NO:199-202为实例性的染色质调节肽。
SEQ ID NO:204-215为在图8、9、10和11中所找到的序列。
发明详述
现在，将会在下文中通过参考附图和实施例(在其中显示了本发明的实施方案)来描述本发明。该描述不是意在是本发明可以实施而所采取的所有不同方式或者可以添加至本发明的所有特征的详细编目。例如，关于一个实施方案所举例说明的特征可以合并入其他实施方案中，并且关于特定实施方案所举例说明的特征可以从那个实施方案中删除。因此，本发明考虑，在本发明的一些实施方案中，可以排除或省略在本文中所阐述的任何特征或特征组合。另外，鉴于本公开内容，在本文中所暗示的对各种实施方案的众多变化和添加对于本领域技术人员来说将会是明显的，其并不背离本发明。因此，下面的描述意在举例说明本发明的一些特别的实施方案，而不是穷尽地详细说明其所有排列、组合和变化。
除非另外定义，在本文中所使用的所有技术和科学术语具有与本发明所属领域的普通技术人员通常所理解的相同的含义。在本文中的发明描述中所使用的术语仅是为了描述特定实施方案而并不意在限制本发明。
在本文中所引用的所有出版物、专利申请、专利和其他参考文献通过提及而以其整体合并，以用于与其中呈现了该参考文献的句子和/或段落有关的教导。
除非上下文另有说明，特别打算的是，在本文中所描述的本发明的各种特征可以以任何组合进行使用。此外，本发明还考虑，在本发明的一些实施方案中，可以排除或省略在本文中所阐述的任何特征或特征组合。为了举例说明，如果说明书陈述到组合物包含组分A、B和C，那么特别打算的是，A、B或C中的任一个或者其组合可以单独地或以任何组合被省略和放弃。
如在本发明的描述和所附的权利要求书中所使用的，单数形式“a”、“an”和“the”意在也包括复数形式，除非上下文另有明确说明。
此外，如在本文中所使用的，“和/或”是指并且涵盖相联系的所列条目中的一个或多个的任何和所有可能的组合，以及组合的缺乏，当以备选项进行解释(“或”)时。
如在本文中所使用的，当涉及可测量的值例如量或浓度等时，术语“大约”意味着涵盖所指定的值的±10％、±5％、±1％、±0.5％或甚至±0.1％的变化以及所指定的值。例如，“大约X”(其中X为可测量的值)意味着包括X，以及X的±10％、±5％、±1％、±0.5％或甚至±0.1％的变化。在本文中对于可测量的值所提供的范围可以包括任何其他范围和/或其中的独个的值。
如在本文中所使用的，短语例如“在X和Y之间”和“在大约X和Y之间”应当被解释为包括X和Y。如在本文中所使用的，短语例如“在大约X和Y之间”意指“在大约X和大约Y之间”，并且短语例如“从大约X至Y”意指“从大约X至大约Y”。
在本文中值的范围的陈述仅意在充当独个地提及落在所述范围内的每个分开的值的速记方法，除非在本文中另有说明，并且每个分开的值被合并入说明书中，就如同它被独个地在本文中进行陈述一样。例如，如果公开了范围10至15，那么也公开了11、12、13和14。
如在本文中所使用的，术语“包含”、“含有”和“包括”明确说明了所陈述的特征、整数、步骤、操作、要素和/或组分的存在，但不排除一个或多个其他特征、整数、步骤、操作、要素、组分和/或其群组的存在或添加。
如在本文中所使用的，过渡性短语“基本上由......组成”意味着，权利要求的范围将会被解释为涵盖在该权利要求中所记述的指定的材料或步骤，以及本质上不影响所要求保护的本发明的基本和新颖特征的那些。因此，当在本发明的权利要求中使用时，术语“基本上由......组成”并不意在被解释为等价于“包含”。
如在本文中所使用的，术语“增加”、“增强”和“改善”(及其语法变化形式)描述了相比于对照而言至少大约25％、50％、75％、100％、150％、200％、300％、400％、500％或更多的提升。
如在本文中所使用的，术语“降低”、“减小”和“减少”(及其语法变化形式)描述了例如，相比于对照而言至少大约5％、10％、15％、20％、25％、35％、50％、75％、80％、85％、90％、95％、97％、98％、99％或100％的减少。在特别的实施方案中，所述降低可以导致没有或基本上没有(即，不显著的量，例如小于大约10％或甚至5％)可检测的活性或量。
“异源(的)”或“重组(的)”核苷酸序列为不与其中引入了它的宿主细胞天然地相关联的核苷酸序列，包括天然出现的核苷酸序列的非天然出现的多个拷贝。
“天然(的)”或“野生型”核酸、核苷酸序列、多肽或氨基酸序列是指天然出现的或内源的核酸、核苷酸序列、多肽或氨基酸序列。因此，例如，“野生型mRNA”为在生物中天然出现的或对参考生物来说内源的mRNA。“同源(的)”核酸序列为与其中引入了它的宿主细胞天然地相关联的核苷酸序列。
如在本文中所使用的，术语“核酸”、“核酸分子”、“核苷酸序列”和“多核苷酸”是指线性或支化的、单链或双链的RNA或DNA，或者其杂合体。该术语还涵盖RNA/DNA杂合体。当以合成方式来产生dsRNA时，较不常见的碱基(例如，肌苷、5-甲基胞嘧啶、6-甲基腺嘌呤、次黄嘌呤和其他)也可以用于反义dsRNA，和核酶配对。例如，包含尿苷和胞苷的C-5丙炔类似物的多核苷酸已显示出以高亲和力结合RNA并且是基因表达的强有力的反义抑制剂。也可以进行其他修饰，例如对于RNA的磷酸二酯主链或者在核糖基团中的2'-羟基的修饰。
如在本文中所使用的，术语“核苷酸序列”是指核苷酸的杂聚物，或者从核酸分子的5'至3'末端的这些核苷酸的顺序，并且包括DNA或RNA分子，包括cDNA、DNA片段或部分、基因组DNA、合成的(例如，以化学方式合成的)DNA、质粒DNA、mRNA和反义RNA，其中的任一种可以是单链或双链的。术语“核苷酸序列”、“核酸”、“核酸分子”、“核酸构建体”、“寡核苷酸”和“多核苷酸”在本文中也可以互换使用，以指核苷酸的杂聚物。在本文中所提供的核酸分子和/或核苷酸序列在本文中以5'至3'方向从左至右呈现，并且使用用于表示核苷酸字符的标准代码来表示，如在美国序列规则37CFR§§1.821-1.825和世界知识产权组织(WIPO)标准ST.25中所阐述的。如在本文中所使用的，“5'区”可以意指最接近多核苷酸的5'末端的多核苷酸的区域。因此，例如，在多核苷酸的5'区中的元件可以位于从位于多核苷酸的5'末端处的第一个核苷酸至位于整个多核苷酸的中途的核苷酸的任何地方。如在本文中所使用的，“3'区”可以意指最接近多核苷酸的3'末端的多核苷酸的区域。因此，例如，在多核苷酸的3'区中的元件可以位于从位于多核苷酸的3'末端处的第一个核苷酸至位于整个多核苷酸的中途的核苷酸的任何地方。
如在本文中所使用的，术语“基因”是指能够用于产生mRNA、反义RNA、miRNA、抗-微小RNA反义寡脱氧核糖核苷酸(AMO)等的核酸分子。基因可以或可以不能够用于产生功能性蛋白质或基因产物。基因可以包括编码区和非编码区(例如，内含子、调控元件、启动子、增强子、终止序列和/或5'和3'非翻译区)两者。基因可以是“分离的”，其意味着这样的核酸，其实质上或基本上没有通常与处于其天然状态的该核酸相关联地发现的组分。此类组分包括其他细胞材料、来自重组生产的培养基和/或在以化学方式合成该核酸中所使用的各种化学品。
术语“突变”是指点突变(例如、错义，或者无义，或者导致移码的单个碱基对的插入或缺失)、插入、缺失和/或截短。当突变为在氨基酸序列内的一个残基被置换为另一个残基或者在序列内的一个或多个残基的缺失或插入时，所述突变典型地通过下述方式来进行描述，即鉴定原始残基，随后为在该序列内该残基的位置和新置换的残基的身份。
如在本文中所使用的，术语“互补(的)”或“互补性”是指通过碱基配对在许可的盐和温度条件下多核苷酸的天然结合。例如，序列“A-G-T”(5'至3')与互补序列“T-C-A”(3'至5')相结合。两个单链分子之间的互补性可以是“部分的”，其中仅一些核苷酸结合，或者它可以是完全的，当在单链分子之间存在完全互补性时。核酸链之间的互补性程度对于核酸链之间的杂交的效率和强度具有显著影响。
如在本文中所使用的，“互补物”可以意指与比较者核苷酸序列的100％互补性，或者它可以意指小于100％的互补性(例如，大约70％、71％、72％、73％、74％、75％、76％、77％、78％、79％、80％、81％、82％、83％、84％、85％、86％、87％、88％、89％、90％、91％、92％、93％、94％、95％、96％、97％、98％、99％等的互补性)。
本发明的核苷酸序列的“部分”或“片段”将会被理解为意指这样的核苷酸序列，其具有相对于参考核酸或核苷酸序列而言减少(例如，减少1、2、3、4、5、6、7、8、9、10、11、12、13、14、15、16、17、18、19、20或更多个核苷酸)的长度，并且包含下述核苷酸序列、基本上由下述核苷酸序列组成和/或由下述核苷酸序列组成：与参考核酸或核苷酸序列同一或几乎同一(例如，70％、71％、72％、73％、74％、75％、76％、77％、78％、79％、80％、81％、82％、83％、84％、85％、86％、87％、88％、89％、90％、91％、92％、93％、94％、95％、96％、97％、98％、99％同一)的邻接核苷酸的核苷酸序列。当合适时，根据本发明的此类核酸片段或部分可以被包括在它是其组成成分的更大的多核苷酸中。作为例子，本发明的指导核酸的重复序列可以包含野生型V型CRISPR-Cas重复序列(例如，野生型CRISPR-Cas重复序列，例如来自Cas12a(Cpf1)、Cas12b(C2c1)、Cas12c(C2c3)、Cas12d(CasY)、Cas12e(CasX)、Cas12g、Cas12h、Cas12i、C2c4、C2c5、C2c8、C2c9、C2c10、Cas14a、Cas14b和/或Cas14c的CRISPR Cas系统的重复序列，等等)的一部分。在一些实施方案中，本发明的指导核酸的重复序列可以包含野生型CRISPR-Cas9重复序列的一部分。
具有同源性的不同核酸或蛋白质在本文中被称为“同源物”。术语“同源物”包括来自相同和其他物种的同源序列，和来自相同和其他物种的直向同源序列。“同源性”是指在位置同一性百分比方面在两个或更多个核酸和/或氨基酸序列之间的相似性水平(即序列相似性或同一性)。同源性也指在不同的核酸或蛋白质之中相似的功能特性的概念。因此，本发明的组合物和方法进一步包含本发明的核苷酸序列和多肽序列的同源物。如在本文中所使用的，“直向同源(的)”是指在物种形成期间产生自共同祖先基因的在不同物种中的同源核苷酸序列和/或氨基酸序列。本发明的核苷酸序列的同源物与本发明的所述核苷酸序列具有实质上的序列同一性(例如，至少大约70％、71％、72％、73％、74％、75％、76％、77％、78％、79％、80％、81％、82％、83％、84％、85％、86％、87％、88％、89％、90％、91％、92％、93％、94％、95％、96％、97％、98％、99％、99.5％或100％)。
如在本文中所使用的，“序列同一性”是指两个经最佳地比对的多核苷酸或多肽序列在整个组分(例如，核苷酸或氨基酸)比对窗中是不变的程度。“同一性”可以通过已知的方法容易地来进行计算，所述已知的方法包括但不限于在下列文献中所描述的那些：Computational Molecular Biology(Lesk,A.M.,编者)Oxford University Press,NewYork(1988)；Biocomputing:Informatics and Genome Projects(Smith,D.W.,编者)Academic Press,New York(1993)；Computer Analysis of Sequence Data,Part I(Griffin,A.M.和Griffin,H.G.,编者)Humana Press,New Jersey(1994)；SequenceAnalysis in Molecular Biology(von Heinje,G.,编者)Academic Press(1987)；和Sequence Analysis Primer(Gribskov,M.和Devereux,J.,编者)Stockton Press,NewYork(1991)。
如在本文中所使用的，术语“序列同一性百分比”或“同一性百分比”是指当以最佳方式比对两个序列时，相比于测试(“受试”)多核苷酸分子(或其互补链)而言在参考(“查询”)多核苷酸分子(或其互补链)的线性多核苷酸序列中相同的核苷酸的百分比。在一些实施方案中，“同一性百分比”可以是指相比于参考多肽而言在氨基酸序列中相同的氨基酸的百分比。
如在本文中所使用的，在两个核酸分子、核苷酸序列或蛋白质序列的情景下，短语“实质上同一的”或“实质上的同一性”是指当就最大相应性进行比较和比对时，具有至少大约70％、71％、72％、73％、74％、75％、76％、77％、78％、79％、80％、81％、82％、83％、84％、85％、86％、87％、88％、89％、90％、91％、92％、93％、94％、95％、96％、97％、98％、99％、99.5％、99.6％、99.7％、99.8％、99.9％或更多核苷酸或氨基酸残基同一性的两个或更多个序列或子序列，如通过使用下列的序列比较算法之一或通过目视检查所测量的。在本发明的一些实施方案中，在长度为大约10个核苷酸至大约20个核苷酸、大约10个核苷酸至大约25个核苷酸、大约10个核苷酸至大约30个核苷酸、大约15个核苷酸至大约25个核苷酸、大约30个核苷酸至大约40个核苷酸、大约50个核苷酸至大约60个核苷酸、大约70个核苷酸至大约80个核苷酸、大约90个核苷酸至大约100个核苷酸或更多个核苷酸的本发明的核苷酸序列的连续核苷酸的区域，和其中任何范围，直至该序列的全长之上存在实质上的同一性。在一些实施方案中，所述核苷酸序列可以在至少大约20个核苷酸(例如，大约20、21、22、23、24、25、26、27、28、29、30、31、32、33、34、35、36、37、38、39、40个核苷酸)之上是实质上同一的。在一些实施方案中，实质上同一的核苷酸或蛋白质序列执行与它与之实质上同一的核苷酸序列(或所编码的蛋白质序列)实质上相同的功能。
对于序列比较，一个序列典型地充当参考序列，测试序列与之进行比较。当使用序列比较算法时，将测试序列和参考序列输入计算机，指派子序列坐标(如果需要)，并且指派序列算法程序参数。然后，序列比较算法基于所指派的程序参数来计算相对于参考序列而言的关于测试序列的序列同一性百分比。
用于比对比较窗的最佳序列比对是本领域技术人员众所周知的，并且可以通过诸如Smith和Waterman的局部同源性算法、Needleman和Wunsch的同源性比对算法、Pearson和Lipman的相似性搜索方法的工具，和任选地通过这些算法的计算机化实施例如GAP、BESTFIT、FASTA和TFASTA(其作为Wisconsin />(Accelrys Inc.,SanDiego,CA)的一部分而可得)来进行。关于测试序列和参考序列的所比对的区段的“同一性分数”为由两个所比对的序列所共享的相同组分的数目除以在参考序列区段(例如，整个参考序列或者参考序列的更小的所定义的部分)中的组分的总数目。将序列同一性百分比表示为同一性分数乘以100。一个或多个多核苷酸序列的比较可以是与全长多核苷酸序列或其部分，或者与更长的多核苷酸序列。对于本发明的目的，“同一性百分比”也可以通过使用BLASTX 2.0版(对于翻译的核苷酸序列)和BLASTN 2.0版(对于多核苷酸序列)来进行测定。
当两个核苷酸序列在严紧条件下相互杂交时，也可以认为这两个序列是实质上互补的。在一些代表性的实施方案中，两个被认为是实质上互补的核苷酸序列在高度严紧条件下相互杂交。
在核酸杂交实验例如Southern和Northern杂交的情景下的“严紧杂交条件”和“严紧杂交洗涤条件”是序列依赖性的，并且在不同的环境参数下是不同的。对于核酸杂交的详尽指南可见于Tijssen Laboratory Techniques in Biochemistry and MolecularBiology-Hybridization with Nucleic Acid Probes,第2章第I部分,"Overview ofprinciples of hybridization and the strategy of nucleic acid probe assays",Elsevier,New York(1993)。一般而言，高度严紧杂交和洗涤条件被选择为在规定的离子强度和pH下低于关于特定序列的热解链点(Tm)大约5℃。
Tm为这样的温度，在该温度下靶序列的50％与完美匹配的探针杂交(在规定的离子强度和pH下)。非常严紧条件被选择为等于关于特定探针的Tm。关于在Southern或Northern印迹中在滤器上具有多于100个互补残基的互补核苷酸序列的杂交的严紧杂交条件的例子在为42℃下具有1mg的肝素的50％甲酰胺，其中杂交过夜进行。高度严紧洗涤条件的例子为在72℃下0.1 5M NaCl进行大约15分钟。严紧洗涤条件的例子为在65℃下0.2xSSC洗涤15分钟(参见，Sambrook(下文)，见SSC缓冲液的描述)。经常，在高严紧性洗涤之前为低严紧性洗涤，以去除背景探针信号。关于例如多于100个核苷酸的双链体的中等严紧性洗涤的例子为在45℃下1x SSC进行15分钟。关于例如多于100个核苷酸的双链体的低严紧性洗涤的例子为在40℃下4-6x SSC进行15分钟。对于短探针(例如，大约10至50个核苷酸)，严紧条件典型地涉及在pH 7.0至8.3下低于大约1.0M Na离子，典型地大约0.01至1.0M Na离子浓度(或其他盐)的盐浓度，并且温度典型地为至少大约30℃。严紧条件也可以通过添加去稳定试剂例如甲酰胺来取得。通常，在特定的杂交测定法中相比于对于不相关的探针所观察的那种而言2x(或更高)的信噪比表明检测到特异性杂交。在严紧条件下不相互杂交的核苷酸序列仍然是实质上同一的，如果它们所编码的蛋白质是实质上同一的。这可以例如当使用由遗传密码所允许的最大密码子简并性来产生核苷酸序列的拷贝时出现。
本发明的多核苷酸和/或重组核酸构建体可以进行密码子优化，以用于进行表达。在一些实施方案中，本发明的多核苷酸、核酸构建体、表达盒和/或载体(例如，其包含/编码CRISPR-Cas效应蛋白(例如，V型CRISPR-Cas效应蛋白)、逆转录酶、翼片核酸内切酶、5'-3'核酸外切酶等)进行密码子优化，以用于在生物中(例如，在特定的物种中)，任选地在动物、植物、真菌、古细菌或细菌中进行表达。在一些实施方案中，经密码子优化的本发明的核酸构建体、多核苷酸、表达盒和/或载体与未进行密码子优化的核酸构建体、多核苷酸、表达盒和/或载体具有大约70％至大约99.9％(例如，70％、71％、72％、73％、74％、75％、76％、77％、78％、79％、80％、81％、82％、83％、84％、85％、86％、87％、88％、89％、90％、91％、92％、93％、94％、95％、96％、97％、98％、99％、99.5％或99.9％)的同一性或更多。
在本文中所描述的实施方案的任一个之中，可以将本发明的多核苷酸或核酸构建体与各种各样的启动子和/或其他调控元件可运转地相关联，以用于在植物中和/或植物的细胞中进行表达。因此，在一些实施方案中，本发明的多核苷酸或核酸构建体可以进一步包含一个或多个与一个或多个核苷酸序列可操作地连接的启动子、内含子、增强子和/或终止子。在一些实施方案中，启动子可以与内含子可操作地相关联(例如，Ubi1启动子和内含子)。在一些实施方案中，与内含子相关联的启动子可以称为“启动子区”(例如，Ubi1启动子和内含子)。
如在本文中关于多核苷酸所使用的，“可操作地连接”或“可操作地相关联”意味着，所示元件在功能上相互联系，并且通常也在物理上相联系。因此，如在本文中所使用的，术语“可操作地连接”或“可操作地相关联”是指在功能上相关联的在单个核酸分子上的核苷酸序列。因此，与第二核苷酸序列可操作地连接的第一核苷酸序列意指当将所述第一核苷酸序列以与所述第二核苷酸序列具有功能关系的方式进行放置时的情况。例如，一个启动子与一个核苷酸序列可操作地相关联，如果所述启动子影响所述核苷酸序列的转录或表达。本领域技术人员将会意识到，控制序列(例如，启动子)不需要与它所与之可操作地相关联的核苷酸序列相邻接，只要所述控制序列发挥功能以指引其表达。因此，例如，间插性的非翻译(然而经转录)的核酸序列可以存在于启动子和核苷酸序列之间，并且所述启动子仍然可以被认为与所述核苷酸序列“可操作地连接”。
如在本文中所使用的，关于多肽，术语“连接”是指一个多肽附接至另一个多肽。多肽可以直接地(例如，经由肽键)或通过连接体与另一个多肽(在N-末端或C-末端处)相连接。
术语“连接体”是本领域公认的，并且是指连接两个分子或部分(例如，融合蛋白的两个结构域，例如DNA结合多肽或结构域和肽标签和/或逆转录酶和与所述肽标签相结合的亲和多肽；或者DNA核酸内切酶多肽或结构域和肽标签和/或逆转录酶和与所述肽标签相结合的亲和多肽)的化学基团或分子。连接体可以由单个连接分子构成，或者可以包含多于一个连接分子。在一些实施方案中，所述连接体可以为有机分子、基团、聚合物或化学部分(例如二价有机部分)。在一些实施方案中，所述连接体可以为氨基酸，或者它可以是肽。在一些实施方案中，所述连接体为肽。
在一些实施方案中，对本发明来说有用的肽连接体的长度可以为大约2至大约100或更多个氨基酸，例如长度为大约2、3、4、5、6、7、8、9、10、11、12、13、14、15、16、17、18、19、20、21、22、23、24、25、26、27、28、29、30、31、32、33、34、35、36、37、38、39、40、41、42、43、44、45、46、47、48、49、50、51、52、53、54、55、56、57、58、59、60、61、62、63、64、65、66、67、68、69、70、71、72、73、74、75、76、77、78、79、80、81、82、83、84、85、86、87、88、89、90、91、92、93、94、95、96、97、98、99、100或更多个氨基酸，例如长度为大约2至大约40，大约2至大约50，大约2至大约60，大约4至大约40，大约4至大约50，大约4至大约60，大约5至大约40，大约5至大约50，大约5至大约60，大约9至大约40，大约9至大约50，大约9至大约60，大约10至大约40，大约10至大约50，大约10至大约60，或者大约2、3、4、5、6、7、8、9、10、11、12、13、14、15、16、17、18、19、20、21、22、23、24、25个氨基酸至大约26、27、28、29、30、31、32、33、34、35、36、37、38、39、40、41、42、43、44、45、46、47、48、49、50、51、52、53、54、55、56、57、58、59、60、61、62、63、64、65、66、67、68、69、70、71、72、73、74、75、76、77、78、79、80、81、82、83、84、85、86、87、88、89、90、91、92、93、94、95、96、97、98、99、100或更多个氨基酸(例如，长度为大约105、110、115、120、130、140、150或更多个氨基酸)。在一些实施方案中，肽连接体可以为GS连接体。
如在本文中所使用的，关于多核苷酸，术语“连接”或“融合”是指一个多核苷酸附接至另一个多核苷酸。在一些实施方案中，两个或更多个多核苷酸分子可以通过连接体相连接，所述连接体可以为有机分子、基团、聚合物或化学部分(例如二价有机部分)。多核苷酸可以经由共价或非共价连接或结合(包括例如沃森-克里克碱基配对)或者通过一个或多个连接核苷酸而连接或融合至另一个多核苷酸(在5'末端或3'末端)。在一些实施方案中，可以将某一结构的多核苷酸基序插入到另一个多核苷酸序列内(例如，在指导RNA中的发夹结构的延伸)。在一些实施方案中，所述连接核苷酸可以为天然出现的核苷酸。在一些实施方案中，所述连接核苷酸可以为非天然出现的核苷酸。
“启动子”为控制或调控与该启动子可操作地相关联的核苷酸序列(例如，编码序列)的转录的核苷酸序列。由启动子控制或调控的编码序列可以编码多肽和/或功能性RNA。典型地，“启动子”是指包含用于RNA聚合酶II的结合位点并且指引转录起始的核苷酸序列。一般而言，启动子被发现处于相对于相应的编码序列的编码区的起始点而言的5'或上游。启动子可以包含作为基因表达的调节物起作用的其他元件，例如启动子区。这些包括TATA盒共有序列，和经常地，CAAT盒共有序列(Breathnach和Chambon,(1981)Annu.Rev.Biochem.50:349)。在植物中，CAAT盒可以被替代为AGGA盒(Messing等人,(1983)Genetic Engineering of Plants,T.Kosuge,C.Meredith和A.Hollaender(编者),PlenumPress,第211-227页)。在一些实施方案中，启动子区可以包含至少一个内含子(参见例如，SEQ ID NO:21、SEQ ID NO:22)。
对本发明来说有用的启动子可以包括例如组成性的、诱导型的、在时间上受调控的、在发育上受调控的、以化学方式受调控的、组织优选的和/或组织特异性的启动子，其用于在制备重组核酸分子例如“合成的核酸构建体”或“蛋白质-RNA复合物”中使用。启动子的这些各种类型是本领域中已知的。
启动子的选择可以取决于对于表达的时间和空间要求而变化，并且也可以基于待转化的宿主细胞而变化。用于许多不同生物的启动子是本领域中众所周知的。基于本领域中存在的详尽知识，可以为特定的目的宿主生物选择合适的启动子。因此，例如，关于在模式生物中高度组成性地表达的基因的上游的启动子知道很多，并且此类知识可以容易地获取并且在其他系统中施行(在合适时)。
在一些实施方案中，在植物中有功能的启动子可以与本发明的构建体一起进行使用。对于驱动在植物中的表达来说有用的启动子的非限制性例子包括RubisCo小亚基基因1的启动子(PrbcS1)、肌动蛋白基因的启动子(Pactin)、硝酸还原酶基因的启动子(Pnr)和重复碳酸酐酶基因1的启动子(Pdca1)(参见，Walker等人,Plant Cell Rep.23:727-735(2005)；Li等人,Gene 403:132-142(2007)；Li等人,Mol Biol.Rep.37:1143-1154(2010))。PrbcS1和Pactin是组成性启动子，而Pnr和Pdca1是诱导型启动子。Pnr被硝酸盐诱导和被铵阻抑(Li等人,Gene 403:132-142(2007))，而Pdca1被盐诱导(Li等人,Mol Biol.Rep.37:1143-1154(2010))。在一些实施方案中，对本发明来说有用的启动子为RNA聚合酶II(PolII)启动子。在一些实施方案中，来自玉蜀黍(Zea mays)的U6启动子或7SL启动子可以是对本发明的构建体有用的。在一些实施方案中，来自玉蜀黍的U6c启动子和/或7SL启动子可以是对于驱动指导核酸的表达来说有用的。在一些实施方案中，来自大豆(Glycine max)的U6c启动子、U6i启动子和/或7SL启动子可以是对本发明的构建体有用的。在一些实施方案中，来自大豆的U6c启动子、U6i启动子和/或7SL启动子可以是对于驱动指导核酸的表达来说有用的。
对于植物来说有用的组成性启动子的例子包括但不限于：夜香树病毒启动子(cmp)(美国专利号7,166,770)、水稻肌动蛋白1启动子(Wang等人,(1992)Mol.Cell.Biol.12:3399-3406；以及美国专利号5,641,876)、CaMV 35S启动子(Odell等人,(1985)Nature313:810-812)、CaMV 19S启动子(Lawton等人,(1987)Plant Mol.Biol.9:315-324)、nos启动子(Ebert等人,(1987)Proc.Natl.Acad.Sci USA 84:5745-5749)、Adh启动子(Walker等人,(1987)Proc.Natl.Acad.Sci.USA 84:6624-6629)、蔗糖合酶启动子(Yang&Russell(1990)Proc.Natl.Acad.Sci.USA 87:4144-4148)和泛素启动子。源自泛素的组成性启动子在许多细胞类型中积累。已从几个植物物种中克隆出了泛素启动子以用于在转基因植物中使用：例如，向日葵(Binet等人,1991.Plant Science 79:87-94)、玉蜀黍(Christensen等人,1989.Plant Molec.Biol.12:619-632)和拟南芥(Norris等人,1993.Plant Molec.Biol.21:895-906)。已在转基因单子叶植物系统中开发了玉蜀黍泛素启动子(UbiP)，并且在专利出版物EP 0 342 926中公开了其序列和为单子叶植物转化所构建的载体。泛素启动子适合于在转基因植物(尤其是单子叶植物)中表达本发明的核苷酸序列。进一步地，可以容易地修饰由McElroy等人(Mol.Gen.Genet.231:150-160(1991))所描述的启动子表达盒以用于表达本发明的核苷酸序列并且特别适合于在单子叶植物宿主中使用。
在一些实施方案中，组织特异性/组织偏爱型启动子可以用于在植物细胞中表达异源多核苷酸。组织特异性或偏爱的表达模式包括但不限于：绿色组织特异性或偏爱的、根特异性或偏爱的、茎特异性或偏爱的、花特异性或偏爱的或者花粉特异性或偏爱的。适合于在绿色组织中进行表达的启动子包括许多调控在光合作用中所牵涉的基因的启动子，并且这些中的许多已从单子叶植物和双子叶植物中克隆出。在一个实施方案中，对本发明来说有用的启动子为来自磷酸烯醇羧化酶基因的玉蜀黍PEPC启动子(Hudspeth&Grula,PlantMolec.Biol.12:579-589(1989))。组织特异性启动子的非限制性例子包括与下列相关联的那些：编码种子贮藏蛋白(例如β-伴大豆球蛋白、cruciferin、油菜籽蛋白和菜豆蛋白)、玉米醇溶蛋白或油体蛋白(例如油质蛋白)或者在脂肪酸生物合成中所牵涉的蛋白质(包括酰基载体蛋白、硬脂酰基-ACP去饱和酶和脂肪酸去饱和酶(fad 2-1))的基因，和其他在胚胎发育期间表达的核酸(例如，Bce4，参见例如，Kridl等人,(1991)Seed Sci.Res.1:209-219；以及EP专利号255378)。对于在植物(特别是玉蜀黍)中表达本发明的核苷酸序列来说有用的组织特异性或组织优先型启动子包括但不限于在根、木髓、叶或花粉中指引表达的那些。此类启动子例如公开在WO 93/07278(其通过提及而以其整体合并入本文)中。对本发明来说有用的组织特异性或组织偏爱型启动子的其他非限制性例子为在美国专利6,040,504中所公开的棉花核酮糖二磷酸羧化酶-加氧酶(rubisco)启动子；在美国专利5,604,121中所公开的水稻蔗糖合酶启动子；由de Framond(FEBS 290:103-106(1991)；属于Ciba-Geigy的EP 0 452 269)所描述的根特异性启动子；在美国专利5,625,136(属于Ciba-Geigy)中所描述的并且驱动玉蜀黍trpA基因的表达的茎特异性启动子；在WO 01/73087中所公开的夜香树黄叶卷曲病毒启动子；和花粉特异性或偏爱型启动子，包括但不限于来自水稻的ProOsLPS10和ProOsLPS11(Nguyen等人,Plant Biotechnol.Reports 9(5):297-306(2015))、来自玉蜀黍的ZmSTK2_USP(Wang等人,Genome 60(6):485-495(2017))、来自番茄的LAT52和LAT59(Twell等人,Development 109(3):705-713(1990))、Zm13(美国专利号10,421,972)、来自拟南芥的PLA2-δ启动子(美国专利号7,141,424)和/或来自玉蜀黍的ZmC5启动子(国际PCT公开号WO1999/042587)。
组织特异性/组织偏爱型启动子的另外例子包括但不限于：根毛特异性顺式元件(RHE)(Kim等人,The Plant Cell 18:2958-2970(2006))、根特异性启动子RCc3(Jeong等人,Plant Physiol.153:185-197(2010))和RB7(美国专利号5459252)、凝集素启动子(Lindstrom等人,(1990)Der.Genet.11:160-167；和Vodkin(1983)Prog.Clin.Biol.Res.138:87-98)、玉米醇脱氢酶1启动子(Dennis等人,(1984)NucleicAcids Res.12:3983-4000)、S-腺苷-L-甲硫氨酸合成酶(SAMS)(Vander Mijnsbrugge等人,(1996)Plant and Cell Physiology,37(8):1108-1115)、玉米集光复合物启动子(Bansal等人,(1992)Proc.Natl.Acad.Sci.USA 89:3654-3658)、玉米热休克蛋白启动子(O'Dell等人,(1985)EMBO J.5:451-458；和Rochester等人,(1986)EMBO J.5:451-458)、豌豆小亚基RuBP羧化酶启动子(Cashmore,“Nuclear genes encoding the small subunit ofribulose-l,5-bisphosphate carboxylase”,第29-39页,Genetic Engineering ofPlants(Hollaender编者),Plenum Press 1983；和Poulsen等人,(1986)Mol.Gen.Genet.205:193-200)、Ti质粒甘露碱合酶启动子(Langridge等人,(1989)Proc.Natl.Acad.Sci.USA 86:3219-3223)、Ti质粒胭脂碱合酶启动子(Langridge等人,(1989),见上)、矮牵牛查耳酮异构酶启动子(van Tunen等人,(1988)EMBO J.7:1257-1263)、豆类富甘氨酸蛋白1启动子(Keller等人,(1989)Genes Dev.3:1639-1646)、截短的CaMV 35S启动子(O'Dell等人,(1985)Nature 313:810-812)、马铃薯patatin启动子(Wenzler等人,(1989)Plant Mol.Biol.13:347-354)、根细胞启动子(Yamamoto等人,(1990)nucleic acids Res.18:7449)、玉蜀黍醇溶蛋白启动子(Kriz等人,(1987)Mol.Gen.Genet.207:90-98；Langridge等人,(1983)Cell 34:1015-1022；Reina等人,(1990)Nucleic Acids Res.18:6425；Reina等人,(1990)Nucleic Acids Res.18:7449；和Wandelt等人,(1989)Nucleic Acids Res.17:2354)、球蛋白-1启动子(Belanger等人,(1991)Genetics 129:863-872)、α-微管蛋白cab启动子(Sullivan等人,(1989)Mol.Gen.Genet.215:431-440)、PEPC酶启动子(Hudspeth&Grula(1989)PlantMol.Biol.12:579-589)、R基因复合物相关联的启动子(Chandler等人,(1989)PlantCell1:1175-1183)和查耳酮合酶启动子(Franken等人,(1991)EMBO J.10:2605-2612)。
对于种子特异性表达来说有用的是豌豆球蛋白启动子(Czako等人,(1992)Mol.Gen.Genet.235:33-40；以及在美国专利号5,625,136中所公开的种子特异性启动子。对于在成熟叶中的表达来说有用的启动子为在衰老开始时进行转换的那些，例如来自拟南芥属(Arabidopsis)的SAG启动子(Gan等人,(1995)Science270:1986-1988)。
另外，可以使用在叶绿体中有功能的启动子。此类启动子的非限制性例子包括在美国专利号7,579,516中所公开的噬菌体T3基因9 5'UTR和其他启动子。对本发明来说有用的其他启动子包括但不限于：S-E9小亚基RuBP羧化酶启动子和Kunitz胰蛋白酶抑制剂基因启动子(Kti3)。
对本发明来说有用的另外的调控元件包括但不限于内含子、增强子、终止序列和/或5'和3'非翻译区。
对本发明来说有用的内含子可以为在植物中鉴定和从植物中分离并然后插入到待在植物转化中使用的表达盒中的内含子。如本领域技术人员将会理解的，内含子可以包含对于自我切除所需要的序列，并且以符合读框的方式掺入到核酸构建体/表达盒中。内含子可以用作间隔子以分开在一个核酸构建体中的多个蛋白质编码序列，或者可以在一个蛋白质编码序列内部使用内含子以例如使mRNA稳定化。如果在蛋白质编码序列之内使用它们，那么“以符合读框的方式”插入它们，包括切除位点。也可以将内含子与启动子相联合以改善或修饰表达。作为例子，对本发明来说有用的启动子/内含子组合包括但不限于玉蜀黍Ubi1启动子和内含子的组合。
对本发明来说有用的内含子的非限制性例子包括来自下列基因的内含子：ADHI基因(例如，Adh1-S内含子1、2和6)、泛素基因(Ubi1)、RuBisCO小亚基(rbcS)基因、RuBisCO大亚基(rbcL)基因、肌动蛋白基因(例如，肌动蛋白-1内含子)、丙酮酸脱氢酶激酶基因(pdk)、硝酸还原酶基因(nr)、重复碳酸酐酶基因1(Tdca1)、psbA基因、atpA基因或其任何组合。实例性的内含子序列可以包括但不限于SEQ ID NO:61和SEQ ID NO:62。
在一些实施方案中，本发明的多核苷酸和/或核酸构建体可以为“表达盒”或者可以被包含在表达盒之内。如在本文中使用的，“表达盒”意指这样的重组核酸分子，其包含例如本发明的核酸构建体(例如，CRISPR-Cas效应蛋白，逆转录酶多肽或结构域，翼片核酸内切酶多肽或结构域(例如，FEN)，和/或5'-3'核酸外切酶)，其中所述核酸构建体与一个或多个控制序列(例如，启动子、终止子等)可操作地相关联。因此，本发明的一些实施方案提供了被设计成用于表达例如本发明的核酸构建体(例如，编码CRISPR-Cas效应蛋白或结构域、逆转录酶多肽或结构域、翼片核酸内切酶多肽或结构域和/或5'-3'核酸外切酶多肽或结构域的本发明的核酸构建体)的表达盒。当本发明的表达盒包含多于一个多核苷酸时，所述多核苷酸可以可操作地连接至驱动所有所述多核苷酸表达的单个启动子或者所述多核苷酸可以可操作地连接至一个或多个分开的启动子(例如，三个多核苷酸可以由一个、两个或三个(以任何组合)启动子来驱动)。当使用两个或更多个分开的启动子时，所述启动子可以是相同的启动子，或者它们可以是不同的启动子。因此，可以将在表达盒中所包含的编码CRISPR-Cas效应蛋白或结构域的多核苷酸、编码逆转录酶多肽或结构域的多核苷酸、编码翼片核酸内切酶多肽或结构域的多核苷酸和/或编码5'-3'核酸外切酶多肽或结构域的多核苷酸各自可操作地连接至分开的启动子，或者它们可以可操作地连接至两个或更多个启动子(以任何组合)。
包含本发明的核苷酸构建体的表达盒可以是嵌合的，这意味着其组分中的至少一个相关于其其他成分中的至少一个而言是异源的(例如，与待在宿主生物中表达的目的多核苷酸可操作地连接的来自宿主生物的启动子，其中所述目的多核苷酸来自不同于该宿主的生物或者通常不与那个启动子相关联地被发现)。表达盒也可以为天然出现但是已经以对于异源表达来说有用的重组形式获得的表达盒。
表达盒任选地可以包括在所选择的宿主细胞中有功能的转录和/或翻译终止区(即，终止区)和/或增强子区。各种各样的转录终止子和/或增强子是本领域中已知的并且是可得的以用于在表达盒中使用。转录终止子负责转录的终止和正确的mRNA多腺苷酸化。终止区和/或增强子区可以对于转录起始区来说是天然的，可以例如对于编码CRISPR-Cas效应蛋白的基因、编码逆转录酶的基因、编码翼片核酸内切酶的基因和/或编码5'-3'核酸外切酶的基因来说是天然的，可以对于宿主细胞来说是天然的，或者可以对于另一个来源(例如，对于启动子，对于编码CRISPR-Cas效应蛋白的基因、编码逆转录酶的基因、编码翼片核酸内切酶的基因和/或编码5'-3'核酸外切酶的基因，对于宿主细胞，或者对于其任何组合来说是外来的或异源的)来说是天然的。
本发明的表达盒也可以包括编码选择标记(其可以用于选择经转化的宿主细胞)的多核苷酸。如在本文中所使用的，“选择标记”意指这样的多核苷酸序列，其当被表达时，赋予表达所述标记的宿主细胞以不同的表型，并且因此允许此类经转化的细胞区别于不具有所述标记的那些细胞。此类多核苷酸序列可以编码可选择或可筛选的标记，这取决于所述标记是否赋予可以通过化学手段，例如通过使用选择试剂(例如，抗生素等)进行选择的性状，或者取决于所述标记是否仅是通过观察或测试，例如通过筛选(例如，荧光)就可以鉴定的性状。合适的选择标记的许多例子是本领域中已知的，并且可以被用于在本文中所描述的表达盒之中。
除了表达盒之外，在本文中所描述的核酸分子/构建体和多核苷酸序列也可以与载体相联系地进行使用。术语“载体”是指用于将核酸转移、递送或引入到细胞中的组合物。载体包含含有待转移、递送或引入的核苷酸序列的核酸构建体。用于在宿主生物的转化中使用的载体是本领域中众所周知的。载体的一般类别的非限制性例子包括病毒载体、质粒载体、噬菌体载体、噬菌粒载体、粘粒载体、F粘粒(fosmid)载体、噬菌体、人工染色体、小环或土壤杆菌二元载体，以双链或单链的线性或环状形式，其可以是或可以不是可自我传递的或可移动的。在一些实施方案中，病毒载体可以包括但不限于，逆转录病毒、慢病毒、腺病毒、腺伴随病毒或单纯疱疹病毒载体。在本文中所定义的载体可以通过整合到细胞基因组中或者在染色体外存在(例如，具有复制起点的自主复制质粒)来转化原核生物或真核生物宿主。另外还包括穿梭载体，其意指能够(天然地或通过设计)在两种不同的宿主生物中复制的DNA载料，所述宿主生物可以选自放线菌和相关物种、细菌和真核生物(例如，高等植物、哺乳动物、酵母或真菌细胞)。在一些实施方案中，在载体中的核酸处于合适的启动子或其他用于在宿主细胞中进行转录的调控元件的控制之下并且与之可操作地连接。所述载体可以为在多种宿主中发挥功能的双功能表达载体。在基因组DNA的情况下，这可以包含其自己的启动子和/或其他调控元件，和在cDNA的情况下，这可以处于合适的启动子和/或其他用于在宿主细胞中进行表达的调控元件的控制之下。因此，可以将本发明的核酸构建体或多核苷酸和/或包含其的表达盒包含在本文中所描述的和本领域中已知的载体之中。
如在本文中所使用的，“接触”及其语法变化形式是指将所希望的反应的组分一起放置在适合于进行该所希望的反应(例如，转化、转录控制、基因组编辑、切口产生和/或切割)的条件下。作为例子，可以使靶核酸与II型或V型CRISPR-Cas效应蛋白和逆转录酶或者编码其的核酸构建体在这样的条件下相接触，通过这些条件所述CRISPR-Cas效应蛋白和所述逆转录酶被表达并且所述CRISPR-Cas效应蛋白与所述靶核酸相结合，并且所述逆转录酶融合至所述CRISPR-Cas效应蛋白或募集至所述CRISPR-Cas效应蛋白(经由例如融合至所述CRISPR-Cas效应蛋白的肽标签和融合至所述逆转录酶的亲和标签)，并且因此所述逆转录酶位于所述靶核酸的附近，由此修饰所述靶核酸。可以使用其他用于募集逆转录酶的方法，其利用其他蛋白质-蛋白质相互作用和还有RNA-蛋白质相互作用和化学相互作用。
如在本文中所使用的，关于靶核酸，“修饰”包括靶核酸的编辑(例如，突变)、共价修饰、交换/置换核酸/核苷酸碱基、缺失、切割、切口产生和/或转录控制。在一些实施方案中，修饰可以包括任何大小的插入/缺失，和/或任何类型的单碱基变化(SNP)。
在目的多核苷酸的情景下，“引入”(及其语法变化形式)意指以这样的方式向宿主生物或所述生物的细胞(例如，宿主细胞，例如植物细胞)呈递目的核苷酸序列(例如，多核苷酸、核酸构建体和/或指导核酸)，从而使得所述核苷酸序列得以进入细胞的内部。
术语“转化”或“转染”可以互换使用，并且如在本文中所使用的，是指将异源核酸引入到细胞中。细胞的转化可以是稳定的或瞬时的。因此，在一些实施方案中，可以用本发明的多核苷酸/核苷酸分子稳定地转化宿主细胞或宿主生物。在一些实施方案中，可以用本发明的核酸构建体瞬时地转化宿主细胞或宿主生物。
在多核苷酸的情景下，“瞬时转化”意味着，多核苷酸被引入到细胞中，并且不整合到细胞的基因组中。
在被引入到细胞中的多核苷酸的情景下，“稳定地引入”意指，所引入的多核苷酸被稳定地掺入到细胞的基因组中，并且因此用所述多核苷酸稳定地转化所述细胞。
如在本文中所使用的，“稳定转化”或“稳定地转化”意味着，核酸分子被引入到细胞中并且整合到细胞的基因组中。如此，所整合的核酸分子能够被其后代，更特别地被多个连续世代的后代遗传。如在本文中所使用的，“基因组”包括核基因组、线粒体基因组和质体基因组，并且因此包括核酸整合到例如叶绿体或线粒体基因组中。如在本文中所使用的，“稳定转化”也指在染色体外维持的转基因，例如作为微型染色体或质粒。
瞬时转化可以通过例如酶联免疫吸附测定法(ELISA)或Western印迹来检测，其可以检测由被引入到生物中的一个或多个转基因所编码的肽或多肽的存在。细胞的稳定转化可以通过例如所述细胞的基因组DNA的Southern印迹杂交测定法来检测，其使用与被引入到生物(例如，植物)中的转基因的核苷酸序列特异性地杂交的核酸序列。细胞的稳定转化可以通过例如所述细胞的RNA的Northern印迹杂交测定法来检测，其使用与被引入到宿主生物中的转基因的核苷酸序列特异性地杂交的核酸序列。细胞的稳定转化也可以通过例如聚合酶链式反应(PCR)或其他扩增反应(如本领域中众所周知的)来检测，其采用与转基因的靶序列杂交的特异性引物序列，从而导致转基因序列的扩增，其可以根据标准方法来检测。转化也可以通过本领域中众所周知的直接测序和/或杂交实验方案来检测。
因此，在一些实施方案中，可以瞬时地表达本发明的核苷酸序列、多核苷酸、核酸构建体和/或表达盒，和/或可以将它们稳定地掺入到宿主生物的基因组中。因此，在一些实施方案中，可以将本发明的核酸构建体(例如，一个或多个编码DNA结合多肽或结构域、核酸内切酶多肽或结构域、逆转录酶多肽或结构域、翼片核酸内切酶多肽或结构域和/或核酸修饰多肽或结构域的表达盒)与指导核酸一起瞬时地引入到细胞中，并且如此没有DNA维持在所述细胞中。
可以通过本领域技术人员已知的任何方法来将本发明的核酸构建体引入到细胞中。在本发明的一些实施方案中，细胞的转化包括核转化。在其他实施方案中，细胞的转化包括质体转化(例如，叶绿体转化)。在进一步的实施方案中，可以经由常规育种技术将本发明的重组核酸构建体引入到细胞中。
用于转化真核和原核生物两者的程序是本领域中众所周知的和常规的，并且在文献各处作了描述(参见例如，Jiang等人,2013.Nat.Biotechnol.31:233-239；Ran等人,Nature Protocols 8:2281-2308(2013))。
因此，可以以许多本领域中众所周知的方式将核苷酸序列引入到宿主生物或其细胞中。本发明的方法不依赖于特定的用于将一个或多个核苷酸序列引入到生物中的方法，只要它们得以进入所述生物的至少一个细胞的内部。当待引入多于一个核苷酸序列时，可以将它们装配作为单个核酸构建体的一部分，或者作为分开的核酸构建体，并且可以位于相同或不同的核酸构建体上。因此，可以在单个转化事件中和/或在分开的转化事件中将核苷酸序列引入到目的细胞中，或者备选地，在相关的情况下，可以将核苷酸序列掺入到植物中，例如作为育种实验方案的一部分。
碱基编辑已显示出是分别将胞嘧啶和腺嘌呤残基改变为胸腺嘧啶和鸟嘌呤的有效方式。这些工具虽然强大，但确实有一些限制，例如旁观者碱基，小的碱基编辑窗口和有限的PAM。
为了在细胞中进行精确的模板式编辑，存在几个基本步骤，其各自具有速率限制，它们在一起由于低的效率而可以严重地妨碍有效地进行编辑的能力。例如，一个步骤要求诱导细胞在靶位点处起始修复事件。这典型地通过由外源提供的序列特异性核酸酶或切口酶引起双链断裂(DSB)或切口来进行。另一个步骤要求同源模板的局部可得性以用于修复。该步骤要求恰好在DSB有能力投入模板式编辑途径时的正确时间模板在DSB的邻近。特别地，在当前的编辑技术中，该步骤被广泛地视为是限速步骤。一个进一步的步骤为序列从模板有效地掺入到断裂的或带切口的靶标中。在本发明之前，该步骤典型地由细胞的内源的DNA修复酶来提供。该步骤的效率是低的并且难以操纵。本发明通过以协调方式使对于执行上述步骤所要求的功能性共局部化而绕开了许多对于模板式编辑过程的效率的主要障碍。
图1显示了通过使用本发明的方法和构建体，从crRNA的逆转录生成DNA序列并且随后整合到切口位点中。延伸型crRNA以蓝色显示并且与第二链切口酶Cpf1(Cas12a)(nCpf1，左上)相结合。如在本文中更详细地描述的，nCpf1可以共价连接至逆转录酶(RT)(经由例如肽)，或者RT可以被募集至nCpf1(例如，经由使用肽标签基序/与所述肽标签相结合的亲和多肽，或经由化学相互作用，如在本文中所描述的)，在这种情况下，可以募集多个逆转录酶蛋白(RTn)。指导RNA的3'末端与DNA在所述切口位点处互补(非粗体配对线，左上)。然后，RT从DNA切口的3'末端开始对DNA进行聚合，从而生成与RNA互补的DNA序列，具有与基因组不互补的核苷酸(粗体配对线，大括号，右上)，随后为互补的核苷酸(非粗体配对线，右上)。解离后，所得的DNA具有延伸型ssDNA，其具有3'突出端，其是与原始DNA(非粗体配对线，右下)在很大程度上相同的序列，但是具有一些非天然核苷酸(粗体配对线，大括号，右下)。在存在掺入到DNA中的错配的核苷酸的情况下，该翼片与具有5'突出端的结构(左下)处于平衡状态。该平衡更多地位于右边的有利的完美配对，但是可以被驱动，可以以各种各样的方式来减少，例如对第二链(例如，非靶标链或上部链)产生切口。左边的结构可以优先地被在DNA滞后链合成中所牵涉的细胞翼片核酸内切酶(其在哺乳动物和植物细胞之间是高度保守的(智人(Homo sapiens)FEN1的氨基酸序列与玉蜀黍和大豆FEN1的氨基酸序列均是超过50％同一的))切割。在一些实施方案中，可以引入翼片核酸内切酶以在包含非天然/错配的核苷酸的3'翼片的方向上驱动所述平衡。在真核细胞中，经常由Dna2蛋白来去除较长的5'翼片，从而再次将所述平衡驱动至3'翼片(所希望的)产物(参见例如，Nucleic Acids Res.2012年8月,40(14):6774-86)。
进一步地，在本发明的方法中，和如在图2中所例示的，为了减少错配修复和为了驱动所述平衡更有利于形成具有经修饰的核苷酸(粗体，大括号)的最终产物，可以将Cpf1切口酶靶向在RT编辑区域之外的区域(闪电箭头)，如在本文中所描述的。nCpf1:crRNA分子可以在编辑泡的任何一侧或两侧。对第一链(例如，图2的靶标链或下部链)(虚线)产生切口向细胞表明，在错配修复和复制期间，新掺入的核苷酸是正确的核苷酸，因此有利于具有新核苷酸的最终产物。
逆转录酶(RT)的变体可以对于编辑系统的温度敏感性和持续能力(processivity)具有显著效应。RT的天然的和经合理和非合理改造的(即，定向进化)变体可以是在植物偏爱的温度下优化活性之中和对于优化持续能力特性谱来说有用的。
与RT多肽的蛋白质结构域融合可以对于编辑系统的温度敏感性和持续能力具有显著效应。RT酶可以通过与ssRNA结合结构域(RBD)相融合来就温度敏感性、持续能力和模板亲和力进行改善。这些RBD可以具有序列特异性、非特异性或序列偏爱(参见例如，SEQ IDNO:37-52)。一系列的亲和力分布可以对于在不同的细胞和体外环境中的编辑来说是有益的。RBD可以通过增加或减小RBD的大小来在特异性和结合自由能方面进行修饰，以便识别更多或更少的核苷酸。多个RBD导致具有作为独个RBD的组合的亲和力分布的蛋白质。向RT酶添加一个或多个RBD可以导致增加的亲和力、增加的或降低的序列特异性和/或启动子协同性。
可以将用于本发明的RT多肽与单链RNA结合蛋白(RBD)相融合。对本发明来说有用的RBD可以为从例如人、小鼠或蝇中获得的RBD。单链结合蛋白可以包含这样的氨基酸序列，其包括但不限于SEQ ID NO:37-52中的任一个。
在逆转录酶将编辑掺入到基因组中后，在新合成的经编辑的序列和它所意欲替换的原始WT序列之间存在序列冗余。这导致在靶位点处的5'或3'翼片，其不得不由细胞进行修复。这两种状态与有利于3'翼片的结合能处于平衡状态，因为当WT序列与其互补物配对时比当经编辑的链与其互补物配对时更多的碱基对是可得的。这对于有效编辑来说是不利的，因为3'翼片的加工(去除)可能去除经编辑的残基并且使靶标恢复至WT序列。然而，细胞翼片核酸内切酶例如FEN1或Dna2可以有效地加工5'翼片。因此，代替依赖于对于细胞来说天然的5'-翼片核酸内切酶的功能，在本发明的一些实施方案中，可以将在靶标处的翼片核酸内切酶的浓度增加至进一步有利于所希望的平衡结果(去除5'翼片中的WT序列，从而经编辑的序列变成稳定地掺入在靶位点处)。这可以通过在细胞中将5'翼片核酸内切酶过表达为游离蛋白质来取得。备选地，可以通过与CRISPR复合物相联合来将FEN或Dna2主动地募集至靶位点，要么通过直接的蛋白质融合，要么通过非共价募集例如用肽标签和亲和多肽对(例如，SunTag抗体/表位对)或化学相互作用，如在本文中所描述的。
本发明进一步提供了用于通过使用本发明的蛋白质/多肽和/或融合蛋白，和编码其的多核苷酸和核酸构建体，和/或包含其的表达盒和/或载体来修饰靶核酸的方法。所述方法可以在体内系统中(例如，在细胞中或在生物中)或在体外系统中(例如，无细胞系统)进行。因此，在一些实施方案中，提供了在植物细胞中修饰靶核酸的方法，所述方法包括：使所述靶核酸与下列组分相接触：(a)V型CRISPR-Cas效应蛋白或II型CRISPR-Cas效应蛋白；(b)逆转录酶，和(c)延伸型指导核酸(例如，延伸型II型或V型CRISPR RNA、延伸型II型或V型CRISPR DNA、延伸型II型或V型crRNA、延伸型II型或V型crDNA；例如tagRNA、tagDNA)，由此修饰所述靶核酸。在一些实施方案中，所述V型CRISPR-Cas效应蛋白或II型CRISPR-Cas效应蛋白、所述逆转录酶和所述延伸型指导核酸可以形成复合物或可以被包含在复合物中，所述复合物能够与所述靶核酸相互作用。在一些实施方案中，本发明的方法可以进一步包括使所述靶核酸与下列组分相接触：(a)第二V型CRISPR-Cas效应蛋白或第二II型CRISPR-Cas效应蛋白；(b)第二逆转录酶；和(c)第二延伸型指导核酸(例如，延伸型CRISPR RNA、延伸型CRISPR DNA、延伸型crRNA、延伸型crDNA；例如，tagDNA、tagRNA)，其中所述第二延伸型指导核酸靶向在所述靶核酸的第一链上的位点(间隔子与之实质上互补/与之相结合)，由此修饰所述靶核酸。在一些实施方案中，本发明的方法可以进一步包括使所述靶核酸与下列组分相接触：(a)第二V型CRISPR-Cas效应蛋白或第二II型CRISPR-Cas效应蛋白；(b)第二逆转录酶；和(c)第二延伸型指导核酸(例如，延伸型CRISPR RNA、延伸型CRISPR DNA、延伸型crRNA、延伸型crDNA；例如，tagDNA、tagRNA)，其中所述第二延伸型指导核酸靶向在所述靶核酸的第二链上的位点(间隔子与之实质上互补/与之相结合)，由此修饰所述靶核酸。在一些实施方案中，本发明的方法包括使所述靶核酸在大约20℃至42℃(例如，大约20、21、22、23、24、25、26、27、28、29、30、31、32、33、34、35、36、37、38、39、40、41或42℃，和其中的任何值或范围)的温度下进行接触。在一些实施方案中，可以使靶核酸与另外的多肽和/或编码其的核酸构建体相接触以便改善错配修复。在一些实施方案中，本发明的方法可以进一步包括使所述靶核酸与下列组分相接触：(a)CRISPR-Cas效应蛋白；和(b)指导核酸，其中(i)所述CRISPR-Cas效应蛋白为切口酶(例如，nCas9、nCas12a)并且对于位于以5'或3'离已经由所述II型或V型CRISPR-Cas效应蛋白产生切口的在第一链上的位点大约10至大约125个碱基对(例如，大约10、11、12、13、14、15、16、17、18、19、20、21、22、23、24、25、26、27、28、29、30、31、32、33、34、35、36、37、38、39、40、41、42、43、44、45、46、47、48、49、50、51、52、53、54、55、56、57、58、59、60、61、62、63、64、65、66、67、68、69、70、71、72、73、74、75、76、77、78、79、80、81、82、83、84、85、86、87、88、89、90、91、92、93、94、95、96、97、98、99、100、101、102、103、104、105、106、107、108、109、110、111、112、113、114、115、116、117、118、119、120、121、122、123、124或125个碱基对，或者其中的任何范围或值)处的在所述靶核酸的第一链上的位点产生切口，或(ii)所述CRISPR-Cas效应蛋白为切口酶(例如，nCas9、nCas12a)并且对于位于离已经由所述II型或V型CRISPR-Cas效应蛋白产生切口的在第一链上的位点大约10至大约125个碱基对处(5'或3')的在所述靶核酸的第二链上的位点产生切口，由此改善错配修复。在一些实施方案中，对于所述靶核酸的第二链(非靶标链)产生切口包括使所述靶核酸与包含具有错配(例如，大约1、2、3、或4个错配；例如，与第二链(非靶标链)大约80-96％互补的)的间隔子的crRNA相接触。在该构造中，产生切口并不改变所述平衡，而是将会利于错配产物种类形成，因为5’翼片在细胞中将会优先被加工。非靶标链的切口产生增强了下一步骤的分辨力，这是当牵涉错配修复时。
因此，在一些实施方案中，至少两种RNA可以用于本发明的方法：tagRNA，其将CRISPR-Cas效应蛋白指导至右边的斑点并且通过使用完美的RNA:DNA匹配来产生双链断裂；和第二RNA(crRNA)，其在相同的链上非常近地与DNA退火。该第二RNA(crRNA)具有间隔子序列，其包含几个错配(不完全互补的，例如，大约1、2、3或4个错配，例如大约80％至大约96％(80、81、82、83、84、85、86、87、88、89、90、91、92、93、94、95、96％)互补性)。这导致所述CRISPR-Cas效应蛋白对于非靶标链产生切口。虽然并不意欲局限于任何特定的理论，但是认为，在产生切口后，存在向后咀嚼(chew back)非靶标链中的一些的细胞核酸酶。在没有非靶标链的情况下，不存在用于“固定”我们正在用本发明的方法(REDRAW)制备的编辑的蓝图，从而导致更大的编辑效率。在该构造中，仅使用一种融合酶，但是它能够结合所述两种不同的RNA(tagRNA和crRNA)。
在一些实施方案中，延伸型指导核酸包含:(i)V型CRISPR核酸或II型CRISPR核酸(II型或V型CRISPR RNA、II型或V型CRISPR DNA、II型或V型crRNA、II型或V型crDNA)和/或CRISPR核酸和tracr核酸(例如，II型或V型tracrRNA、II型或V型tracrDNA)；和(ii)包含引物结合位点和逆转录酶模板(RT模板)的延伸部分。在一些实施方案中，所述延伸部分可以融合至所述CRISPR核酸的5'末端或3'末端(例如，5'至3'：重复序列-间隔子-延伸部分，或者延伸部分-重复序列-间隔子)，和/或融合至所述tracr核酸的5'或3'末端。在一些实施方案中，延伸型指导核酸的延伸部分从5'至3'包含RT模板(RTT)和引物结合位点(PBS)(例如，5’-crRNA-间隔子-RTT(编辑编码的)-PBS-3’)或者从5'至3'包含PBS和RTT，取决于所述延伸部分相对于所述指导的CRISPR RNA而言的位置(例如，5’-crRNA-间隔子-PBS-RTT(编辑编码的)-3’)。在一些实施方案中，靶核酸是双链的并且包含第一链和第二链，并且所述引物结合位点与所述靶核酸的第二链(非靶标，上部链)相结合。在一些实施方案中，靶核酸是双链的并且包含第一链和第二链，并且所述引物结合位点与所述靶核酸的第一链相结合(例如，结合至所述靶标链，所述CRISPR-Cas效应蛋白被募集至的同一链，下部链)。在一些实施方案中，靶核酸是双链的并且包含第一链和第二链，并且所述引物结合位点与所述靶核酸的第二链(非靶标链，与所述CRISPR-Cas效应蛋白被募集至的链相反的链)相结合。因此，在一些实施方案中，所述编辑性逆转录酶(RT)添加至靶标链(所述CRISPR RNA的间隔子所与之互补的并且所述CRISPR-Cas效应蛋白被募集至的链)，并且在一些实施方案中，所述编辑性逆转录酶(RT)添加至非靶标链(与下述链互补的链：所述CRISPR RNA的间隔子所与之互补的并且所述CRISPR-Cas效应蛋白被募集至的链)。
所述RT模板编码待被掺入到所述靶核酸(编辑)中的修饰。编辑的修饰可以位于在RT模板内的任何位置(相对于所述靶核酸的原间隔子邻近基序(PAM)的位置而言的位置定位)。因此，例如，图27显示了具有位于相对于在所述靶核酸中的原间隔子邻近基序(PAM)(TTTG)的位置而言的位置-1至19(-1、1、2、3、4、5、6、7、8、9、10、11、12、13、14、15、16、27、18或19)处的编辑的RT模板。在每种情况下，观察到精确编辑。在一些实施方案中，RT模板可以包含位于核苷酸位置-1、1、2、3、4、5、6、7、8、9、10、11、12、13、14、15、16、27、18或19处的编辑。在一些实施方案中，RT模板可以包含位于相对于所述靶核酸的原间隔子邻近基序(PAM)的位置而言的所述RT模板的核苷酸位置4至核苷酸位置17(例如，位置4、5、6、7、8、9、10、11、12、13、14、15、16或17)处的编辑。在一些实施方案中，RT模板可以包含位于相对于所述靶核酸的原间隔子邻近基序(PAM)的位置而言的所述RT模板的核苷酸位置10至核苷酸位置17(例如，位置10、11、12、13、14、15、16或17)处的编辑。在一些实施方案中，RT模板可以包含位于相对于所述靶核酸的原间隔子邻近基序(PAM)的位置而言的所述RT模板的核苷酸位置12至核苷酸位置15(例如，位置12、13、14或15)处的编辑。
在一些实施方案中，提供了修饰具有第一链和第二链的靶核酸的方法，所述方法包括：使所述靶核酸与下列组分相接触：(a)V型CRISPR-Cas效应蛋白或II型CRISPR-Cas效应蛋白；(b)逆转录酶，和(c)延伸型指导核酸(例如，延伸型II型或V型CRISPR RNA、延伸型II型或V型CRISPR DNA、延伸型II型或V型crRNA、延伸型II型或V型crDNA)，其中所述延伸型指导核酸包含：(i)II型或V型CRISPR核酸(II型或V型CRISPR RNA、II型或V型CRISPR DNA、II型或V型crRNA、II型或V型crDNA)和/或CRISPR核酸和tracr核酸(例如，II型或V型tracrRNA、II型或V型tracrDNA)；和(ii)包含引物结合位点和逆转录酶模板(RT模板)的延伸部分，并且所述II型或V型CRISPR核酸包含与所述第一链(例如，靶标链)相结合(即，与在所述靶核酸的第一链中的一部分的连续核苷酸互补)的间隔子，并且所述引物结合位点与所述第一链(靶标链)相结合，由此修饰所述靶核酸。在一些实施方案中，II型CRISPR-Cas效应蛋白可以为Cas9多肽，任选地spCas9。在一些实施方案中，V型CRISPR-Cas效应蛋白可以为Cas12a多肽或cas12b多肽。在一些实施方案中，II型或V型CRISPR-Cas效应蛋白、逆转录酶和延伸型指导核酸可以形成复合物或被包含在复合物中。在一些实施方案中，接触可以进一步包括使所述靶核酸与5'-3'核酸外切酶相接触。
在一些实施方案中，所述靶核酸可以另外与5'翼片核酸内切酶(FEN)(任选地，FEN1和/或Dna2多肽)相接触，由此通过去除不包含待被掺入到所述靶核酸中的5′翼片来改善错配修复。在一些实施方案中，FEN和/或Dna2可以在所述靶核酸存在下过表达。在一些实施方案中，FEN可以为包含融合至V型CRISPR-Cas效应蛋白或结构域的FEN结构域的融合蛋白，由此将所述FEN募集至所述靶核酸。在一些实施方案中，Dna2可以为包含融合至V型CRISPR-Cas效应蛋白或结构域的Dna2结构域的融合蛋白，由此将所述Dna2募集至所述靶核酸。
在一些实施方案中，II型或V型CRISPR-Cas效应蛋白可以为包含融合(连接)至肽标签(例如，表位或多聚化表位)的V型CRISPR-Cas效应蛋白结构域的II型或V型CRISPR-Cas融合蛋白，并且FEN可以为包含融合至与所述肽标签相结合的亲和多肽的FEN结构域的FEN融合蛋白，由此将所述FEN募集至所述II型或V型CRISPR-Cas效应蛋白结构域和所述靶核酸。在一些实施方案中，II型或V型CRISPR-Cas效应蛋白可以为包含融合(连接)至肽标签(例如，表位或多聚化表位)的II型或V型CRISPR-Cas效应蛋白结构域的II型或V型CRISPR-Cas融合蛋白，并且Dna2可以为包含融合至与所述肽标签相结合的亲和多肽的Dna2结构域的Dna2融合蛋白，由此将所述Dna2募集至所述II型或V型CRISPR-Cas效应蛋白结构域和所述靶核酸。在一些实施方案中，V型CRISPR-Cas效应蛋白可以为包含融合(连接)至肽标签(例如，表位或多聚化表位)的II型或V型CRISPR-Cas效应蛋白结构域的II型或V型CRISPR-Cas融合蛋白，并且FEN可以为包含融合至与所述肽标签相结合的亲和多肽的FEN结构域的FEN融合蛋白，由此将所述FEN募集至所述II型或V型CRISPR-Cas效应蛋白结构域和所述靶核酸。在一些实施方案中，II型或V型CRISPR-Cas效应蛋白可以为包含融合(连接)至肽标签(例如，表位或多聚化表位)的II型或V型CRISPR-Cas效应蛋白结构域的II型或V型CRISPR-Cas融合蛋白，并且Dna2可以为包含融合至与所述肽标签相结合的亲和多肽的Dna2结构域的Dna2融合蛋白，由此将所述Dna2募集至所述II型或V型CRISPR-Cas效应蛋白结构域和所述靶核酸。在一些实施方案中，靶核酸可以与两个或更多个FEN融合蛋白和/或Dna2融合蛋白相接触。
在一些实施方案中，本发明的方法可以进一步包括使所述靶核酸与5'-3'核酸外切酶相接触，由此通过去除不包含待被掺入到所述靶核酸中的编辑的5'翼片(未编辑的链)来改善错配修复。在一些实施方案中，5'-3'核酸外切酶可以融合至II型或V型CRISPR-Cas效应蛋白，任选地融合至II型或V型CRISPR-Cas融合蛋白。在一些实施方案中，5'-3'核酸外切酶可以为包含融合至肽标签的5'-3'核酸外切酶的融合蛋白，并且II型或V型CRISPR-Cas效应蛋白可以为包含融合至能够与所述肽标签相结合的亲和多肽的II型或V型CRISPR-Cas效应蛋白结构域的融合蛋白，由此改善错配修复。在一些实施方案中，5'-3'核酸外切酶可以为包含融合至能够与所述肽标签相结合的亲和多肽的5'-3'核酸外切酶的融合蛋白，并且II型或V型CRISPR-Cas效应蛋白可以为包含融合至肽标签的II型或V型CRISPR-Cas效应蛋白结构域的融合蛋白。在一些实施方案中，5'-3'核酸外切酶可以为包含融合至能够与RNA募集基序相结合的亲和多肽的5'-3'核酸外切酶的融合蛋白，并且所述延伸型指导核酸连接至RNA募集基序，由此经由亲和多肽与RNA募集基序之间的相互作用来将所述5'-3'核酸外切酶募集至所述靶核酸。5'-3'核酸外切酶可以为任何已知的或后来发现的在目的生物、细胞或体外系统中有功能的5'-3'核酸外切酶。在一些实施方案中，5'-3'核酸外切酶可以包括但不限于：RecE核酸外切酶(RecE，例如SEQ ID NO:129)，RecJ核酸外切酶(RecJ，例如SEQ ID NO:130)，T5核酸外切酶(T5_Exo，例如SEQ ID NO:131)，和/或T7核酸外切酶(T7_Exo，例如SEQ ID NO:132)，λ核酸外切酶(λ_Exo，例如SEQ ID NO:133)，大肠杆菌核酸外切酶sbcB(SEQ ID NO:134)，和/或人核酸外切酶(Exo，例如SEQ ID NO:135)。在一些实施方案中，可以使用在两侧侧翼为核定位序列(NLS)(其来自例如大肠杆菌(菌株K12))的RecE核酸外切酶C-末端片段(SEQ ID NO:98)。在一些实施方案中，可以使用在两侧侧翼为核定位序列(NLS)(其来自例如大肠杆菌(菌株K12))的RecJ核酸外切酶(SEQ ID NO:99)。在一些实施方案中，可以使用在两侧侧翼为核定位序列(NLS)的T5核酸外切酶(SEQ ID NO:100)。在一些实施方案中，可以使用在两侧侧翼为核定位序列(NLS)(其来自例如埃希氏菌噬菌体7)的T7核酸外切酶(SEQ ID NO:101)。在一些实施方案中，5'-3'核酸外切酶包括但不限于：RecE(例如，SEQ ID NO:129)、RecJ(例如，SEQ ID NO:130)、T5_Exo(例如，SEQ ID NO:131)、T7_Exo(例如，SEQ ID NO:132)、sbcB(SEQ ID NO:134)和/或Exo(SEQ ID NO:135)。
在一些实施方案中，本发明的方法可以进一步包括减少双链断裂。在一些实施方案中，减少双链断裂可以通过在所述靶核酸的区域中引入非同源末端连接(NHEJ)的化学抑制剂或者通过引入靶向NHEJ蛋白的CRISPR指导核酸或siRNA以瞬时敲低所述NHEJ蛋白的表达来进行。在一些实施方案中，NHEJ的抑制剂可以融合至本发明的逆转录酶(RT)或CRISPR-Cas效应蛋白，任选地融合至所述RT或CRISPR-Cas效应蛋白的N-末端。在一些实施方案中，NHEJ的抑制剂包括但不限于埃希氏菌噬菌体Mu Gam(SEQ ID NO:147)。
在一些实施方案中，II型或V型CRISPR-Cas效应蛋白可以为融合蛋白，和/或所述逆转录酶可以为融合蛋白，其中所述II型或V型CRISPR-Cas融合蛋白、所述逆转录酶融合蛋白和/或所述延伸型指导核酸可以融合至一个或多个允许将所述逆转录酶募集至所述II型或V型CRISPR-Cas效应蛋白的组分。在一些实施方案中，所述一个或多个组分经由蛋白质-蛋白质相互作用、蛋白质-RNA相互作用和/或化学相互作用来进行募集。
因此，在一些实施方案中，V型CRISPR-Cas效应蛋白可以为包含融合(连接)至肽标签(例如，表位或多聚化表位)的V型CRISPR-Cas效应蛋白结构域的V型CRISPR-Cas效应融合蛋白，并且所述逆转录酶可以为包含融合(连接)至与所述肽标签相结合的亲和多肽的逆转录酶结构域的逆转录酶融合蛋白，其中所述V型CRISPR-Cas效应蛋白与所述指导核酸相互作用，所述指导核酸与所述靶核酸相结合，由此将所述逆转录酶募集至所述V型CRISPR-Cas效应蛋白和所述靶核酸。在一些实施方案中，所述II型CRISPR-Cas效应蛋白为包含融合(连接)至肽标签(例如，表位或多聚化表位)的II型CRISPR-Cas效应蛋白结构域的II型CRISPR-Cas融合蛋白，并且所述FEN为包含融合至与所述肽标签相结合的亲和多肽的FEN结构域的FEN融合蛋白，和/或所述II型CRISPR-Cas效应蛋白为包含融合至肽标签的II型CRISPR-Cas效应蛋白结构域的II型CRISPR-Cas融合蛋白，并且所述Dna2多肽为包含融合至与所述肽标签相结合的亲和多肽的Dna2结构域的Dna2融合蛋白，任选地其中所述靶核酸与两个或更多个FEN融合蛋白和/或两个或更多个Dna2融合蛋白相接触，由此将所述FEN和/或Dna2募集至所述II型CRISPR-Cas效应蛋白结构域和所述靶核酸。在一些实施方案中，可以将两个或更多个逆转录酶融合蛋白募集至所述II型或V型CRISPR-Cas效应蛋白，由此使所述靶核酸与两个或更多个逆转录酶融合蛋白相接触。
肽标签可以包括但不限于：GCN4肽标签(例如，Sun-Tag)、c-Myc亲和标签、HA亲和标签、His亲和标签、S亲和标签、甲硫氨酸-His亲和标签、RGD-His亲和标签、FLAG八肽、strep标签或strep标签II、V5标签和/或VSV-G表位。对本发明可以使用任何这样的表位，其可以与多肽相连接并且对于其存在有可以被连接至另一个多肽的相应的亲和多肽。在一些实施方案中，肽标签可以包含1或2或更多个拷贝的肽标签(例如，表位、多聚化表位(例如，串联重复序列))
17、18、19、20、21、22、23、24、25或更多个肽标签)。在一些实施方案中，与肽标签相结合的亲和多肽可以为抗体。在一些实施方案中，所述抗体可以为scFv抗体。在一些实施方案中，与肽标签相结合的亲和多肽可以是合成的(例如，为亲和相互作用而进化的)，其包括但不限于亲和体(affibody)、anticalin、monobody和/或DARPin(参见例如，Sha等人,Protein Sci.26(5):910-924(2017))；Gilbreth(Curr Opin Struc Biol 22(4):413-420(2013)；美国专利号9,982,053，其各自通过提及而以其整体合并以用于与亲和体、anticalin、monobody和/或DARPin相关的教导)。实例性的肽标签序列和其亲和多肽包括但不限于SEQ ID NO:23-25的氨基酸序列。
在一些实施方案中，延伸型指导核酸可以连接至RNA募集基序，并且所述逆转录酶可以为逆转录酶融合蛋白，其中所述逆转录酶融合蛋白包含融合至与所述RNA募集基序相结合的亲和多肽的逆转录酶结构域，其中所述延伸型指导与所述靶核酸相结合并且所述RNA募集基序与所述亲和多肽相结合，由此将所述逆转录酶融合蛋白募集至所述延伸型指导并且使所述靶核酸与所述逆转录酶结构域相接触。在一些实施方案中，可以将两个或更多个逆转录酶融合蛋白募集至延伸型指导核酸，由此使所述靶核酸与两个或更多个逆转录酶融合蛋白相接触。实例性的RNA募集基序和其亲和多肽包括但不限于SEQ ID NO:26-36的序列。
在一些实施方案中，RNA募集基序可以位于所述延伸型指导核酸的延伸部分的3'末端处(例如，5'-3'，重复序列-间隔子-延伸部分(RT模板-引物结合位点)-RNA募集基序)。在一些实施方案中，RNA募集基序可以被包埋在所述延伸部分中。
在本发明的一些实施方案中，延伸型指导RNA和/或指导RNA可以连接至一个或连接至两个或更多个RNA募集基序(例如，1、2、3、4、5、6、7、8、9、10或更多个基序，例如，至少10至大约25个基序)，任选地其中所述两个或更多个RNA募集基序可以是相同的RNA募集基序或不同的RNA募集基序。在一些实施方案中，RNA募集基序和相应的亲和多肽可以包括但不限于：端粒酶Ku结合基序(例如，Ku结合发夹)和相应的亲和多肽Ku(例如，Ku异二聚体)，端粒酶Sm7结合基序和相应的亲和多肽Sm7，MS2噬菌体操纵基因茎-环和相应的亲和多肽MS2外壳蛋白(MCP)，PP7噬菌体操纵基因茎-环和相应的亲和多肽PP7外壳蛋白(PCP)，SfMu噬菌体Com茎-环和相应的亲和多肽Com RNA结合蛋白，PUF结合位点(PBS)和亲和多肽Pumilio/fem-3mRNA结合因子(PUF)，和/或合成的RNA-适体和作为相应的亲和多肽的适体配体。在一些实施方案中，所述RNA募集基序和相应的亲和多肽可以为MS2噬菌体操纵基因茎-环和亲和多肽MS2外壳蛋白(MCP)。在一些实施方案中，所述RNA募集基序和相应的亲和多肽可以为PUF结合位点(PBS)和亲和多肽Pumilio/fem-3mRNA结合因子(PUF)。
在一些实施方案中，用于募集多肽和核酸的组分可以为通过化学相互作用发挥功能的那些，其可以包括但不限于：雷帕霉素可诱导的FRB–FKBP的二聚化；生物素-链霉抗生物素蛋白；SNAP标签；Halo标签；CLIP标签；由化合物诱导的DmrA-DmrC异二聚体；双功能配体(例如，两个蛋白质结合化学品融合在一起，例如二氢叶酸还原酶(DHFR))。
在本发明的一些实施方案中，CRISPR-Cas效应蛋白(例如，CRISPR-Cas效应蛋白，第一CRISPR-Cas效应蛋白、第二CRISPR-Cas效应蛋白、第三CRISPR-Cas效应蛋白和/或第四CRISPR-Cas效应蛋白)可以来自I型CRISPR-Cas系统、II型CRISPR-Cas系统、III型CRISPR-Cas系统、IV型CRISPR-Cas系统和/或V型CRISPR-Cas系统。在一些实施方案中，所述CRISPR-Cas核酸酶来自II型CRISPR-Cas系统或V型CRISPR-Cas系统。
在本发明的一些实施方案中，CRISPR-Cas效应蛋白可以为Cas9、C2c1、C2c3、Cas12a(也称为Cpf1)、Cas12b、Cas12c、Cas12d、Cas12e、Cas13a、Cas13b、Cas13c、Cas13d、Casl、CaslB、Cas2、Cas3、Cas3'、Cas3”、Cas4、Cas5、Cas6、Cas7、Cas8、Cas9(也称为Csnl和Csx12)、Cas10、Csyl、Csy2、Csy3、Csel、Cse2、Cscl、Csc2、Csa5、Csn2、Csm2、Csm3、Csm4、Csm5、Csm6、Cmrl、Cmr3、Cmr4、Cmr5、Cmr6、Csbl、Csb2、Csb3、Csxl7、Csxl4、Csx10、Csx16、CsaX、Csx3、Csxl、Csxl5、Csfl、Csf2、Csf3、Csf4(dinG)和/或Csf5核酸酶，任选地其中所述CRISPR-Cas核酸酶可以为Cas9、Cas12a(Cpf1)、Cas12b、Cas12c(C2c3)、Cas12d(CasY)、Cas12e(CasX)、Cas12g、Cas12h、Cas12i、C2c4、C2c5、C2c8、C2c9、C2c10、Cas14a、Cas14b和/或Cas14c核酸酶。
在一些实施方案中，CRISPR-Cas效应蛋白可以为作为切口酶发挥功能的蛋白质(例如，Cas9切口酶或Cas12a切口酶)。在一些实施方案中，对本发明来说有用的CRISPR-Cas效应蛋白可以在其核酸酶活性位点(例如，RuvC、HNH，例如Cas12a核酸酶结构域的RuvC位点，例如Cas9核酸酶结构域的RuvC位点和/或HNH位点)中包含突变。在其核酸酶活性位点中具有突变并且因此不再包含核酸酶活性的CRISPR-Cas效应蛋白通常被称为“死的”或“失活的”，例如dCas。在一些实施方案中，在其核酸酶活性位点中具有突变的CRISPR-Cas核酸酶结构域或多肽可以具有相比于没有该突变的相同的CRISPR-Cas核酸酶而言受损的活性或降低的活性。在一些实施方案中，对本发明来说有用的CRISPR-Cas效应蛋白可以为双链核酸酶。在一些实施方案在中，具有双链核酸酶活性的CRISPR-Cas效应蛋白可以为II型或V型CRISPR-Cas效应蛋白。在一些实施方案中，具有双链核酸酶活性的V型CRISPR-Cas效应蛋白为Cas12a多肽。在一些实施方案中，具有双链核酸酶活性的II型CRISPR-Cas效应蛋白为Cas9多肽。
在一些实施方案中，CRISPR-Cas效应蛋白可以为V型CRISPR-Cas效应蛋白。在一些实施方案中，V型CRISPR-Cas效应蛋白可以包含Cas12a(Cpf1)、Cas12b(C2c1)、Cas12c(C2c3)、Cas12d(CasY)、Cas12e(CasX)、Cas12g、Cas12h、Cas12i、C2c4、C2c5、C2c8、C2c9、C2c10、Cas14a、Cas14b和/或Cas14c效应蛋白和/或结构域。
在一些实施方案中，Cas12a(Cpf1)可以包括但不限于：LbCas12a、Lb2Cas12a、Lb3Cas12a、AsCas12a、BpCas12a、CMtCas12a、EeCas12a、FnCas12a、LiCas12a、MbCas12a、PbCas12a、PcCas12a、PdCas12a、PeCas12a、PmCas12a、SsCas12a、enAsCas12a，任选地其中所述Cas12a包含一个或多个在本文中所描述的突变。在一些实施方案中，Cas12b(C2c1)可以包括但不限于BhCas12b，任选地其中所述Cas12b包含一个或多个在本文中所描述的突变。
在一些实施方案中，V型CRISPR-Cas效应蛋白可以包括但不限于：来自氨基酸球菌属物种(Acidaminococcus sp.)的V型CRISPR-Cas效应蛋白(AsCas12a)、来自毛螺菌科(Lachnospiraceae)细菌的V型CRISPR-Cas效应蛋白(例如，LbCas12a)或来自亨盖特氏丁酸弧菌(Butyrivibrio hungatei)的V型CRISPR-Cas效应蛋白(BhCas12b)，或者其经修饰的V型CRISPR-Cas效应蛋白。在一些实施方案中，来自氨基酸球菌属物种的V型CRISPR-Cas效应蛋白可以包含与SEQ ID NO:2具有至少80％同一性的序列。在一些实施方案中，来自毛螺菌科细菌的V型CRISPR-Cas效应蛋白可以包含与SEQ ID NO:1、SEQ ID NO:7、SEQ ID NO:8或SEQ ID NO:9中的任一个具有至少80％同一性的氨基酸序列。在一些实施方案中，来自亨盖特氏丁酸弧菌的V型CRISPR-Cas效应蛋白可以包含与SEQ ID NO:151的氨基酸序列具有至少80％同一性的序列。在一些实施方案中，经修饰的来自氨基酸球菌属物种的V型CRISPR-Cas效应蛋白可以包含与SEQ ID NO:18、SEQ ID NO:19、SEQ ID NO:20、SEQ ID NO:149或SEQ ID NO:150中的任一个具有至少80％同一性的氨基酸序列。在一些实施方案中，经修饰的来自毛螺菌科细菌的V型CRISPR-Cas效应蛋白可以包含与SEQ ID NO:148具有至少80％同一性的序列。
在一些实施方案中，II型CRISPR-Cas效应蛋白可以包括但不限于Cas9效应蛋白，任选地其中所述Cas9效应蛋白可以来自链球菌属(Streptococcus)，任选地来自酿脓链球菌(Streptococcus pyogenes)。在一些实施方案中，Cas9效应蛋白可以为经修饰的Cas9效应蛋白。在一些实施方案中，Cas9效应蛋白可以包含与SEQ ID NO:106或SEQ ID NO:107中的任一个具有至少80％同一性的多肽序列。在一些实施方案中，Cas9效应蛋白可以由与SEQID NO:108-122中的任一个具有至少80％同一性的多核苷酸序列编码。
在一些实施方案中，V型CRISPR-Cas系统可以包含仅利用V型CRISPR核酸的效应蛋白。在一些实施方案中，V型CRISPR-Cas系统可以包含这样的效应蛋白，其利用CRISPR核酸和反式激活性CRISPR(tracr)核酸两者，这与II型CRISPR-Cas系统相似。因此，在一些实施方案中，对本发明来说有用的V型CRISPR-Cas效应蛋白可以仅与相应的CRISPR核酸一起发挥功能(例如，Cas12a、Cas12a、Cas12i、Cas12h、Cas14b、Cas14c、C2c10、C2c9、C2c8、C2c4)。在一些实施方案中，对本发明来说有用的V型CRISPR-Cas效应蛋白可以与相应的CRISPR核酸和tracr核酸一起发挥功能(例如，Cas12b、Cas12c、Cas12e、Cas12g、Cas14a)。
对本发明来说有用的CRISPR核酸可以包含至少一个能够与相应的V型CRISPR-Cas效应蛋白相互作用的重复序列和至少一个间隔子序列，其中所述至少一个间隔子序列能够结合靶核酸(例如，所述靶核酸的第一链或第二链)。在一些实施方案中，CRISPR核酸的重复序列可以位于所述间隔子序列的5'。在一些实施方案中，CRISPR核酸可以包含多于一个重复序列，其中所述重复序列连接至所述间隔子的5'末端和3'末端两者。在一些实施方案中，对本发明来说有用的CRISPR核酸可以包含两个或更多个重复序列和一个或多个间隔子序列，其中每个间隔子序列在5'末端和3'末端处与重复序列相连接。
对本发明来说有用的tracr核酸可以包含与相应的CRISPR核酸的重复序列实质上互补并且与之杂交的第一部分，和与相应的II型或V型CRISPR-Cas效应蛋白相互作用的第二部分。
在一些实施方案中，对本发明来说有用的V型CRISPR-Cas效应蛋白可以作为双链DNA核酸酶发挥功能。在一些实施方案中，V型CRISPR-Cas效应蛋白可以作为单链DNA切口酶发挥功能，任选地其中对于所述第一链产生切口。在一些实施方案中，V型CRISPR-Cas效应蛋白可以作为单链DNA切口酶发挥功能，任选地其中对于所述第二链产生切口。在一些实施方案中，所述V型CRISPR-Cas效应蛋白可以为作为切口酶发挥功能的Cas12a效应蛋白，任选地其中对于所述第一链(靶标链)产生切口。在一些实施方案中，所述V型CRISPR-Cas效应蛋白可以为作为切口酶发挥功能的Cas12a效应蛋白，任选地其中对于所述第二链产生切口。在一些实施方案中，所述V型CRISPR-Cas效应蛋白可以为通过使用包含战略错配(strategic mismatch)的crRNA作为切口酶发挥功能的Cas12a效应蛋白。因此，例如，crRNA可以包含具有一个至大约四个错配(例如，1、2、3或4个错配)(例如，80-96％互补的)的间隔子。
在一些实施方案中，Cas12a效应蛋白可以为具有在LQMRNS基序中的精氨酸的突变的Cas12a切口酶。在该基序中的精氨酸的突变可以是至任何氨基酸的，由此提供了Cas12a切口酶。在一些实施方案中，所述突变可以是至丙氨酸。在一些实施方案中，所述突变可以是至丙氨酸、天冬酰胺、天冬氨酸、半胱氨酸、谷氨酸、谷氨酰胺、甘氨酸、组氨酸、异亮氨酸、亮氨酸、赖氨酸、甲硫氨酸、苯丙氨酸、脯氨酸、丝氨酸、苏氨酸、色氨酸、酪氨酸或缬氨酸。在一些实施方案中，所述突变可以是至丙氨酸的突变。在一些实施方案中，所述突变不包括至赖氨酸或组氨酸的突变。在一些实施方案中，Cas12a效应蛋白可以为包含下述突变的LbCas12a切口酶：R1138突变，任选地R1138A突变(参见参考核苷酸序列SEQ ID NO:9)；R1137突变，任选地R1137A突变(参见参考核苷酸序列SEQ ID NO:1)；或R1124突变，任选地R1124A突变(参见参考核苷酸序列SEQ ID NO:7)。在一些实施方案中，Cas12a效应蛋白可以为包含下述突变的AsCas12a切口酶：R1226突变，任选地R1226A突变(参见参考核苷酸序列SEQ ID NO:2)。在一些实施方案中，Cas12a效应蛋白可以为包含下述突变的FnCas12a切口酶：R1218突变，任选地R1218A突变(参见参考核苷酸序列SEQ ID NO:6)。在一些实施方案中，Cas12a效应蛋白可以为包含下述突变的PdCas12a切口酶：R1241突变，任选地R1241A突变(参见参考核苷酸序列SEQ ID NO:14)。
在一些实施方案中，对本发明来说有用的V型CRISPR-Cas效应蛋白可以包含降低的单链DNA切割活性(ss DNA酶活性)(例如，所述V型CRISPR-Cas效应蛋白可以经修改(突变)以降低ss DNA酶活性，例如比野生型或未修饰的V型CRISPR-Cas效应蛋白低大约10、15、20、25、30、35、40、45、50、55、60、65、70、75、80、85、90、95、96、97、98、99或100％的ss DNA酶活性)。
在一些实施方案中，对本发明来说有用的V型CRISPR-Cas效应蛋白可以包含降低的自加工RNA酶活性(例如，所述V型CRISPR-Cas效应蛋白可以经修饰(突变)以降低自加工RNA酶活性，例如比野生型或未修饰的V型CRISPR-Cas效应蛋白低大约10、15、20、25、30、35、40、45、50、55、60、65、70、75、80、85、90、95、96、97、98、99或100％的自加工RNA酶活性)。在一些实施方案中，降低自加工RNA酶活性的突变可以为在通过参考SEQ ID NO:1或SEQ ID NO:9的核苷酸位置编号而言的残基位置759处的组氨酸的突变，任选地组氨酸至丙氨酸的突变(H759A)。实例性的具有降低的单链DNA切割活性的V型CRISPR-Cas效应蛋白可以包括但不限于LbCas12a(H759A)(SEQ ID NO:148)。在一些实施方案中，对本发明来说有用的具有H759A突变的Cas12a CRISPR-Cas效应蛋白可以包含与SEQ ID NO:148的氨基酸序列具有至少80％序列同一性的序列。在一些实施方案中，具有H759A突变的Cas12a CRISPR-Cas效应蛋白可以为LbCas12a CRISPR-Cas效应蛋白，任选地其中所述LbCas12a CRISPR-Cas效应蛋白包含与SEQ ID NO:148的氨基酸序列的至少90％序列同一性。
在一些实施方案中，对本发明来说有用的V型CRISPR-Cas效应蛋白或结构域可以在其核酸酶活性位点(例如，d V型CRISPR-Cas效应蛋白或结构域的RuvC，例如Cas12a核酸酶结构域的RuvC位点)中包含突变。在其核酸酶活性位点中具有突变并且因此不再包含核酸酶活性的CRISPR-Cas核酸酶通常被称为“失活的”或“死的”，例如dCas、dCas12a。在一些实施方案中，在其核酸酶活性位点中具有突变的CRISPR-Cas核酸酶结构域或多肽可以具有相比于没有该突变的相同的CRISPR-Cas核酸酶而言受损的活性或降低的活性。在一些实施方案中，失活的V型CRISPR-Cas效应蛋白可以作为切口酶发挥功能(第一链切口酶和/或第二链切口酶)。
在一些实施方案中，对本发明来说有用的V型CRISPR-Cas效应蛋白或结构域可以包含一个或多个氨基酸残基的修饰，其降低所述V型CRISPR-Cas效应蛋白的DNA结合亲和力。在一些实施方案中，所述修饰可以为氨基酸置换。在一些实施方案中，可以使与DNA主链相互作用的带正电荷的残基进行突变，任选地其中可以将与DNA主链相互作用的带正电荷的残基突变为丙氨酸(例如，用丙氨酸置换)。在Cas12a效应蛋白中将带正电荷的残基置换为丙氨酸可以包括但不限于通过参考SEQ ID NO:1或SEQ ID NO:148的氨基酸位置编号而言的K167A、K272A和/或K349A的氨基酸置换。在一些实施方案中，所述V型CRISPR-Cas效应蛋白为包含通过参考SEQ ID NO:148的氨基酸位置编号而言的K167A、K272A、K349A、K167A+K272A、K167A+K349A、K272A+K349A或K167A+K272A+K349A的氨基酸置换的Cas12a CRISPR-Cas效应蛋白，任选地其中所述V型CRISPR-Cas效应蛋白为LbCas12a。
在一些实施方案中，V型CRISPR-Cas效应蛋白可以为V型CRISPR-Cas融合蛋白，其中所述V型CRISPR-Cas融合蛋白包含融合至逆转录酶的V型CRISPR-Cas效应蛋白结构域。在一些实施方案中，所述逆转录酶可以融合至所述V型CRISPR-Cas效应多肽的C-末端。在一些实施方案中，所述逆转录酶可以融合至所述V型CRISPR-Cas效应多肽的N-末端。
在一些实施方案中，V型CRISPR-Cas效应蛋白可以为V型CRISPR-Cas融合蛋白，其中所述V型CRISPR-Cas融合蛋白包含融合至产生切口的酶(例如，Fok1、BFi1，例如经改造的Fok1或BFiI)的V型CRISPR-Cas效应蛋白结构域，任选地其中所述V型CRISPR-Cas效应蛋白结构域可以为融合至所述产生切口的酶的失活的V型CRISPR-Cas结构域。
在一些实施方案中，II型CRISPR-Cas效应蛋白可以为II型CRISPR-Cas融合蛋白，其中所述II型CRISPR-Cas融合蛋白包含融合至逆转录酶的II型CRISPR-Cas效应蛋白结构域。在一些实施方案中，所述逆转录酶可以融合至所述II型CRISPR-Cas效应多肽的C-末端。在一些实施方案中，所述逆转录酶可以融合至所述II型CRISPR-Cas效应多肽的N-末端。在一些实施方案中，II型CRISPR-Cas效应蛋白可以为II型CRISPR-Cas融合蛋白，其中所述II型CRISPR-Cas融合蛋白包含融合至产生切口的酶(例如，Fok1、BFi1，例如经改造的Fok1或BFiI)的II型CRISPR-Cas效应蛋白结构域，任选地其中所述II型CRISPR-Cas效应蛋白结构域可以为融合至所述产生切口的酶的失活的II型CRISPR-Cas结构域。
在一些实施方案中，对本发明来说有用的逆转录酶可以为野生型逆转录酶。在一些实施方案中，对本发明来说有用的逆转录酶可以为合成的逆转录酶，参见例如，Heller等人,Nucleic acids Research,47(7)3619-3630(2019))。实例性的逆转录酶多肽包括但不限于与SEQ ID NO:53或SEQ ID NO:172的氨基酸序列具有实质上的同一性(例如，至少大约70％、71％、72％、73％、74％、75％、76％、77％、78％、79％、80％、81％、82％、83％、84％、85％、86％、87％、88％、89％、90％、91％、92％、93％、94％、95％、96％、97％、98％、99％、99.5％或100％同一性)的那些。
在一些实施方案中，可以就(V型或II型)基因编辑活性对逆转录酶的活性进行修饰，以提供最佳的与V型或II型CRISPR-Cas效应多肽相关联的活性(例如，相比于未进行修饰的参考逆转录酶而言大约5、10、15、20、25、30、35、40、45、50、55、60、65、70、75、80、85、90、95、96、97、98、99或100％的活性增加，当与V型CRISPR-Cas效应多肽相关联时)。此类突变包括影响或改善RT起始、持续能力、酶动力学、温度敏感性和/或错误率的那些。
在一些实施方案中，对本发明来说有用的逆转录酶可以进行修饰以改善所述逆转录酶的转录功能。逆转录酶的转录功能可以通过改善所述逆转录酶的持续能力来进行改善，例如，增加所述逆转录酶在与模板的单个结合事件期间(例如，在它从模板上脱落之前)聚合更多DNA碱基的能力(例如，相比于未进行修饰的参考逆转录酶而言将持续能力增加大约5、10、15、20、25、30、35、40、45、50、55、60、65、70、75、80、85、90、95、96、97、98、99或100％)。
在一些实施方案中，逆转录酶的转录功能可以通过增加所述逆转录酶的模板亲和力来进行改善(例如，相比于未进行修饰的参考逆转录酶而言将模板亲和力增加大约5、10、15、20、25、30、35、40、45、50、55、60、65、70、75、80、85、90、95、96、97、98、99或100％)。
在一些实施方案中，逆转录酶的转录功能可以通过就在所希望的温度下的经改善的性能而改善所述逆转录酶的热稳定性来进行改善(例如，相比于未进行修饰的参考逆转录酶而言将热稳定性增加大约5、10、15、20、25、30、35、40、45、50、55、60、65、70、75、80、85、90、95、96、97、98、99或100％)。在一些实施方案中，所述经改善的热稳定性在大约20℃至42℃(例如，大约20、21、22、23、24、25、26、27、28、29、30、31、32、33、34、35、36、37、38、39、40、41或42℃，和其中的任何值或范围)的温度下。在一些实施方案中，具有经改善的热稳定性的逆转录酶包括但不限于：通过参考SEQ ID NO:172的氨基酸位置编号而言的M-MuLV三突变体D200N+L603W+T330P或M-MuLV五突变体(5M)D200N+L603W+T330P+T306K+W313F(例如，SEQID NO:53)。参见例如，Baranauskas等人(Protein Eng.Des.Sel.25,657-668(2012))；Anzalone等人(Nature 576:149-157(2019))。在逆转录酶中的另外的氨基酸修饰可以包括通过参考SEQ ID NO:172的氨基酸位置编号而言的L139P、D200N、W388R、E607K、T306K、W313F、F155Y、H638G、Q221R、V223M和/或D524N的氨基酸置换。
在一些实施方案中，对本发明来说有用的逆转录酶可以包括但不限于下列氨基酸置换的组合：(1)L139P、D200N、W388R和E607K；(2)L139P、D200N、T306K、W313F、W388R和E607K；(3)5M(T355A/Q357M/K358R/A359G/S360A)、F155Y和H638G；(4)5M(T355A/Q357M/K358R/A359G/S360A)、Q221R和V223M；或(5)5M(T355A/Q357M/K358R/A359G/S360A)和D524N，通过参考SEQ ID NO:172的氨基酸位置编号而言。
在本发明的一些实施方案中，逆转录酶可以融合至一个或多个单链RNA结合结构域(RBD)。对本发明来说有用的RBD可以包括但不限于：SEQ ID NO:37-52(SEQ ID NO:37、SEQ ID NO:38、SEQ ID NO:39、SEQ ID NO:40、SEQ ID NO:41、SEQ ID NO:42、SEQ ID NO:43、SEQ ID NO:44、SEQ ID NO:45、SEQ ID NO:46、SEQ ID NO:47、SEQ ID NO:48、SEQ IDNO:49、SEQ ID NO:50、SEQ ID NO:51和/或SEQ ID NO:52)，由此改善所述逆转录酶的热稳定性、持续能力和模板亲和力。
本发明的多肽/蛋白质/结构域(例如，CRISPR-Cas效应蛋白，例如II型或V型CRISPR-Cas效应蛋白)、逆转录酶、5'翼片核酸内切酶和/或5'-3'核酸外切酶)可以由一个或多个多核苷酸来编码，所述多核苷酸任选地可操作地连接至一个或多个启动子和/或其他调控序列(例如，终止子、操纵子和/或增强子等)。在一些实施方案中，本发明的多核苷酸可以被包含在一个或多个表达盒和/或载体中。在一些实施方案中，所述至少一个调控序列可以为例如启动子、操纵子、终止子或增强子。在一些实施方案中，所述至少一个调控序列可以为启动子。在一些实施方案中，所述调控序列可以为内含子。在一些实施方案中，所述至少一个调控序列可以为例如可操作地与内含子相关联的启动子或者包含内含子的启动子区。在一些实施方案中，所述至少一个调控序列可以为例如泛素启动子和其相关联的内含子(例如，蒺藜苜蓿(Medicago truncatula)启动子和/或玉蜀黍及其相关联的内含子)(例如，包含内含子的ZmUbi1；包含内含子的MtUb2，例如SEQ ID NO:21或22)。
在一些实施方案中，本发明提供了编码II型CRISPR-Cas效应蛋白或结构域或者V型CRISPR-Cas效应蛋白或结构域的多核苷酸、编码CRISPR-Cas效应蛋白或结构域的多核苷酸、编码逆转录酶多肽或结构域的多核苷酸、编码5'-3'核酸外切酶多肽或结构域的多核苷酸和/或编码翼片核酸内切酶多肽或结构域的多核苷酸，其可操作地与一个或多个包含内含子或与内含子相关联的启动子区相关联，任选地其中所述启动子区可以为泛素启动子和内含子(例如，苜蓿或玉蜀黍泛素启动子和内含子，例如SEQ ID NO:21或22)。
在一些实施方案中，编码II型或V型CRISPR-Cas效应蛋白的多核苷酸和/或编码逆转录酶的多核苷酸可以被包含在相同或分开的表达盒中，任选地当编码所述II型或V型CRISPR-Cas效应蛋白的多核苷酸和编码所述逆转录酶的多核苷酸被包含在相同的表达盒中时，编码所述II型或V型CRISPR-Cas效应蛋白的多核苷酸和编码所述逆转录酶的多核苷酸可以可操作地连接至单一启动子或者以任何组合可操作地连接至两个或更多个分开的启动子。在一些实施方案中，编码CRISPR-Cas效应蛋白的多核苷酸可以被包含在表达盒中，其中编码所述CRISPR-Cas效应蛋白的多核苷酸可以可操作地连接至启动子。
在一些实施方案中，延伸型指导核酸和/或指导核酸可以被包含在表达盒中，任选地其中所述表达盒被包含在载体中。在一些实施方案中，包含所述延伸型指导核酸的表达盒和/或载体可以为与包含编码所述II型或V型CRISPR-Cas效应蛋白的多核苷酸和/或编码所述逆转录酶的多核苷酸的那种相同或不同的表达盒和/或载体。在一些实施方案中，包含所述指导核酸的表达盒和/或载体可以为与包含编码所述CRISPR-Cas效应蛋白的多核苷酸的那种相同或不同的表达盒和/或载体。
在一些实施方案中，编码5'翼片核酸内切酶的多核苷酸和/或编码5'-3'核酸外切酶的多核苷酸可以被包含在一个或多个表达盒中，所述一个或多个表达盒可以为相同或不同的表达盒。在一些实施方案中，包含编码5'翼片核酸内切酶的多核苷酸和/或编码5'-3'核酸外切酶的多核苷酸的表达盒可以为与包含编码II型或V型CRISPR-Cas效应蛋白的多核苷酸、编码II型或V型CRISPR-Cas效应蛋白的多核苷酸和/或编码逆转录酶的多核苷酸的那种相同或不同的表达盒。
在本发明的一些实施方案中，可以对编码CRISPR-Cas效应蛋白(例如，II型CRISPR-Cas效应蛋白、V型CRISPR-Cas效应蛋白)、逆转录酶、翼片核酸内切酶、5'-3'核酸外切酶和包含其的融合蛋白的多核苷酸以及包含所述多核苷酸的核酸构建体、表达盒和/或载体进行密码子优化以用于在生物(例如，动物(例如，哺乳动物、昆虫、鱼类等)、植物(例如，双子叶植物、单子叶植物)、细菌、古细菌、病毒等)中进行表达。在一些实施方案中，可以对所述多核苷酸、表达盒和/或载体进行密码子优化以用于在植物(任选地双子叶植物或单子叶植物)中进行表达。对于其来说本发明可以是有用的示例性哺乳动物包括但不限于：灵长类(人和非人(例如，黑猩猩、狒狒、猴子、大猩猩等))、猫、狗、雪貂、沙鼠、仓鼠、牛、猪、马、山羊、驴或绵羊。在一些实施方案中，可以对所述多核苷酸、表达盒和/或载体进行密码子优化以用于在真菌(包括但不限于接合菌门(Zygomycota)、子囊菌门(Ascomycota)、担子菌门(Basidiomycota)和半知菌门(Deuteromycota)(不完全真菌))中进行表达，任选地其中所述真菌可以为子囊菌，任选地酵母(例如，酿酒糖酵母(Saccharomyces cerevisiae))。
在一些实施方案中，为在生物中进行表达而进行优化的本发明的多核苷酸、核酸构建体、表达盒或载体可以是与编码其但未就在植物中进行表达而进行密码子优化的核酸构建体、表达盒或载体大约70％至100％(例如，大约70％、71％、72％、73％、74％、75％、76％、77％、78％、79％、80％、81％、82％、83％、84％、85％、86％、87％、88％、89％、90％、91％、92％、93％、94％、95％、96％、97％、98％、99％、99.5％或100％)同一的。
在一些实施方案中，可以提供多核苷酸、核酸构建体、表达盒和载体用于实施本发明的方法。因此，在一些实施方案中，提供了经密码子优化以用于在生物中进行表达的表达盒，其以5'至3'包含：(a)编码启动子序列的多核苷酸；(b)编码V型CRISPR-Cas核酸酶(例如，Cpf1(Cas12a)、dCas12a等)或II型CRISPR-Cas核酸酶(例如，Cas9、dCas9等)的多核苷酸，其进行了密码子优化以用于在所述生物中进行表达；(c)连接体序列；和(d)编码逆转录酶的多核苷酸，其进行了密码子优化以用于在所述生物中进行表达。在一些实施方案中，所述生物为动物、植物、真菌、古细菌或细菌。在一些实施方案中，所述生物为植物，并且所述编码V型CRISPR-Cas核酸酶的多核苷酸进行了密码子优化以用于在植物中进行表达，并且所述启动子序列为植物特异性启动子序列(例如，ZmUbi1、MtUb2、RNA聚合酶II(Pol II))。
在一些实施方案中，可以提供多核苷酸、核酸构建体、表达盒和载体用于实施本发明的方法。因此，在一些实施方案中，提供了经密码子优化以用于在植物中进行表达的表达盒，其以5'至3'包含：(a)编码植物特异性启动子序列(例如，ZmUbi1、MtUb2、RNA聚合酶II(Pol II))的多核苷酸；(b)经植物密码子优化的编码II型或V型CRISPR-Cas效应蛋白(例如，Cpf1(Cas12a)、dCas12a等)的多核苷酸；(c)连接体序列；和(d)经植物密码子优化的编码逆转录酶的多核苷酸。
在一些实施方案中，本发明的多肽可以为包含一个或多个经由连接体相互连接的多肽的融合蛋白。在一些实施方案中，所述连接体可以为氨基酸或肽连接体。在一些实施方案中，肽连接体的长度可以为大约2至大约100个氨基酸(残基)，如在本文中所描述的。在一些实施方案中，肽连接体可以为例如GS连接体。
在一些实施方案中，本发明提供了经密码子优化以用于在植物中进行表达的表达盒，其包含：(a)编码植物特异性启动子序列(例如，ZmUbi1、MtUb2)的多核苷酸，和(b)延伸型指导核酸序列，其中所述延伸型指导核酸包含延伸部分，所述延伸部分包含在其3'末端处的引物结合位点和待被掺入到所述靶核酸中的编辑(例如，在逆转录酶模板中的编辑)(例如，5'-3'–crRNA-RTT-PBS)(例如，tag核酸；例如，tagRNA)，任选地其中所述延伸型指导核酸被包含在表达盒中，任选地其中所述延伸型指导核酸可操作地连接至Pol II启动子。在一些实施方案中，当所述指导核酸的延伸部分在所述crRNA的5'末端处附接至CRISPRRNA时，所述延伸部分包含在其5'末端处的引物结合位点和在3'末端处的待被掺入到所述靶核酸中的编辑(例如，逆转录酶模板)(5'-3'–PBS-RTT-crRNA)。
在一些实施方案中，本发明的表达盒可以进行密码子优化以用于在双子叶植物中或在单子叶植物中进行表达。在一些实施方案中，本发明的表达盒可以于在植物或植物细胞中修饰靶核酸的方法之中进行使用，所述方法包括将一个或多个本发明的表达盒引入到植物或植物细胞中，由此在所述植物或植物细胞中修饰所述靶核酸以产生包含所述经修饰的靶核酸的植物或植物细胞。在一些实施方案中，所述方法可以进一步包括使包含所述经修饰的靶核酸的植物细胞进行再生，以产生包含所述经修饰的靶核酸的植物。
在一些实施方案中，本发明的表达盒可以进行密码子优化以用于在动物(例如，哺乳动物)中进行表达。在一些实施方案中，本发明的表达盒可以于在动物细胞(例如，哺乳动物细胞)中修饰靶核酸的方法之中进行使用，所述方法包括将一个或多个本发明的表达盒引入到动物细胞中，由此在所述动物细胞中修饰所述靶核酸以产生包含所述经修饰的靶核酸的动物细胞。
对本发明来说有用的CRISPR Cas9多肽或CRISPR Cas9结构域(例如，II型CRISPRCase效应蛋白)可以为任何已知的或后来鉴定出的Cas9核酸酶。在一些实施方案中，CRISPRCas9多肽可以为来自例如链球菌属物种(Streptococcus spp.)(例如，酿脓链球菌、嗜热链球菌(S.thermophilus))(例如，spCas9)、乳杆菌属物种(Lactobacillus spp.)、双歧杆菌属物种(Bifidobacterium spp.)、坎德勒氏菌属物种(Kandleria spp.)、明串珠菌属物种(Leuconostoc spp.)、酒球菌属物种(Oenococcus spp.)、片球菌属物种(Pediococcusspp.)、魏斯氏菌属物种(Weissella spp.)和/或奥尔森氏菌属物种(Olsenella spp.)的Cas9多肽。
Cas12a为V型成簇规律间隔短回文重复序列(Clustered Regularly InterspacedShort Palindromic Repeats；CRISPR)-Cas效应蛋白或结构域。Cas12a在几个方面不同于更熟知的II型CRISPR Cas9效应蛋白。例如，Cas9识别处于其指导RNA(gRNA，sgRNA)结合位点(原间隔子(protospacer)，靶核酸，靶DNA)3'处的富含G的原间隔子邻近基序(PAM)(3'-NGG)，而Cas12a识别位于靶核酸5'处的富含T的PAM(5'-TTN、5'TTTN)。事实上，Cas9和Cas12a结合其指导RNA所采取的定向相关于其N和C末端而言是非常几乎相反的。进一步地，Cas12a效应蛋白使用单一指导RNA(gRNA，CRISPR阵列，crRNA)，而不是在天然Cas9系统中发现的双重指导RNA(sgRNA(例如，crRNA和tracrRNA))，并且Cas12a加工其自己的gRNA。另外，Cas12a的核酸酶活性产生错开的DNA双链断裂，而不是由Cas9的核酸酶活性产生的平端，并且Cas12a依赖于单个RuvC结构域来切割两条DNA链，而Cas9利用HNH结构域和RuvC结构域来进行切割。
对本发明来说有用的CRISPR Cas12a效应蛋白或结构域可以为任何已知的或后来鉴定出的Cas12a核酸酶(以前称为Cpf1)(参见例如，美国专利号9,790,490，其通过提及而合并，为了其关于Cpf1(Cas12a)序列的公开内容)。术语“Cas12a”、“Cas12a多肽”或“Cas12a结构域”是指包含Cas12a或其片段(其包含Cas12a的指导核酸结合结构域，和/或Cas12a的有活性的、无活性的或部分有活性的DNA切割结构域)的RNA-指导的效应蛋白。在一些实施方案中，对本发明来说有用的Cas12a可以在核酸酶活性位点(例如，Cas12a结构域的RuvC位点)中包含突变。在其核酸酶活性位点中具有突变并且因此不再包含核酸酶活性的Cas12a效应蛋白或结构域通常被称为死的或失活的Cas12a(例如，dCas12a)。
在一些实施方案中，可以按照本发明进行优化或修饰(例如，失活)的Cas12a效应多肽可以包括但不限于SEQ ID NO:1-20(例如，SEQ ID NO:1、2、3、4、5、6、7、8、9、10、11、12、13、14、15、16、17、18、19或20)或者SEQ ID NO:148、149、150或151中的任一个的氨基酸序列，或者编码其的多核苷酸。
在一些实施方案中，可以按照本发明进行优化或修饰(例如，灭活)的Cas9效应多肽可以包括但不限于SEQ ID NO:106或SEQ ID NO:107中的任一个的氨基酸序列，或者编码其的多核苷酸。在一些实施方案中，可以按照本发明进行优化或修饰(例如，灭活)的Cas9效应多肽可以包含由SEQ ID NO:108-122中的任一个的核酸序列所编码的氨基酸序列。
如在本文中所使用的，“指导核酸”、“指导RNA”、“gRNA”、“CRISPR RNA/DNA”、“crRNA”或“crDNA”意指这样的核酸，其包含至少一个与靶DNA(例如，原间隔子)互补(并且与之杂交)的间隔子序列，和至少一个相应于特定的CRISPR-Cas效应蛋白的重复序列(例如，对于V型CRISPR Cas效应蛋白，所述重复序列或其片段或部分来自V型Cas12a CRISPR-Cas系统；对于II型CRISPR Cas效应蛋白，所述重复序列或其片段或部分来自II型Cas9CRISPR-Cas系统)。因此，对本发明来说有用的CRISPR-Cas系统的重复序列可以相应于例如Cas9、C2c3、Cas12a(也称为Cpf1)、Cas12b、Cas12c、Cas12d、Cas12e、Cas13a、Cas13b、Cas13c、Cas13d、Casl、CaslB、Cas2、Cas3、Cas3'、Cas3”、Cas4、Cas5、Cas6、Cas7、Cas8、Cas9(也称为Csnl和Csx12)、Cas10、Csyl、Csy2、Csy3、Csel、Cse2、Cscl、Csc2、Csa5、Csn2、Csm2、Csm3、Csm4、Csm5、Csm6、Cmrl、Cmr3、Cmr4、Cmr5、Cmr6、Csbl、Csb2、Csb3、Csxl7、Csxl4、Csx10、Csx16、CsaX、Csx3、Csxl、Csxl5、Csfl、Csf2、Csf3、Csf4(dinG)和/或Csf5的CRISPR-Cas效应蛋白或其片段，其中所述重复序列可以连接至所述间隔子序列的5'末端和/或3'末端。本发明的指导核酸的设计可以基于I型、II型、III型、IV或V型CRISPR-Cas系统。在一些实施方案中，本发明的指导核酸的设计基于V型CRISPR-Cas系统。在一些实施方案中，本发明的指导核酸的设计基于II型CRISPR-Cas系统。
在一些实施方案中，指导核酸(例如，crRNA，例如Cas12a crRNA、Cas12b crRNA、Cas9 crRNA等)可以包含(从5'至3')：重复序列(全长或其部分(“柄”)；例如，假结样结构)；和间隔子序列。在一些实施方案中，延伸型指导核酸(例如，tagRNA，例如Cas12a延伸型指导核酸、Cas12b延伸型指导核酸、Cas9延伸型指导核酸等)可以包含(从5'至3')：重复序列(全长或其部分(“柄”)；例如，假结样结构)，间隔子序列，加上包含引物结合位点和逆转录酶模板(RT模板)(RTT)的3'或5'延伸部分(例如，tagRNA延伸)。
在一些实施方案中，指导核酸可以包含多于一个重复序列-间隔子序列(例如，2、3、4、5、6、7、8、9、10或更多个重复序列-间隔子序列)(例如，重复-间隔子-重复；例如，重复-间隔子-重复-间隔子-重复-间隔子-重复-间隔子-重复-间隔子，等等)。本发明的指导核酸是合成的、人造的并且未在自然界中发现。指导核酸可以是相当长的，并且可以用作适配体(像在MS2募集策略中那样)或者其他从间隔子上悬挂的RNA结构。在一些实施方案中，如在本文中所描述的，指导核酸可以包括用于编辑的模板和引物结合位点。在一些实施方案中，指导核酸可以在其5'末端或3'末端处包含与编辑模板(逆转录酶模板)互补的区域或序列，由此将所述编辑模板募集至所述靶核酸(即，延伸型指导核酸)。在一些实施方案中，指导核酸可以在其5'末端或3'末端处包含与在所述靶核酸上的引物互补的区域或序列(引物结合位点)，由此将所述引物结合位点募集至所述靶核酸(即，延伸型指导核酸)。
如在本文中所使用的，“重复序列”是指例如，野生型CRISPR Cas基因座(例如，Cas9基因座、Cas12a基因座、C2c1基因座等)的任何重复序列，或者与由本发明的核酸构建体所编码的CRISPR-Cas核酸酶一起有功能的合成的crRNA的重复序列。对本发明来说有用的重复序列可以为任何已知的或后来鉴定出的CRISPR-Cas基因座(例如，I型、II型、III型、IV型、V型或VI型)的重复序列，或者它可以为被设计成在I、II、III、IV、V或VI型CRISPR-Cas系统中发挥功能的合成的重复序列。因此，在一些实施方案中，重复序列可以与来自野生型I型CRISPR-Cas基因座、II型CRISPR-Cas基因座、III型CRISPR-Cas基因座、IV型CRISPR-Cas基因座、V型CRISPR-Cas基因座和/或VI型CRISPR-Cas基因座的重复序列相同或实质上相同。在一些实施方案中，对本发明来说有用的重复序列可以为任何已知的或后来鉴定出的V型CRISPR-Cas基因座的重复序列，或者它可以为被设计成在V型CRISPR-Cas系统中发挥功能的合成的重复序列。重复序列可以包含发夹结构和/或茎-环结构。在一些实施方案中，重复序列可以在其5'末端处形成假结样结构(即，“柄”)。因此，在一些实施方案中，重复序列可以与来自野生型V型CRISPR-Cas基因座或野生型II型CRISPR-Cas基因座的重复序列同一或实质上同一。来自野生型CRISPR-Cas基因座的重复序列可以通过已建立的算法来确定，例如使用通过CRISPRdb提供的CRISPRfinder(参见Grissa等人,Nucleic Acids Res.35(Web Server issue):W52-7；或BMC Informatics 8:172(2007)(doi:10.1186/1471-2105-8-172))。在一些实施方案中，重复序列或其部分在其3'末端处连接至间隔子序列的5'末端，从而形成重复序列-间隔子序列(例如，指导RNA、crRNA)。
在一些实施方案中，重复序列包含至少10个核苷酸、基本上由至少10个核苷酸组成或者由至少10个核苷酸组成，取决于特定的重复序列以及包含该重复序列的指导RNA是经加工的还是未加工的(例如，大约10、11、12、13、14、15、16、17、18、19、20、21、22、23、24、25、26、27、28、29、30、31、32、33、34、35、36、37、38、39、40、41、42、43、44、45、46、47、48、49、50至100个或更多个核苷酸，或者其中的任何范围或值；例如，大约)。在一些实施方案中，重复序列包含下述核苷酸、基本上由下述核苷酸组成或者由下述核苷酸组成：大约10至大约20、大约10至大约30、大约10至大约45、大约10至大约50、大约15至大约30、大约15至大约40、大约15至大约45、大约15至大约50、大约20至大约30、大约20至大约40、大约20至大约50、大约30至大约40、大约40至大约80、大约50至大约100或更多个核苷酸。
与间隔子序列的5'末端相连接的重复序列可以包含重复序列的一部分(例如，野生型重复序列的5、6、7、8、9、10、11、12、13、14、15、16、17、18、19、20、21、22、23、24、25、26、27、28、29、30、31、32、33、34、35或更多个邻接核苷酸)。在一些实施方案中，与间隔子序列的5'末端相连接的重复序列的一部分的长度可以为大约五个至大约十个连续核苷酸(例如，大约5、6、7、8、9、10个核苷酸)，并且与野生型CRISPR Cas重复核苷酸序列的相同区域(例如，5'末端)具有至少90％同一性(例如，至少大约90％、91％、92％、93％、94％、95％、96％、97％、98％、99％或更多)。在一些实施方案中，重复序列的一部分可以在其5'末端处包含假结样结构(例如，“柄”)。
如在本文中所使用的，“间隔子序列”为与靶核酸(例如，靶DNA)(例如，原间隔子)互补的核苷酸序列。所述间隔子序列可以与靶核酸完全互补或实质上互补(例如，至少大约70％互补(例如，大约70％、71％、72％、73％、74％、75％、76％、77％、78％、79％、80％、81％、82％、83％、84％、85％、86％、87％、88％、89％、90％、91％、92％、93％、94％、95％、96％、97％、98％、99％、99.5％、99.6％、99.7％、99.8％、99.9％或更多))。因此，在一些实施方案中，所述间隔子序列可以相比于所述靶核酸而言具有一个、两个、三个、四个或五个错配，所述错配可以是邻接的或非邻接的。在一些实施方案中，所述间隔子序列可以与靶核酸具有70％互补性。在一些实施方案中，所述间隔子核苷酸序列可以与靶核酸具有80％互补性。在另外其他实施方案中，所述间隔子核苷酸序列可以与所述靶核酸(原间隔子)具有85％、90％、91％、92％、93％、94％、95％、96％、97％、98％、99％、99.5％、99.6％、99.7％、99.8％、99.9％或更多互补性，等等。在一些实施方案中，所述间隔子序列与所述靶核酸100％互补。在一些实施方案中，间隔子序列可以具有大约15个核苷酸至大约30个核苷酸(例如，15、16、17、18、19、20、21、22、23、24、25、26、27、28、29或30个核苷酸，或者其中的任何范围或值)的长度。因此，在一些实施方案中，间隔子序列可以在长度为至少大约15个核苷酸至大约30个核苷酸的靶核酸(例如，原间隔子)的区域上具有完全的互补性或实质上的互补性。在一些实施方案中，所述间隔子的长度为大约20个核苷酸。在一些实施方案中，所述间隔子的长度为大约23个核苷酸。
在一些实施方案中，指导RNA的间隔子序列的5'区可以与靶DNA同一，而所述间隔子的3'区可以与所述靶DNA(例如，V型CRISPR-Cas)实质上互补，或者指导RNA的间隔子序列的3'区可以与靶DNA同一，而所述间隔子的5'区可以与所述靶DNA(例如，II型CRISPR-Cas)实质上互补，并且因此所述间隔子序列与所述靶DNA的总互补性可以小于100％。因此，例如，在用于V型CRISPR-Cas系统的指导中，在例如具有20个核苷酸的间隔子序列的5'区(即，种子区)中的前1、2、3、4、5、6、7、8、9、10个核苷酸可以与所述靶DNA 100％互补，而在所述间隔子序列的3'区中的其余核苷酸与所述靶DNA实质上互补(例如，至少大约70％互补)。在一些实施方案中，所述间隔子序列的5'末端的前1至8个核苷酸(例如，前1、2、3、4、5、6、7、8个核苷酸，以及其中的任何范围)可以与所述靶DNA 100％互补，而在所述间隔子序列的3'区中的其余核苷酸与所述靶DNA实质上互补(例如，至少大约50％互补(例如，50％、55％、60％、65％、70％、71％、72％、73％、74％、75％、76％、77％、78％、79％、80％、81％、82％、83％、84％、85％、86％、87％、88％、89％、90％、91％、92％、93％、94％、95％、96％、97％、98％、99％、99.5％、99.6％、99.7％、99.8％、99.9％或更多)。
作为一个进一步的例子，在用于II型CRISPR-Cas系统的指导中，在例如具有20个核苷酸的间隔子序列的3'区(即，种子区)中的前1、2、3、4、5、6、7、8、9、10个核苷酸可以与所述靶DNA 100％互补，而在所述间隔子序列的5'区中的其余核苷酸与所述靶DNA实质上互补(例如，至少大约70％互补)。在一些实施方案中，所述间隔子序列的3'末端的前1至10个核苷酸(例如，前1、2、3、4、5、6、7、8、9、10个核苷酸，以及其中的任何范围)可以与所述靶DNA100％互补，而在所述间隔子序列的5'区中的其余核苷酸与所述靶DNA实质上互补(例如，至少大约50％互补(例如，至少大约50％、55％、60％、65％、70％、71％、72％、73％、74％、75％、76％、77％、78％、79％、80％、81％、82％、83％、84％、85％、86％、87％、88％、89％、90％、91％、92％、93％、94％、95％、96％、97％、98％、99％、99.5％、99.6％、99.7％、99.8％、99.9％或更多，或者其中的任何范围或值)。
在一些实施方案中，间隔子的种子区的长度可以为大约8至大约10个核苷酸，长度为大约5至大约6个核苷酸，或者长度为大约6个核苷酸。
在一些实施方案中，本发明的延伸型指导核酸可以为延伸型指导核酸、第一延伸型指导核酸和/或第二延伸型指导核酸。在一些实施方案中，对本发明来说有用的延伸型指导核酸可以包含：(a)CRISPR核酸(例如，CRISPR RNA、CRISPR DNA、crRNA、crDNA)和/或CRISPR核酸和tracr核酸；和(b)包含引物结合位点和逆转录酶模板(RT模板)的延伸部分，其中所述RT模板编码待被掺入到所述靶核酸中的修饰(如在本文中所描述的)(例如，编码位于RT模板之内的任何位置中的编辑，其具有相对于所述靶核酸的原间隔子邻近基序(PAM)的位置而言的位置定位，任选地位于核苷酸位置-1至核苷酸位置19，核苷酸位置10至核苷酸位置17，或核苷酸位置12至核苷酸位置15处的编辑)。在一些实施方案中，CRISPR核酸可以为II型或V型CRISPR核酸，和/或tracr核酸可以为任何相应于合适的II型或V型CRISPR核酸的tracr。延伸型指导核酸也可以称为所靶向的等位基因指导核酸、所靶向的等位基因指导DNA、所靶向的等位基因指导RNA(tagRNA)。在一些实施方案中，对本发明来说有用的CRISPR核酸可以为V型CRISPR核酸。在一些实施方案中，对本发明来说有用的tracr核酸可以为V型CRISPR tracr核酸。在一些实施方案中，对本发明来说有用的CRISPR核酸可以为II型CRISPR核酸。在一些实施方案中，对本发明来说有用的tracr核酸可以为II型CRISPRtracr核酸。在一些实施方案中，CRISPR核酸和/或tracr核酸可以来自例如Cas9、C2c3、Cas12a(也称为Cpf1)、Cas12b、Cas12c、Cas12d、Cas12e、Cas13a、Cas13b、Cas13c、Cas13d、Casl、CaslB、Cas2、Cas3、Cas3'、Cas3”、Cas4、Cas5、Cas6、Cas7、Cas8、Cas9(也称为Csnl和Csx12)、Cas10、Csyl、Csy2、Csy3、Csel、Cse2、Cscl、Csc2、Csa5、Csn2、Csm2、Csm3、Csm4、Csm5、Csm6、Cmrl、Cmr3、Cmr4、Cmr5、Cmr6、Csbl、Csb2、Csb3、Csxl7、Csxl4、Csx10、Csx16、CsaX、Csx3、Csxl、Csxl5、Csfl、Csf2、Csf3、Csf4(dinG)和/或Csf5系统。
在一些实施方案中，所述延伸型指导的延伸部分可以以5'至3'包含RT模板和引物结合位点(当所述延伸型指导连接至所述CRISPR核酸的3'末端时)。在一些实施方案中，所述延伸型指导的延伸部分可以以5'至3'包含引物结合位点和RT模板(RTT)(当所述延伸型指导连接至所述CRISPR核酸的5'末端时)。在一些实施方案中，RT模板的长度可以为大约1个核苷酸至大约100个核苷酸(例如，大约1、2、3、4、5、6、7、8、9、10、11、12、13、14、15、16、17、18、19、20、21、22、23、24、25、26、27、28、29、30、31、32、33、34、35、36、37、38、39、40、41、42、43、44、45、46、47、48、49、50、51、52、53、54、55、56、57、58、59、60、61、62、63、64、65、66、67、68、69、70、71、72、73、74、75、76、77、78、79、80、81、82、83、84、85、86、87、88、89、90、91、92、93、94、95、96、97、98、99、100或更多个核苷酸，和其中的任何范围或值)，例如，长度为大约35个核苷酸至大约100个核苷酸、大约35个核苷酸至大约80个核苷酸、大约35个核苷酸至大约75个核苷酸、大约40个核苷酸至大约75个核苷酸、大约45个核苷酸至大约75个核苷酸、大约45个核苷酸至大约60个核苷酸和其中任何范围或值。在一些实施方案中，RT模板的长度可以为至少30个核苷酸，任选地长度为大约30、31、32、33、34、35、36、37、38、39或40个核苷酸至大约41、42、43、44、45、46、47、48、49、50、51、52、53、54、55、56、57、58、59、60、61、62、63、64、65、66、67、68、69、70、71、72、73、74、75、76、77、78、79或80个核苷酸，或者其中的任何范围或值。在一些实施方案中，RT模板的长度可以为大约8、36、40、44、47、50、52、55、63、72或74个核苷酸。在所述RTT的长度之内包含编辑。所述编辑可以位于所述RTT之内的任何地方，其中所述编辑的位置可以相对于所述靶核酸的原间隔子邻近基序(PAM)的位置而言进行描述。在一些实施方案中，RT模板可以包含位于核酸位置-1、1、2、3、4、5、6、7、8、9、10、11、12、13、14、15、16、27、18或19处的编辑。在一些实施方案中，RT模板可以包含位于所述RT模板的核苷酸位置4至核苷酸位置17(例如，位置4、5、6、7、8、9、10、11、12、13、14、15、16或17)处的编辑，相对于所述靶核酸的原间隔子邻近基序(PAM)的位置而言。在一些实施方案中，RT模板可以包含位于所述RT模板的核苷酸位置10至核苷酸位置17(例如，位置10、11、12、13、14、15、16或17)处的编辑，相对于所述靶核酸的原间隔子邻近基序(PAM)的位置而言。在一些实施方案中，RT模板可以包含位于所述RT模板的核苷酸位置12至核苷酸位置15(例如，位置12、13、14或15)处的编辑，相对于所述靶核酸的原间隔子邻近基序(PAM)的位置而言。
如在本文中所使用的，延伸型指导核酸(例如，tagRNA)的延伸部分的“引物结合位点(PBS)”是指可以与在靶核酸上的区域或“引物”相结合(即与所述靶核酸引物互补)的连续核苷酸的序列。作为例子，CRISPR Cas效应蛋白(例如，II型或V型，例如Cas9或Cas12a)对于所述DNA产生切口/进行切割，经切割的DNA的3'末端充当用于所述延伸型指导核酸的PBS部分的引物。所述PBS被设计成与所述靶核酸的链的3'末端互补，并且可以被设计成与靶标链或非靶标链相结合。引物结合位点可以与所述引物完全互补，或者它可以与在所述靶核酸上的引物实质上互补(例如，至少70％互补(例如，大约70％、71％、72％、73％、74％、75％、76％、77％、78％、79％、80％、81％、82％、83％、84％、85％、86％、87％、88％、89％、90％、91％、92％、93％、94％、95％、96％、97％、98％、99％、99.5％、99.6％、99.7％、99.8％、99.9％或更多))。在一些实施方案中，延伸部分的引物结合位点的长度可以为大约1个核苷酸至大约100个核苷酸(例如，大约1、2、3、4、5、6、7、8、9、10、11、12、13、14、15、16、17、18、19、20、21、22、23、24、25、26、27、28、29、30、31、32、33、34、35、36、37、38、39、40、41、42、43、44、45、46、47、48、49、50、51、52、53、54、55、56、57、58、59、60、61、62、63、64、65、66、67、68、69、70、71、72、73、74、75、76、77、78、79、80、81、82、83、84、85、86、87、88、89、90、91、92、93、94、95、96、97、98、99、100或更多个核苷酸，或者其中的任何值或范围)，或者大约4个核苷酸至大约85个核苷酸、大约10个核苷酸至大约80个核苷酸、大约20个核苷酸至大约80个寡核苷酸、大约25个核苷酸至大约80个核苷酸、大约30个核苷酸至大约80个核苷酸、大约40个核苷酸至大约80个核苷酸、大约45个核苷酸至大约80个核苷酸、大约45个核苷酸至大约75个核苷酸或大约45个核苷酸至大约60个核苷酸，或者其中的任何范围或值。在一些实施方案中，PBS的长度可以为至少30个核苷酸，任选地大约30、31、32、33、34、35、36、37、38、39或40个核苷酸至大约41、42、43、44、45、46、47、48、49、50、51、52、53、54、55、56、57、58、59、60、61、62、63、64、65、66、67、68、69、70、71、72、73、74、75、76、77、78、79或80个核苷酸，或者其中的任何范围或值。在一些实施方案中，PBS的长度可以为大约8、16、24、32、40、48、56、64、72或80个核苷酸。
在一些实施方案中，RTT可以具有大约35个核苷酸至大约75个核苷酸的长度，并且PBS可以具有大约30个核苷酸至大约80个核苷酸的长度，任选地其中所述PBS可以包含大约8、16、24、32、40、48、56、64、72或80个核苷酸的长度，并且所述RTT可以包含大约36、40、44、47、50、52、55、63、72或74个核苷酸的长度，或者RTT长度和/或PBS长度的其任何组合。
在一些实施方案中，延伸型指导核酸(例如，延伸型指导核酸，第一延伸型指导核酸，第二延伸型指导核酸)可以包含结构化RNA基序，任选地其中所述结构化RNA基序可以位于所述延伸型指导核酸的3'末端处。在一些实施方案中，所述结构化RNA基序可以包括但不限于：AsCpf1BB(SEQ ID NO:189)、BoxB(SEQ ID NO:190)、假结(诱饵)(SEQ ID NO:95、SEQID NO:203)、假结(tEvoPreQ1)(SEQ ID NO:191)、fmp结(SEQ ID NO:192)、mp结(SEQ IDNO:193)、MS2(SEQ ID NO:194)、PP7(SEQ ID NO:195)、SLBP(SEQ ID NO:196)、TAR(SEQ IDNO:197)和/或ThermoPh(SEQ ID NO:198)。在一些实施方案中，结构化RNA基序可以为假结，任选地其中所述假结位于所述延伸型指导核酸的3'末端处。假结为在发夹的环中的RNA的单链区域与在RNA链中别处的互补核苷酸链段进行碱基配对后形成的RNA结构基序。在一些实施方案中，对本发明来说有用的假结可以为天然出现的假结或合成的假结。如在本文中所使用的，术语“假结”包括但不限于发夹、多环、相吻环、共轴堆叠、三链体、假结样结构、假结化发夹和/或诱饵假结化发夹或其他RNA结构基序。在一些实施方案中，所述假结可以位于所述延伸型指导核酸的3'末端处。在一些实施方案中，当所述延伸型指导包含5'-3'crRNA-RTT-PBS时，假结可以位于所述RTT的5'处或所述PBS的3'处。在一些实施方案中，所述假结可以位于所述延伸型指导核酸的3'末端处。在一些实施方案中，当所述延伸型指导在crRNA的5'末端处包含延伸(延伸部分)时，假结可以位于所述RTT的3'处或所述PBS的5'处。在一些实施方案中，对延伸型指导来说有用的假结可以包括但不限于：包含UAAUUUCUACUAAGUGUAGAU(SEQ ID NO:158)的核酸序列的tEvoPreQ1假结，包含TTGACGCGGTTCTATCTAGTTACGCGTTAAACCAACUAGAAA(SEQ ID NO:191)的核酸序列的假结EvoPreQ1，或包含TAAGTCTCCATAGAATGGAGG(SEQ ID NO:95)和/或UAAGUCUCCAUAGAAUGGAGG(SEQ ID NO:203)的核酸序列的假结。本发明的延伸型指导核酸可以被包含在表达盒中，任选地其中所述表达盒被包含在载体中。
在一些实施方案中，延伸型指导的延伸部分可以融合至II型或V型CRISPR核酸的5'末端或3'末端(例如，5'至3'：重复-间隔子-延伸部分，或延伸部分-重复-间隔子)，和/或融合至tracr核酸的5'或3'末端。在一些实施方案中，当延伸部分位于所述crRNA的5'时，修饰所述V型CRISPR-Cas效应蛋白以降低(或消除)自加工RNA酶活性。在一些实施方案中，当延伸部分位于所述crRNA的3'时，也可以利用进行了修饰以降低(或消除)自加工RNA酶活性的V型CRISPR-Cas效应蛋白。
在一些实施方案中，延伸型指导核酸的延伸部分可以经由连接体连接至所述II型或V型CRISPR核酸和/或所述II型或V型tracrRNA。在一些实施方案中，连接体的长度可以为大约1至大约100个核苷酸或更多个(例如，长度为大约1、2、3、4、5、6、7、8、9、10、11、12、13、14、15、16、17、18、19、20、21、22、23、24、25、26、27、28、29、30、31、32、33、34、35、36、37、38、39、40、41、42、43、44、45、46、47、48、49、50、51、52、53、54、55、56、57、58、59、60、61、62、63、64、65、66、67、68、69、70、71、72、73、74、75、76、77、78、79、80、81、82、83、84、85、86、87、88、89、90、91、92、93、94、95、96、97、98、99、100或更多个核苷酸，和其中的任何范围)(例如，长度为大约2至大约40，大约2至大约50，大约2至大约60，大约4至大约40，大约4至大约50，大约4至大约60，大约5至大约40，大约5至大约50，大约5至大约60，大约9至大约40，大约9至大约50，大约9至大约60，大约10至大约40，大约10至大约50，大约10至大约60，大约40至大约100，大约50至大约100，或者大约2、3、4、5、6、7、8、9、10、11、12、13、14、15、16、17、18、19、20、21、22、23、24、25个核苷酸至大约26、27、28、29、30、31、32、33、34、35、36、37、38、39、40、41、42、43、44、45、46、47、48、49、50、51、52、53、54、55、56、57、58、59、60、61、62、63、64、65、66、67、68、69、70、71、72、73、74、75、76、77、78、79、80、81、82、83、84、85、86、87、88、89、90、91、92、93、94、95、96、97、98、99、100或更多个核苷酸(例如，长度为大约105、110、115、120、130、140、150或更多个核苷酸))。
如在本文中所使用的，“靶核酸”、“靶DNA”、“靶核苷酸序列”、“靶区域”或“基因组中的靶区域”是指与在本发明的指导RNA中的间隔子序列完全互补(100％互补)或实质上互补(例如，至少70％互补(例如，70％、71％、72％、73％、74％、75％、76％、77％、78％、79％、80％、81％、82％、83％、84％、85％、86％、87％、88％、89％、90％、91％、92％、93％、94％、95％、96％、97％、98％、99％或更多))的生物的基因组的区域(例如，所述间隔子与所述靶核酸的靶标链实质上互补)。对CRISPR-Cas系统来说有用的靶区域可以紧接地位于在生物的基因组(例如，植物基因组)中的PAM序列的3'(例如，V型CRISPR-Cas系统)或紧接地位于其5'(例如，II型CRISPR-Cas系统)。靶区域可以选自紧接地位于靶标链上的PAM序列邻近的至少15个连续核苷酸(例如，16、17、18、19、20、21、22、23、24、25、26、27、28、29、30个核苷酸，等等)的任何区域。
“原间隔子序列”是指靶双链DNA，和特别地是指与CRISPR重复序列-间隔子序列(例如，指导RNA、CRISPR阵列、crRNA)的间隔子序列完全或实质上互补(并且杂交)的靶核酸/靶DNA(例如，基因组(例如，核基因组、质体基因组、线粒体基因组)中的靶区域，或者基因组外序列，例如质粒、微型染色体等)的那一部分。因此，所述原间隔子序列与所述靶核酸的靶标链互补。在一些实施方案中，靶核酸可以具有第一链和第二链(双链DNA)。在一些实施方案中，如在本文中关于靶核酸所使用的，术语“第一链”可以指靶标链或下部链。在一些实施方案中，如在本文中关于靶核酸所使用的，术语“第二链”为与所述第一链互补的链(例如，上部链或非靶标链)。
如在本领域中所理解的和如在本文中所使用的，“靶标链”是指所述间隔子与之互补的并且所述CRISPR-Cas效应蛋白被募集至的双链DNA的那一条链，而“非靶标链”是指在双链核酸中与靶标链相对的那一条链。在本发明的一些实施方案中，将双链核酸的非靶标链(所述CRISPR-Cas效应蛋白被募集至的那一条链的相对链)通过CRISPR-Cas效应蛋白来产生切口并且通过逆转录酶来进行编辑。在一些实施方案中，将双链核酸的靶标链(所述CRISPR-Cas效应蛋白被募集至的同一链)通过CRISPR-Cas效应蛋白来产生切口并且通过逆转录酶来进行编辑。
在V型CRISPR-Cas(例如，Cas12a)系统和II型CRISPR-Cas(Cas9)系统的情况下，所述原间隔子序列的侧翼为(例如，紧邻)原间隔子邻近基序(PAM)。对于IV CRISPR-Cas系统，所述PAM位于在非靶标链上的5'末端和靶标链的3'末端(作为例子，参见下面)。
在II型CRISPR-Cas(例如，Cas9)系统的情况下，所述PAM紧接地位于所述靶区域的3'。关于I型CRISPR-Cas系统的PAM位于所述靶标链的5'。关于III型CRISPR-Cas系统，没有已知的PAM。Makarova等人描述了对于CRISPR系统的所有类别、类型和亚型的命名法(Nature Reviews Microbiology 13:722-736(2015))。指导结构和PAM由R.Barrangou(Genome Biol.16:247(2015))进行了描述。
规范的Cas12a PAM是富含T的。在一些实施方案中，规范的Cas12a PAM序列可以为5'-TTN、5'-TTTN或5'-TTTV。在一些实施方案中，规范的Cas9(例如，酿脓链球菌)PAM可以为5'-NGG-3'。在一些实施方案中，可以使用非规范的PAM，但是它可能是效率较低的。
另外的PAM序列可以由本领域技术人员通过已建立的实验和计算方法来确定。因此，例如，实验方法包括靶向侧翼为所有可能的核苷酸序列的序列，并且鉴定不经历靶向的序列成员，例如通过靶质粒DNA的转化(Esvelt等人,2013.Nat.Methods 10:1116-1121；Jiang等人,2013.Nat.Biotechnol.31:233-239)。在一些方面，计算方法可以包括进行天然间隔子的BLAST搜索以鉴定在噬菌体或质粒中的原始靶DNA序列，并且比对这些序列以确定邻近靶序列的保守序列(Briner和Barrangou.2014.Appl.Environ.Microbiol.80:994-1001；Mojica等人,2009.Microbiology 155:733-740)。
在一些实施方案中，本发明进一步提供了修饰靶核酸的方法，所述方法包括：使所述靶核酸在第一位点处与下列组分相接触：(a)(i)第一CRISPR-Cas效应蛋白；和(ii)第一延伸型指导核酸(例如，第一延伸型CRISPR RNA、第一延伸型CRISPR DNA、第一延伸型crRNA、第一延伸型crDNA)；和(b)(i)第二CRISPR-Cas效应蛋白；(ii)第一逆转录酶；和(ii)第一指导核酸，由此修饰所述靶核酸。在一些实施方案中，本发明的方法可以进一步包括使所述靶核酸与下列组分相接触：(a)第三CRISPR-Cas效应蛋白；和(b)第二指导核酸，其中所述第三CRISPR-Cas效应蛋白对于位于离已经由所述第二CRISPR-Cas效应蛋白产生切口的在第二链上的第二位点大约10至大约125个碱基对处(5'或3')的在所述靶核酸的第一链上的位点产生切口，由此改善错配修复。在一些实施方案中，本发明的方法可以进一步包括使所述靶核酸与下列组分相接触：(a)第四CRISPR-Cas效应蛋白；(b)第二逆转录酶；和(c)第二延伸型指导核酸(例如，第二延伸型CRISPR RNA、第二延伸型CRISPR DNA、第二延伸型crRNA、第二延伸型crDNA)，其中所述第二延伸型指导核酸靶向在所述靶核酸的第一链上的位点(间隔子与之实质上互补/与之相结合)，由此修饰所述靶核酸。对本发明来说有用的CRISPR-Cas效应蛋白(例如，第一、第二、第三、第四)可以为在本文中所描述的任何的I型、II型、III型、IV型或V型CRISPR-Cas效应蛋白，以任何组合。在一些实施方案中，所述CRISPR-Cas效应蛋白可以为Cas9、C2c3、Cas12a(也称为Cpf1)、Cas12b、Cas12c、Cas12d、Cas12e、Cas13a、Cas13b、Cas13c、Cas13d、Casl、CaslB、Cas2、Cas3、Cas3'、Cas3”、Cas4、Cas5、Cas6、Cas7、Cas8、Cas9(也称为Csnl和Csx12)、Cas10、Csyl、Csy2、Csy3、Csel、Cse2、Cscl、Csc2、Csa5、Csn2、Csm2、Csm3、Csm4、Csm5、Csm6、Cmrl、Cmr3、Cmr4、Cmr5、Cmr6、Csbl、Csb2、Csb3、Csxl7、Csxl4、Csx10、Csx16、CsaX、Csx3、Csxl、Csxl5、Csfl、Csf2、Csf3、Csf4(dinG)和/或Csf5。
在一些实施方案中，对所述第一CRISPR-Cas效应蛋白来说有用的延伸型指导核酸可以包含：(a)CRISPR核酸(CRISPR RNA、CRISPR DNA、crRNA、crDNA)；和(b)包含引物结合位点和逆转录酶模板(RT模板)的延伸部分，其中所述RT模板编码待被掺入到所述靶核酸中的修饰(如在本文中所描述的)(例如，编码位于RT模板之内的任何位置中的编辑，其具有相对于所述靶核酸的原间隔子邻近基序(PAM)的位置而言的位置定位，任选地位于核苷酸位置-1至核苷酸位置19，核苷酸位置10至核苷酸位置17，或核苷酸位置12至核苷酸位置15处的编辑)。
在一些实施方案中，所述延伸型指导核酸的CRISPR核酸包含能够与在所述靶核酸的第一链上的第一位点相结合(与之具有实质上的同源性)的间隔子序列。
在一些实施方案中，对CRISPR-Cas效应蛋白来说有用的指导核酸包含CRISPR核酸(CRISPR RNA、CRISPR DNA、crRNA、crDNA)。在一些实施方案中，所述第一指导核酸的CRISPR核酸包含与处于在所述靶核酸的第一链上的第一位点上游(3')的在所述靶核酸的第一链上的第二位点相结合的间隔子序列。
在一些实施方案中，所述第二CRISPR-Cas效应蛋白可以为包含融合至所述逆转录酶的CRISPR-Cas效应蛋白结构域的CRISPR-Cas融合蛋白。
在一些实施方案中，所述第二CRISPR-Cas效应蛋白可以为包含融合至肽标签的CRISPR-Cas效应蛋白结构域的CRISPR-Cas融合蛋白，并且所述逆转录酶可以为包含融合至能够结合所述肽标签的亲和多肽的逆转录酶结构域的逆转录酶融合蛋白。
在一些实施方案中，所述第一指导核酸可以连接至RNA募集基序，并且所述逆转录酶可以为包含融合至能够结合所述RNA募集基序的亲和多肽的逆转录酶结构域的逆转录酶融合蛋白。
在一些实施方案中，可以进一步使所述靶核酸与5'-3'核酸外切酶相接触，任选地其中所述5'-3'核酸外切酶融合至所述第一CRISPR-Cas效应蛋白。在一些实施方案中，5'-3'核酸外切酶可以为包含融合至肽标签的5'-3'核酸外切酶的融合蛋白，并且所述第一CRISPR-Cas效应蛋白可以为包含融合至能够与所述肽标签相结合的亲和多肽的CRISPR-Cas效应蛋白结构域的融合蛋白。在一些实施方案中，5'-3'核酸外切酶可以为包含融合至能够与所述肽标签相结合的亲和多肽的5'-3'核酸外切酶的融合蛋白，并且所述第一CRISPR-Cas效应蛋白可以为包含融合至肽标签的CRISPR-Cas效应蛋白结构域的融合蛋白。在一些实施方案中，5'-3'核酸外切酶可以为包含融合至能够结合RNA募集基序的亲和多肽的5'-3'核酸外切酶的融合蛋白，并且所述延伸型指导核酸连接至RNA募集基序。
在一些实施方案中，本发明进一步提供了使靶核酸与一个或多个单链DNA结合蛋白(ssDNA BP)相接触。单链DNA结合蛋白(ssDNA BP)可以是对于稳定在本发明的方法期间生成的单链DNA来说有用的。虽然不希望受任何特定理论的束缚，但是ssDNA BP可以保护DNA链免于降解或者防止它们变得对于RT-介导的引发和聚合来说不可得。对本发明来说有用的单链DNA结合蛋白可以包括但不限于从实例性的ssDNA BP中获得的那些，其包括但不限于来自人、细菌或噬菌体的那些。在一些实施方案中，ssDNA BP包括但不限于：hRad51(任选地、hRad51_S208E_A209D)(SEQ ID NO:123)、hRad52(SEQ ID NO:124)、BsRecA(SEQ IDNO:125)、EcRecA(SEQ ID NO:126)、T4ssB(SEQ ID NO:127)和/或Brex27(SEQ ID NO:128)。在一些实施方案中，可以使靶核酸与一个或多个ssDNA BP相接触，其中所述ssDNA BP可以融合至CRISPR-Cas效应蛋白(例如，CRISPR-Cas效应蛋白，第一CRISPR-Cas效应蛋白、第二CRISPR-Cas效应蛋白、第三CRISPR-Cas效应蛋白和/或第四CRISPR-Cas效应蛋白)的C-末端或N-末端。ssDNA BP可以融合至所述CRISPR-Cas效应蛋白/结构域的C-末端或N-末端。在一些实施方案中，所述ssDNA BP融合至II型CRISPR-Cas效应蛋白/结构域和/或V型CRISPR-Cas效应蛋白/结构域。
在一些实施方案中，本发明的方法可以进一步包括通过引入非同源末端连接(NHEJ)的化学抑制剂，通过引入靶向NHEJ蛋白的CRISPR指导核酸或siRNA以瞬时敲低所述NHEJ蛋白的表达，或者通过引入阻止NHEJ的多肽来减少双链断裂。在一些实施方案中，所述阻止NHEJ的多肽可以包括但不限于Gam蛋白，任选地其中所述Gam蛋白为埃希氏菌噬菌体MuGam蛋白(例如，SEQ ID NO:147)。
在一些实施方案中，提供了延伸型指导核酸，所述延伸型指导核酸包含：(i)V型CRISPR核酸或II型CRISPR核酸(II型或V型CRISPR RNA、II型或V型CRISPR DNA、II型或V型crRNA、II型或V型crDNA)和/或V型CRISPR核酸或II型CRISPR核酸和tracr核酸(例如，II型或V型tracrRNA、II型或V型tracrDNA)；和(ii)包含引物结合位点和逆转录酶模板(RT模板)(RTT)的延伸部分。在一些实施方案中，所述延伸型指导核酸进一步包含结构化RNA基序，任选地其中所述结构化RNA基序位于所述延伸型指导核酸的3'末端处。在一些实施方案中，所述结构化RNA基序可以包括但不限于：AsCpf1BB(SEQ ID NO:189)、BoxB(SEQ ID NO:190)、假结(诱饵)(SEQ ID NO:95、SEQ ID NO:203)、假结(tEvoPreQ1)(SEQ ID NO:191)、fmp结(SEQ ID NO:192)、mp结(SEQ ID NO:193)、MS2(SEQ ID NO:194)、PP7(SEQ ID NO:195)、SLBP(SEQ ID NO:196)、TAR(SEQ ID NO:197)和/或ThermoPh(SEQ ID NO:198)。在一些实施方案中，所述结构化RNA基序为假结，任选地其中所述假结位于所述延伸型指导核酸的3'末端处。在一些实施方案中，对本发明来说有用的假结可以为天然出现的假结或合成的假结。假结在本文中也可以称为假结样结构、假结化发夹和/或诱饵假结化发夹。在一些实施方案中，所述假结可以位于所述延伸型指导核酸的3'末端处。在一些实施方案中，当所述延伸型指导包含5'-3'crRNA-RTT-PBS时，所述假结可以位于所述RTT的5'处或所述PBS的3'处。在一些实施方案中，当所述延伸型指导包含在所述crRNA的5'末端处的延伸(延伸部分)时，假结可以位于所述RTT的3'处或所述PBS的5'处。在一些实施方案中，假结可以位于所述延伸型指导核酸的5’末端处，随后按5'-3'为PBS，然后为RTT，在crRNA中的天然假结(例如，在重复序列中)，随后为互补区域(例如，间隔子序列)。
在一些实施方案中，对所述延伸型指导来说有用的假结可以包括但不限于：包含SEQ ID NO:158的核酸序列的tEvoPreQ1假结、包含SEQ ID NO:191的核酸序列的EvoPreQ1假结和/或包含SEQ ID NO:95或SEQ ID NO:203的核酸序列的假结。本发明的延伸型指导核酸可以被包含在表达盒中，任选地其中所述表达盒被包含在载体中。
在一些实施方案中，提供了复合物，所述复合物包含：(a)II型CRISPR-Cas效应蛋白或V型CRISPR-Cas效应蛋白；(b)逆转录酶；和(c)延伸型指导核酸(例如，延伸型CRISPRRNA、延伸型CRISPR DNA、延伸型crRNA、延伸型crDNA；例如，tagDNA、tagRNA)。
在一些实施方案中，复合物的II型或V型CRISPR-Cas效应蛋白可以为包含融合至肽标签的II型或V型CRISPR-Cas效应蛋白结构域的融合蛋白。在一些实施方案中，所述复合物的II型或V型CRISPR-Cas效应蛋白可以为包含融合至能够结合肽标签的亲和多肽的II型或V型CRISPR-Cas效应蛋白结构域的融合蛋白。在一些实施方案中，所述复合物的II型或V型CRISPR-Cas效应蛋白可以为包含融合至能够结合RNA募集基序的亲和多肽的II型或V型CRISPR-Cas效应蛋白结构域的融合蛋白。
在一些实施方案中，所述复合物的逆转录酶可以为包含融合至肽标签的逆转录酶结构域的融合蛋白。在一些实施方案中，所述复合物的逆转录酶可以为包含融合至能够结合肽标签的亲和多肽的逆转录酶结构域的融合蛋白。在一些实施方案中，所述复合物的逆转录酶可以为包含融合至能够结合RNA募集多肽的亲和多肽的逆转录酶结构域的融合蛋白。在一些实施方案中，所述复合物可以进一步包含指导核酸(例如，延伸型CRISPR RNA、延伸型CRISPR DNA、延伸型crRNA、延伸型crDNA)。在一些实施方案中，所述复合物可以进一步包含延伸型指导核酸(例如，延伸型CRISPR RNA、延伸型CRISPR DNA、延伸型crRNA、延伸型crDNA)。
在一些实施方案中，所述复合物的延伸型指导核酸可以进一步包含假结。在一些实施方案中，在所述复合物的延伸型指导核酸中所包含的假结可以位于所述延伸型指导核酸的3'末端处。在一些实施方案中，对本发明的复合物的延伸型指导核酸来说有用的假结可以为天然出现的假结或合成的假结。假结在本文中也可以称为假结样结构、假结化发夹和/或诱饵假结化发夹。在一些实施方案中，所述假结可以位于所述延伸型指导核酸的3'末端处。在一些实施方案中，当所述延伸型指导包含5'-3'crRNA-RTT-PBS时，所述假结可以位于所述RTT的5'处或所述PBS的3'处。在一些实施方案中，假结可以包括但不限于：包含SEQID NO:158的核酸序列的tEvoPreQ1假结、包含SEQ ID NO:191的核酸序列的EvoPreQ1假结或包含SEQ ID NO:95或SEQ ID NO:203的核酸序列的假结。
在一些实施方案中，本发明的复合物可以被包含在表达盒中，任选地其中所述表达盒被包含在载体中。在一些实施方案中，包含本发明的复合物的表达盒可以进行密码子优化以用于在生物中进行表达(如在本文中所描述的)，任选地其中所述生物为动物(例如人)、植物、真菌、古细菌、细菌或病毒。
本发明进一步提供了经密码子优化以用于在生物中进行表达的表达盒，其以5'至3'包含：(a)编码启动子序列的多核苷酸；(b)编码V型CRISPR-Cas核酸酶(例如，Cpf1(Cas12a)、dCas12a等)或II型CRISPR-Cas核酸酶(例如，Cas9、dCas9等)的多核苷酸，其进行了密码子优化以用于在所述生物中进行表达；(c)连接体序列；和(d)编码逆转录酶的多核苷酸，其进行了密码子优化以用于在所述生物中进行表达，任选地其中所述生物为动物(例如，人)、植物、真菌、古细菌、细菌或病毒。进一步提供了经密码子优化以用于在植物中进行表达的表达盒，其以5'至3'包含：(a)编码植物特异性启动子序列(例如，ZmUbi1、MtUb2、RNA聚合酶II(Pol II))的多核苷酸；(b)经植物密码子优化的编码V型CRISPR-Cas核酸酶(例如，Cpf1(Cas12a)、dCas12a等)的多核苷酸；(c)连接体序列；和(d)经植物密码子优化的编码逆转录酶的多核苷酸。在一些实施方案中，连接体序列可以为在本文中所描述的氨基酸或肽连接体。在一些实施方案中，在表达盒中的逆转录酶可以融合至一个或多个ssRNA结合结构域(RBD)。
本发明进一步提供了经密码子优化以用于在植物中进行表达的表达盒，其包含：(a)编码植物特异性启动子序列(例如，ZmUbi1、MtUb2)的多核苷酸，和(b)延伸型RNA指导序列，其中所述延伸型指导核酸包含延伸部分，所述延伸部分在其3'末端处包含引物结合位点和待被掺入到所述靶核酸中的编辑(例如，逆转录酶模板)，任选地其中所述延伸型指导核酸被包含在表达盒中，任选地其中所述延伸型指导核酸可操作地连接至Pol II启动子。
在一些实施方案中，所述表达盒包含延伸型指导核酸，其进一步包含结构化RNA基序，任选地其中所述结构化RNA基序位于所述延伸型指导核酸的3'末端处。在一些实施方案中，所述结构化RNA基序可以包括但不限于：AsCpf1BB(SEQ ID NO:189)、BoxB(SEQ ID NO:190)、假结(诱饵)(SEQ ID NO:95、SEQ ID NO:203)、假结(tEvoPreQ1)(SEQ ID NO:191)、fmp结(SEQ ID NO:192)、mp结(SEQ ID NO:193)、MS2(SEQ ID NO:194)、PP7(SEQ ID NO:195)、SLBP(SEQ ID NO:196)、TAR(SEQ ID NO:197)和/或ThermoPh(SEQ ID NO:198)。在一些实施方案中，所述结构化RNA基序为假结，任选地其中所述假结位于所述延伸型指导核酸的3'末端处。在一些实施方案中，对所述延伸型指导来说有用的假结可以包括但不限于包含SEQ ID NO:158、SEQ ID NO:191、SEQ ID NO:95和/或SEQ ID NO:203的核酸序列的假结。
在一些实施方案中，对本发明的表达盒来说有用的植物特异性启动子可以与内含子相关联或者为包含内含子的启动子区(例如，包含内含子的ZmUbi1；包含内含子的MtUb2)。
在一些实施方案中，所述表达盒可以进行密码子优化以用于在双子叶植物中进行表达。在一些实施方案中，所述表达盒可以进行密码子优化以用于在单子叶植物中进行表达。
在一些实施方案中，本发明提供了用于在植物或植物细胞中修饰靶核酸的方法，其包括将一个或多个本发明的表达盒引入到所述植物或植物细胞中，由此在所述植物或植物细胞中修饰所述靶核酸，从而产生包含所述经修饰的靶核酸的植物或植物细胞。在一些实施方案中，本发明的方法进一步包括从包含所述经修饰的靶核酸的植物细胞再生出植物，从而产生包含所述经修饰的靶核酸的植物。在一些实施方案中，本发明的方法包括使所述靶核酸在大约20℃至42℃(例如，大约20、21、22、23、24、25、26、27、28、29、30、31、32、33、34、35、36、37、38、39、40、41或42℃，和其中的任何值或范围)的温度下进行接触。
在一些实施方案中，本发明提供了包含一个或多个本发明的多核苷酸、指导核酸、核酸构建体、表达盒或载体的细胞。
当与指导核酸相组合地进行使用时，本发明的多核苷酸/核酸构建体/表达盒可以用于修饰靶核酸。在使靶核酸与指导核酸相接触之前、同时或之后，可以使所述靶核酸与本发明的多核苷酸/核酸构建体/表达盒相接触。在一些实施方案中，本发明的多核苷酸和指导核酸可以被包含在相同的表达盒或载体中，并且因此可以同时使靶核酸与本发明的多核苷酸和指导核酸相接触。在一些实施方案中，本发明的多核苷酸和指导核酸可以在不同的表达盒或载体中，并且因此可以使靶核酸在与指导核酸相接触之前、同时或之后与本发明的多核苷酸相接触。
可以使用本发明的多核苷酸和方法来修饰(例如，突变，例如碱基编辑、切割、产生切口等)任何生物的靶核酸，所述生物包括但不限于真核生物或原核生物，例如植物、动物、细菌、古细菌、真菌和/或病毒。可以使用本发明的多核苷酸来修饰(例如，突变，例如碱基编辑、切割、产生切口等)任何动物或其细胞，所述动物包括但不限于昆虫、鱼类、鸟类、两栖动物、爬行动物和/或哺乳动物。对于其来说本发明可以是有用的示例性哺乳动物包括但不限于：灵长类(人和非人(例如，黑猩猩、狒狒、猴子、大猩猩等))、猫、狗、雪貂、沙鼠、仓鼠、牛、猪、马、山羊、驴或绵羊。在一些实施方案中，真菌靶生物可以包括但不限于接合菌门、子囊菌门、担子菌门和半知菌门(不完全真菌)，任选地其中所述真菌靶生物可以为子囊菌，任选地酵母。在一些实施方案中，真菌靶生物可以来自糖酵母属(Saccharomyces)，任选地酿酒糖酵母。
可以使用本发明的多核苷酸来修饰(例如，突变，例如碱基编辑、切割、产生切口等)任何植物或植物部分的靶核酸。可以使用本发明的核酸构建体来修饰任何植物(或被分组为例如属或更高的目类别的植物组别)，包括被子植物，裸子植物，单子叶植物，双子叶植物，C3、C4、CAM植物，苔藓植物，蕨类植物和/或拟蕨植物、微观藻类，和/或大型藻类。对本发明来说有用的植物和/或植物部分可以是任何植物物种/变种/栽培种的植物或植物部分。如在本文中所使用的，术语“植物部分”包括但不限于胚、花粉、胚珠、种子、叶、茎、苗、花、枝条、果实、谷粒、穗、穗轴、壳、柄、根、根尖、花药、植物细胞(包括在植物和/或植物部分中是完整的植物细胞)、植物原生质体、植物组织、植物细胞组织培养物、植物愈伤组织、植物团块等。如在本文中所使用的，“苗”是指包括叶和茎的地上部分。进一步地，如在本文中所使用的，“植物细胞”是指植物的结构和生理学单元，其包含细胞壁，并且也可以是指原生质体。植物细胞可以处于分离的单细胞的形式，或者可以是培养的细胞，或者可以是经更高地组织的单元(例如，植物组织或植物器官)的一部分。
对本发明来说有用的植物的非限制性例子包括：草坪草(例如，早熟禾、剪股颖、黑麦草、羊茅)，羽毛芦苇草，发草，芒，芦竹，柳枝稷，蔬菜作物，包括洋蓟，球茎甘蓝，芝麻菜，韭葱，芦笋，莴苣(例如，头型莴苣、叶莴苣、长叶莴苣)，黄体芋，甜瓜类(例如，香瓜、西瓜、克兰省甜瓜、香蜜瓜、罗马甜瓜)，芸苔类作物(例如，抱子甘蓝、卷心菜、花椰菜、球花甘蓝、羽衣甘蓝、无头甘蓝、大白菜、小白菜)，菜蓟，胡萝卜，中国卷心菜(napa)，秋葵，洋葱，芹菜，欧芹，鹰嘴豆，欧洲防风，菊苣，胡椒，马铃薯，葫芦类(例如，西葫芦，黄瓜，密生西葫芦，笋瓜，南瓜，蜜瓜，西瓜，罗马甜瓜)，萝卜，干球洋葱，芜菁甘蓝，茄子，婆罗门参，苣荬菜，青葱，荷兰莴苣，大蒜，菠菜，绿洋葱，笋瓜，绿叶蔬菜(greens)，甜菜类(糖用甜菜和饲用甜菜)，甘薯，莙荙菜，辣根，番茄，芜菁，和香料；水果作物，例如苹果，杏，樱桃，油桃，桃，梨，李子，洋李，樱桃，榅桲，无花果，坚果类(例如，栗子、美洲山核桃、阿月浑子、榛子、阿月浑子、花生、胡桃、澳洲坚果、杏仁等)，柑橘类(例如，克莱门氏小柑橘、金橘、橙子、葡萄柚、红橘、桔子、柠檬、来檬等)，蓝莓，黑覆盆子，博伊森树莓，酸果蔓，茶藨子，鹅莓，罗甘莓，覆盆子，草莓，黑莓，葡萄(酿酒葡萄和食用葡萄)，鳄梨，香蕉，猕猴桃，柿子，石榴，菠萝，热带水果，梨果，甜瓜，芒果，番木瓜，和荔枝，大田作物植物例如三叶草，苜蓿，梯牧草，月见草，白芒花(meadow foam)，玉米/玉蜀黍(大田玉米、甜玉米、爆花用玉米)，蛇麻草，希蒙得木，荞麦，红花，昆诺阿藜，小麦，稻，大麦，黑麦，粟，高粱，燕麦，黑小麦，高粱，烟草，木棉花，豆科植物(豆类(例如，菜豆和干豆)、兵豆、豌豆、大豆)，油料植物(油菜、卡诺拉油菜、芥菜、罂粟、橄榄、向日葵、椰子、蓖麻油植物、可可豆、落花生、油棕)，浮萍，拟南芥属(Arabidopsis)，纤维植物(棉花、亚麻、大麻、黄麻)，大麻属(Cannabis)(例如，大麻(Cannabis sativa)、印度大麻(Cannabis indica)和莠草大麻(Cannabis ruderalis))，樟科(肉桂、樟树)，或植物例如咖啡、甘蔗、茶和天然橡胶植物；和/或花坛植物例如有花植物、仙人掌、肉质植物和/或观赏植物(例如，蔷薇、郁金香、紫罗兰)，以及树木例如林木(阔叶树和常绿树，例如针叶树；例如，榆树、白蜡树、橡树、槭树、冷杉、云杉、雪松、松树、桦树、柏树、桉树、柳树)，以及灌木和其他圃育苗木。在一些实施方案中，本发明的核酸构建体和/或编码其的表达盒和/或载体可以用于修饰玉蜀黍、大豆、小麦、卡诺拉油菜、稻、番茄、胡椒、向日葵、覆盆子、黑莓、黑覆盆子和/或樱桃。
本发明进一步包括用于实施本发明的方法的试剂盒。本发明的试剂盒可以包含用于混合、测量、分选、标记等的试剂、缓冲液和器具，以及说明书等，如将会适合于修饰靶核酸的。
在一些实施方案中，本发明提供了试剂盒，其包含一种或多种本发明的核酸构建体和/或包含其的表达盒和/或载体，以及任选的关于其使用的说明书。在一些实施方案中，试剂盒可以进一步包含CRISPR-Cas指导核酸(或延伸型指导核酸)(相应于由本发明的多核苷酸所编码的CRISPR-Cas效应蛋白)和/或包含其的表达盒和/或载体。在一些实施方案中，所述指导核酸/延伸型指导核酸可以提供在与一个或多个本发明的多核苷酸相同的表达盒和/或载体上。在一些实施方案中，指导核酸/延伸型指导核酸可以提供在与包含一个或多个本发明的多核苷酸的那种分开的表达盒或载体上。
在一些实施方案中，所述试剂盒可以进一步包含编码指导核酸的核酸构建体，其中所述构建体包含用于将与靶核酸序列同一或互补的核酸序列克隆到所述指导核酸的主链中的克隆位点。
在一些实施方案中，本发明的核酸构建体可以为可以在所编码的多核苷酸内编码一个或多个内含子的mRNA。在一些实施方案中，包含一个或多个本发明的多核苷酸的表达盒和/或载体可以进一步编码一个或多个对于鉴定转化体来说有用的选择标记(例如，编码抗生素抗性基因、除草剂抗性基因等的核酸)。
现在将会参考下面的实施例来描述本发明。应当意识到的是，这些实施例并不意在限制本发明的权利要求书的范围，而是意在作为某些实施方案的示例。对于技术人员而言而出现的在所例示的方法中的任何变化都意在落于本发明的范围之内。
实施例
等位基因的RNA编码的DNA替换(RNA-encoded DNA-replacement of alleles；REDRAW)利用V型Cas效应子(从游离的DNA 3′末端(退火位点，AS)开始从DNA:RNA杂合体进行聚合的酶)和延伸型指导核酸(即，所靶向的等位基因指导RNA(tagRNA))。这三种大分子串联工作以：i)通过使用CRISPR效应子和tagRNA的crRNA部分来将CRISPR酶定位至目的基因组位点，ii)对于DNA产生切口或进行切割以产生游离的3′末端，iii)提供与DNA的游离的3′末端退火的tagRNA的一部分，iv)提供为RNA依赖性DNA聚合酶提供模板的tagRNA的一部分，和v)允许通过酶碰撞、天然终止或遇到稳定的发夹来终止逆转录。
我们使用非靶标链(NTS)切口酶版本的LbCas12a_R1138A和来自莫洛尼鼠类白血病病毒(M-MuLV)的RT测试了REDRAW系统。基于与以前所描述的AsCas12a_R1226A突变的比对，预期LbCas12a_R1138A为NTS切口酶。我们在图XXX中证实，LbCas12a_R1138A的确为切口酶。所使用的LbCas12a为RNA酶(+)或具有阻止RNA酶活性的突变(H759A)。使用LbCas12a_R1138A_H759A突变体来阻止tagRNA的自加工，当制造5′延伸时或者当掺入3′发夹时(例如，包含发夹元件的假结)。
所测试的tagRNA包括包含5′或3′延伸的crRNA。测试了各种退火位点长度，允许从产生切口的非靶标链形成更短或更长的DNA:RNA杂合体。也测试了RNA模板的各种长度。最后，还将两个不同的发夹掺入到LbCas12a crRNA序列、假结化发夹设计和诱饵假结化发夹设计中。
实施例1
LbCas12a_R1138A切口酶测定法
合成核酸构建体，其包含LbCas12a，随后为核浆素NLS和6x组氨酸标签(GeneWiz)(SEQ ID NO:57)，并且将其在NcoI和XhoI之间克隆到pET28a载体中，从而生成pWISE450(SEQ ID NO:58)。存在有添加至在Met-1和Ser-2之间的序列的另外的甘氨酸以有利于克隆。在本文中所呈现的编号排除了该额外的甘氨酸。然后，按照制造商的说明书，使用QuickChange II位点定向诱变试剂盒(Agilent)来制备R1138A突变。然后，将这些表达质粒转化到BL21(DE3)Star感受态大肠杆菌细胞(ThermoFisher Scientific)中。
使所述BL21(DE3)Star细胞在Luria Broth和50ug/ml的卡那霉素中在37℃下生长，直至达到A600＝0.5的光密度。添加异丙基-β-d-1-硫代吡喃型半乳糖苷(IPTG)至0.5mM，并且在18℃下诱导蛋白质过夜。以5,000x g使细胞形成粒状沉淀。按照制造商的实验方案，使用两个柱子来完成纯化：HisTrap柱，随后为MonoS柱(GE Healthcare)。
由Synthego合成CRISPR RNA(crRNA)，具有序列AAUUUCUACUAAGUGUAGAUGGAAUCCC UUCUGCAGCACCUGG(SEQ ID NO:59)(其中指导部分加有下划线)。
待被切割的质粒为pUC19，其具有下述的所插入的序列：TTTCGGAATCCCTTCTGCAGCACCTGG(SEQ ID NO:60)，其中加有下划线的该序列的部分为由LbCas12a所识别的PAM序列，其余部分(无下划线的)为原间隔子序列。将所述pUC19质粒转化到XL1-Blue(Agilent)(大肠杆菌)中，并随后使用Qiagen质粒旋转小型试剂盒来进行纯化。
所述核酸酶测定法通过下述方式来完成：混合10:10:1比例的LbCas12a_R1138:crRNA:质粒，在New England Biolabs缓冲液2.1中在37℃下温育15分钟，在80℃下热灭活20分钟，并且加载到具有包埋的SYBR-Safe染料(Invitrogen)的1％ TAE-琼脂糖凝胶上以对DNA进行染色。如在图4中所显示的，在体外测定法中，LbCas12a_R1138A为切口酶。如在泳道2和3中所显示的，超螺旋的2.8kB质粒以2.0kB的表观大小进行走样(泳道2)，直至由野生型LbCas12a生成双链断裂(泳道3)。突变体酶LbCas12a_R1138A占优势地生成以5.0kB的表观大小进行走样的带切口的产物。泳道4-6显示，增加该突变体酶的浓度不改变比例，直至使用极端高的酶浓度，其导致该质粒(256nM)的全面核酸酶消化。
REDRAW编辑器质粒设计和构建-细菌筛选
通过固相合成来合成REDRAW(等位基因的RNA编码的DNA替换)表达构建体，并且将其在NcoI和XhoI限制位点之间克隆到表达载体pET28a(+)中。所述REDRAW表达载体包含ColE1复制起点，卡那霉素抗性标记，和REDRAW编辑器(在T7启动子的控制之下)和终止子。所述REDRAW编辑器包含Cas12a切口酶(R1138A)或RNA酶死的Cas12a切口酶(R1138A，H759A)，其用XTEN或5R连接体融合至Mu-LV逆转录酶MuLV(5M)的(参见例如，SEQ ID NO:97)(鼠类白血病病毒逆转录酶，其具有五个突变-D200N+L603W+T330P+T306K+W313F)(Anzalone等人,Nature 576(.7785):149-157(2019))。所有的REDRAW编辑器序列均是经大肠杆菌密码子优化的。所测试的REDRAW编辑器构造显示在图5中。在图5中所提供的两个构造具有处于逆转录酶的N-末端的Cas12a，和两个构造具有处于逆转录酶的C-末端的Cas12a。所测试的构造用Cas12a变体建立，所述Cas12a变体具有另外的H759A突变以阻止包含5’延伸的tagRNA的加工。
tagRNA质粒设计和构建-细菌筛选
使用将Cas12a间隔子和支架序列与逆转录酶模板和引物结合位点(对于每个靶标来说独特的)装配在一起的算法来设计tagRNA(所靶向的等位基因指导RNA)文库的序列。在表1中所显示的设计参数横跨宽范围的引物结合位点和逆转录酶模板长度。在表3中所显示的所希望的变化被设计成在成功编辑后赋予对于抗生素的抗性。
表1.在第一文库中的所测试的tagRNA的构造
类型 PBS RTT 在文库中的靶标
5’延伸 10-20nt，1nt步级 10-150nt，5nt步级 2个基因组的，3个质粒的
3’延伸 10-20nt，1nt步级 10-150nt，5nt步级 2个基因组的，3个质粒的
图6显示了在第一文库中的tagRNA的构造。包含RTT和PBS的5’和3’延伸均被包括在该文库中。
以与第一文库相似的方式设计第二文库，同时另外还评价在3’tagRNA延伸构造中正好位于间隔子的3’处的发夹的存在是否将会改善REDRAW编辑。在表2中所显示的设计参数再次询问了宽范围的引物结合位点(PBS)和逆转录酶模板(RTT)长度，但是也集中于来自第一文库的被发现有功能的RTT长度的区域。包含RTT和PBS的5’和3’延伸均被包括在该文库中。另外，包含诱饵发夹的变体也被包括在该第二tagRNA文库中。由于所希望的是将会与天然LbCas12a支架序列相似但是不会被Cas12a蛋白识别和切割的发夹，因而在HIV-1RNA基因组中发现了现有的具有与LbCas12a发夹相似的建构的发夹，并且通过添加UA序列来对其进行修饰以形成假结，如在图7中所显示的。
表2.在第二文库中的所测试的tagRNA的构造
用于细菌筛选的tagRNA质粒构建
通过固相合成和将夹持片段克隆到pTwist Amp Medium Copy(TWIST)中来生成用于tagRNA文库的基础质粒。所述质粒包含p15A复制起点和氨苄青霉素抗性标记。tagRNA从合成的BbaJ23119启动子组成性地表达并且通过T7终止子来终止。合成所评价的第一tagRNA文库，并且由外部销售商(Genewiz)将其克隆到所述tagRNA基础载体中。对于第二文库，合成寡聚物，和然后按照制造商的说明书通过使用NEBHiFi Assembly试剂盒将其克隆到所述tagRNA基础载体中。通过来自所述文库的72个克隆的菌落PCR和桑格测序来调查文库多样性，以确保宽范围的PBS、RTT和靶标被包括在所述文库中并且没有实质的偏倚。
报道质粒设计和构建
通过下述方式来构建包含CloDF13复制起点、氯霉素抗性标记和壮观霉素抗性标记(aadA)的基础报道质粒：PCR扩增CloDF13复制起点和氯霉素抗性标记，并且将其与经PCR扩增的aadA抗性标记相连接。然后，通过下述方式来构建三个包含aadA变体的报道质粒：切出在BamHI和BglII限制位点之间的野生型aadA基因，并且连接在于残基位置Thr61、Leu115或Asp132处包含终止密码子的合成的基因区块中。在构建后通过桑格测序来对所有报道质粒进行验证。另外，在将它们在REDRAW tagRNA筛选实验中进行使用之前，包含在编码序列中具有终止密码子的aadA变体的报道质粒被验证为是壮观霉素和链霉素敏感的。
用于REDRAW编辑的靶标-细菌筛选
在REDRAW编辑实验中测试了五个靶标，其在下面显示在表3中。在所有情况下均使用两个基因组靶标和三个质粒靶标。在所述靶标中的任一个处的成功REDRAW编辑导致对于抗生素(萘啶酮酸或链霉素)的抗性，从而将宿主生物(大肠杆菌)的存活与REDRAW编辑的成功联系起来。
表3.用于细菌REDRAW编辑的靶标
REDRAW tagRNA实验-细菌筛选
用于所有细菌REDRAW tagRNA筛选实验的宿主生物为大肠杆菌BL21(DE3)。在进行选择实验之前，按照制造商的说明书将每种REDRAW表达构建体转化到化学感受态BL21(DE3)中并且铺板到具有卡那霉素的LB琼脂平板上。然后，从转化平板中挑取单个克隆，并且按照以前开发的方法制备电感受态细胞的批次(Sambrook和Russell,Transformationof E.coli by electroporation.Cold Spring Harbor Protocols 2006.1(2006):pdb-prot3933)。然后，将含有每种REDRAW表达构建体的感受态细胞用10ng的每种报道质粒进行电穿孔，在SOC中在37℃和225rpm下恢复1小时，并且铺板到具有卡那霉素和氯霉素的LB琼脂平板上。然后，从转化平板中挑取来自这些平板的单个克隆，并且再次制备电感受态细胞的批次(Sambrook和Russell,Transformation of E.coli by electroporation.ColdSpring Harbor Protocols 2006.1(2006):pdb-prot3933)。下面的表4汇总了为了第一tagRNA文库测试而制备的电感受态细胞的批次。
表4–为了tagRNA文库1选择实验而制备的电感受态细胞
SV40＝NLS，MMLV-RT＝逆转录酶，XTEN＝连接体，nLbCas12a＝切口酶Cas12
选择实验通过首先将100ng的tagRNA文库电穿孔到50uL的每个批次的电感受态细胞中来进行。在37℃下以225rpm摇动来使转化物恢复1小时。在1小时的恢复后，移出1uL的回收物，与99uL的LB相混合，并且铺板到具有合适的抗生素的LB琼脂平板上以检查转化效率。然后，将剩余量的每种转化物添加至29mL的LB+抗生素(用于基因组选择的LB Kan/Carb，和用于质粒选择的LB Kan/Carb/Cam)和0.5mM IPTG。使表达培养物在37℃下进行生长，其中以225rpm摇动过夜。
次日，测量每种表达培养物的OD600。对于每种表达培养物，将1OD铺板到5个平板上(大约0.2OD/平板)，所述平板包含关于REDRAW表达载体(Kan)、tagRNA质粒(Carb)、报道质粒的抗生素，0.5mM IPTG，和另外的选择抗生素(萘啶酮酸或链霉素)。将平板在37℃下温育过夜，并且次日早上观察生长。如果未观察到菌落，那么将平板在37℃下再温育24小时。
挑取在选择平板上观察到的菌落，再划线到具有合适的抗生素的平板上，然后使其经历菌落PCR以扩增用于编辑的靶向基因和用于桑格测序的tagRNA。由Genewiz对于菌落PCR产物进行桑格测序。
第二文库的评价以与第一tagRNA文库相同(具有一处修改)的方式来进行。代替制备20个批次的电感受态细胞，制备一个大批次的含有第二tagRNA文库的电感受态BL21(DE3)。然后，将REDRAW表达构建体(100ng)或REDRAW表达构建体+报道质粒(每种100ng)转化到含有tagRNA文库的电感受态细胞中。以相同方式重复所有的随后步骤。
用第一tagRNA文库来评价REDRAW编辑-细菌筛选
在下面的表5中汇总了对于第一tagRNA文库从选择实验中获得的菌落的数目。对于基因组选择(选择1-8)中的任一个，未观察到菌落。对于质粒选择中的每一个，观察到菌落。
表5.第一tagRNA文库选择实验结果
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对于选择9、12、15和18(aadA Thr61靶标)，观察到细菌菌苔。从这些平板中分离出的菌落是假阳性的。对于选择10、11、13、14、16和17(aadA Leu115靶标和aadA Asp132靶标)，在平板上观察到少量的菌落。在这些平板上的菌落具有tagRNA和靶标(通过菌落PCR扩增出的)两者，并且被送去进行桑格测序以确证所制备的编辑和鉴定负责所述编辑的tagRNA。所有从选择11、14、17和20(aadA Asp132靶标)中评价的菌落均是假阳性的。多个来自选择10(aadA Leu115靶标)的菌落具有所设计的编辑和相关联的tagRNA。经编辑的靶标的测序结果显示在图8中，其证实了在去功能的aadA基因中的TGA→CTG编辑，从而恢复了抗生素抗性。
鉴定出的负责所述编辑的tagRNA的序列与在图8中所显示的编辑相关联：
5’–GTTTCAAAGATTAAATAATTTCTACTAAGTGTAGATTACGGCTCCGCAGTGGATGGCGGTAATTTCTACTAAGTGTAGATGCGGCGCGTTGTTTCATCAAGGCGTACGGTCACCGTAACCAGCAAATCAATATCACTGTGTGGCTTCAGGCCGCCATCCACTGCGG–3’(SEQ ID NO:87)。
来自选择10的蛋白质构造如下：SV40-nCas12a-XTEN-MMLV-RT-SV40。
用第二tagRNA文库来评价REDRAW编辑-基因组选择结果
在下面的表6中汇总了对于第二tagRNA文库从基因组选择实验中获得的菌落的数目。在rpsL选择平板上观察到菌落。
表6.第二tagRNA文库实验结果–在用于基因组选择的选择平板上的菌落
对于选择2.1-2.4和2.9-2.12(gyrA基因组靶标)，在平板上未观察到菌落。对于选择2.5-2.8和2.13-2.16(rpsL基因组靶标)，在这些平板上观察到少量的菌落。将在这些平板上的菌落再划线以验证对于所有抗生素的抗性。然后，将来自这些平板的菌落用于生成tagRNA和靶标的PCR产物以用于桑格测序。桑格测序用于确证所制备的编辑和鉴定负责所述编辑的tagRNA。所有来自选择2.6-2.8和2.13-2.16的菌落都是假阳性的。一个来自选择2.5的菌落具有所设计的编辑AAA至CGT，其赋予链霉素抗性(参见图9)。
鉴定出的与在图9中所显示的编辑相关联的tagRNA的序列为：5’–TATTTCTATAAGTGTAGATTACTCGTGTATATATACTCCG CACCGAGGTTGGTACGAACACCGGGAGTCTTTAACACGACCGCCACGGATCAGGATCACGGAGTGCTCCTGCAGGTTGTGACCTTCACCACCGATGTAGGAAGTCACTTCGAAACCGTTAGTCAGACGAACACGGCATACTTTACGCAGCGCGGAGTTCGGTTTACGAGGAGTGGTAGTATATACACGAGT–3’SEQ ID NO:92。
来自选择2.5的蛋白质构造如下：SV40-MMLV-RT-XTEN-nRVRLbCas12a(H759A)-SV40。
用第二tagRNA文库来评价REDRAW编辑–质粒选择结果
在下面的表7中汇总了对于第二tagRNA文库从质粒选择实验中获得的菌落的数目。
表7.
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对于Leu115和Asp132选择，在平板上观察到菌落。选择2.18、2.19、2.22、2.25、2.28、2.31、2.39和2.40在选择平板上具有菌落。将这些菌落再划线以验证对于所有抗生素的抗性。然后，将它们用于生成tagRNA和靶标的PCR产物以用于桑格测序。桑格测序用于确证所制备的编辑和鉴定负责所述编辑的tagRNA。所有来自选择2.18、2.19、2.22、2.28、2.39和2.40的菌落都是假阳性的。四个来自选择2.25的菌落和两个来自选择2.31的菌落具有所设计的编辑和相关联的tagRNA，如在图10和图11中所显示的。所述四个来自选择2.25的菌落具有相同的编辑和tagRNA。所述两个来自选择2.31的菌落也具有相同的编辑和tagRNA。
鉴定出的来自选择2.25的与图10中的编辑相关联的tagRNA的序列为：
5’–TAATTTCTACTAAGTGTAGATTACGGCTCCGCAGTGGATGGCGGTAAGTCTCCATAGAATGGAGGACAGCGCGGAGAATCTCGCTCTCTCCAGGGGAAGCCGAAGTTTCCAAAAGGTCGTTGATCAAAGCGCGGCGCGTTGTTTCATCAAGGCGTACGGTCACCGTAACCAGCAAATCAATATCACTGTGTGGCTTCAGGCCGCCATCCACTGCGGAT–3’SEQ ID NO:93。
来自选择2.25的蛋白质构造如下：SV40-nCas12a-XTEN-MMLV-RT-SV40。
鉴定出的来自选择2.31的与图11中的编辑相关联的tagRNA的序列为：
5’–TAATTTCAACTAAGTGTAGATTACGGCTCCGCAGTGGATGGCGGTAAGTCTCCATAGAATGGAGGGCGGAGAATCTCGCTCTCTCCAGGGGAAGCCGAAGTTTCCAAAAGGTCGTTGATCAAAGCGCGGCGCGTTGTTTCATCAAGGCGTACGGTCACCGTAACCAGCAAATCAATATCACTGTGTGGCTTCAGGCCGCCATCCACTGCGGAT–3’SEQ ID NO:94。
来自选择2.31的蛋白质构造如下：SV40-MMLV-RT-XTEN-nLbCas12a(H759)-SV40。
在细菌细胞中的所观察到的REDRAW编辑的汇总
下面的表8提供了在大肠杆菌中的所观察到的REDRAW编辑情况的汇总。对于每个实例描述了蛋白质构造(REDRAW编辑器)、被编辑的靶标、tagRNA延伸的定位(Cas12a发夹和指导的5’或3’)、PBS长度和RTT长度。
表8.在大肠杆菌中观察到的REDRAW编辑的汇总
*诱饵发夹序列:TAAGTCTCCATAGAATGGAGG SEQ ID NO:95。
实施例2.在人细胞中的精确编辑活性
在图12中显示了一个进一步的与逆转录酶相联合地使用Cas12a的活性形式的方法，并且将其概述在下面。
·核酸酶活性的Cas12a经由间隔子–靶位点相互作用而被募集至所述位点。
·Cas12a制造双链断裂。任选地，提供5’至3’核酸外切酶以降解非模板链。
·通过使用tagRNA而出现引发。引物结合位点(PBS)编码与模板链DNA互补的在切割位点右边的序列。
·逆转录酶(MMuLV-RT(5M))在靶核酸上从引发位点或引物开始进行延伸(虚线＝延伸)，从而在新合成的链内编辑所希望的变化。
·经由错配修复和DNA连接的DNA中间体的分解生成经编辑的新的DNA链。
方法：
设计延伸型指导RNA以靶向HEK293T细胞中的两个基因组位点，DMNT1和FANCF1。检定了引物结合位点(PBS)和逆转录酶模板(RTT)长度的多样化的组合。指导RNA在靶指导的PAM区域中编码两碱基变化，相应于在-2和-3位置处的TT至AA(将TTTV PAM计数为-4至-1位置)。将指导延伸融合至指导RNA的5’或3’末端。
按照制造商的实验方案，使用LipofectamineTM 3000来将编码RNA酶-死突变体LbCas12a(H758A)、逆转录酶(MMuLV-RT(5M))和任选地核酸外切酶(T5核酸外切酶、T7核酸外切酶、RecE和RecJ之一)和延伸型指导RNA的质粒转染到以70％汇合生长的HEK293T细胞中。在3天后收获细胞，并且通过下一代测序来定量基因编辑。
结果：
我们观察到所意在的对于所述两个所靶向的位点的精确编辑。取决于指导设计，我们观察到在FANCF1位点处的直至0.5％编辑(图13)和在DMNT1位点处的直至1.7％编辑(图14)。核酸外切酶的使用改善了在一些指导设计中的编辑效率。
表9.用于靶向FANCF1位点的指导设计(图13)
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表10.用于靶向DMNT1位点的指导设计(图14)。
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表11.实例性的延伸型指导核酸(tagRNA)
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核酸外切酶转染对于在DMNT1位点处的精确编辑活性的效应显示在图15中(标准化至无核酸外切酶处理；pUC19＝1)。核酸外切酶改善了用一些指导构造的编辑。
实施例3.REDRAW蛋白质构造中的变动
使用不同的对于LbCas12a和RT(5M)的蛋白质建构/构建体测试了本发明的方法(即，REDRAW)，包括：(1)其中所述逆转录酶(RT(5M))通过在细胞中过表达RT来提供；(2)构建体，其中SunTag(GCN4，例如SEQ ID NO:23、SEQ ID NO:24)融合至所述CRISPR-Cas效应蛋白(例如，LbCpf1)，和所述RT(RT(5M))被募集至所述编辑位点，通过将它融合至与融合至CRISPR-Cas效应蛋白的SunTag相结合的抗体(例如，单链可变片段(scFv)抗体)；和(3)其中所述逆转录酶(RT(5M))融合至所述CRISPR-Cas效应蛋白(例如，LbCpf1(LbCas12a)，例如LbCpf1(H759A))的N-末端或C-末端(例如，RT(5M)-LbCpf1(H759A)或LbCpf1(H759A)-RT(5M)))。结果显示在图16中。所有构建体都显示出RT(5M)和tagRNA依赖性REDRAW活性的证据，其中使用了四种不同的tagRNA构建体。在该实施例中，用SunTag的主动募集并未增强活性。可能的是，未加标签的构建体的过表达允许在细胞中的足够水平的蛋白质浓度，并且因此，在这些条件下，这不是对于生成精确编辑的限速方面。
MS2/MCP系统
除了上面所测试的建构外，还就与本发明的构建体和方法一起进行使用评价了MS2/MCP系统。MS2发夹RNA结构与MCP蛋白相结合。可以将MS2发夹添加至所述tagRNA。在该实施例中，将MS2发夹结构添加至所述tagRNA的3’末端，和将MCP融合至RT(5M)以便将RT(5M)募集至靶位点。
使LbCas12a H759A与RT(5M)一起瞬时表达，没有MCP(以反式控制)或具有MCP-RT(5M)(融合构建体)。使用两种tagRNA(tagRNA5和tagRNA6)来测试该建构。我们还比较了不同的tagRNA版本，其中用MS2序列在其3’末端处修饰tagRNA5和tagRNA6。结果显示在图37中。通过比较MCP-RT(5M)和RT(5M)，MS2 tagRNA和MCP-RT(5M)未导致精确编辑效率的增加。MCP融合在这些实验条件下可能无法增加精确编辑效率，因为RT浓度不是限速的。然而，对于在其3'末端处具有MS2的tagRNA，注意到编辑效率的增加。在tagRNA的3’末端处的MS2结构可以稳定所述tagRNA并且减少其降解。
实施例4.用于与本发明的方法(REDRAW)一起使用的5’-3’核酸外切酶
通过在双链断裂的两个末端处都降解DNA，5’-3’核酸外切酶对本发明的方法来说可以是有用的。因此，5’-3’核酸外切酶可以：(1)通过降解与将会被延长的DNA链正常地碱基配对的链而允许更稳固的RNA-DNA双链体形成(关于RT-介导的聚合的底物)；和/或(2)通过降解将会由RT改写的区域而允许细胞偏向于使用RT-合成的DNA用于DNA修复中。参见例如，图17中的示意图。
在该实施例中，所包括的经测试的核酸外切酶为在表11中所列出的那些。
表12.所测试的5'-3'核酸外切酶
将所述5'-3'核酸外切酶融合至LbCas12a(H759A)的C-末端。将融合构建体与逆转录酶(5M)构建体和表达合适的编码精确突变的tagRNA的质粒一起转染到HEK293T细胞中。转染3天后收获细胞，并且通过使用高通量测序(HTS)来分析DNA。结果显示在图18中。在此，RT以反式进行表达(没有募集)，和所述5'-3'核酸外切酶融合至LbCpf1 H759A的C-末端。相比于其中不存在核酸外切酶的构建体(仅LbCpf1 H759A)而言，特别地，T7_Exo的融合在所测试的四个tagRNA中的三个之中改善了REDRAW精确编辑。
图19提供了另外的用本发明的方法(REDRAW)并且在上面所记录的相同条件下进行的5'-3'核酸外切酶测试。特别地，图19显示了使用5'-3'核酸外切酶sbcB(SEQ ID NO:134)或5'-3'核酸外切酶Exo(SEQ ID NO:135)(各自融合至Cas多肽(LbCpf1)的C-末端)的采用REDRAW的精确编辑百分比。RT(5M)(SEQ ID NO:97)以反式进行表达(无募集)。与T7_Exo(SEQ ID NO:132)形成对比，核酸外切酶sbcB和Exo未改善REDRAW。
还用本发明的方法以反式测试了5'-3'核酸外切酶。结果提供在图20中。LbCpf1和RT(5M)(SEQ ID NO:97)以融合蛋白来提供。图20的右侧显示了用融合至LbCpf1(RT(5M)-LbCpf1(H759A))的N-末端的RT的结果，和该图的左侧显示了使用融合至LbCpf1(LbCpf1(H759A)-RT(5M))的C-末端的RT的结果。图20显示，当以反式表达5'-3'核酸外切酶(没有融合至LbCpf1)时，相比于没有5'-3'核酸外切酶的处理而言编辑率不增加。因此，当将所述5'-3'核酸外切酶融合至所述CRISPR-Cas效应蛋白时，观察到来自与本发明的方法一起使用5'-3'核酸外切酶的益处。
实施例5.调节用于与REDRAW一起使用的Cas12a的DNA结合亲和力的突变
设想了降低CRISPR-Cas效应蛋白的DNA结合亲和力以允许所述CRISPR-Cas效应蛋白与所述靶位点的更好的解离，由此诱导双链DNA断裂。这可以允许更快地形成促进通过RT的序列替换的中间体并且增加用本发明的方法(REDRAW)的编辑效率。
在该实施例中，将与DNA主链相互作用的在Cas12a(LbCas12a)中的带正电荷的残基突变为丙氨酸。特别地，将下述三个突变即K167A、K272A、K349A(参考SEQ ID NO:1或SEQID NO:148的氨基酸位置编号)作为单突变体、双突变体或三突变体(K167A、K272A、K349A、K167A+K272A、K167A+K349A、K272A+K349A和K167A+K272A+K349A)克隆到LbCas12a H759A中。在这种情况下，该H759A突变(SEQ ID NO:148)用于使LbCas12a的RNA加工能力失活以推动5’tagRNA延伸至crRNA。
将包含各种结合亲和力突变组合的LbCas12a与编码RT(5M)和tagRNA(其编码精确编辑)的质粒一起转染到HEK293T细胞中。转染后三天收获细胞，并且通过使用高通量测序(HTS)来分析DNA。某些突变组合显示出改善了本发明的方法的精确编辑(图21)。
实施例6.用于与REDRAW一起使用的单链DNA结合蛋白(ssDNA BP)
单链DNA结合蛋白(ssDNA BP)的使用可以潜在地改善REDRAW，通过稳定在反应期间生成的ssDNA，通过保护DNA链免于降解并且使其对于RT-介导的引发和聚合来说是可得的。用本发明的方法来测试ssDNA BP的选择(参见表12)
表13.所评价的单链DNA结合蛋白
名称 来源物种 SEQ ID NO
hRad51(S208E_A209D) 人 123
hRad52 人 124
BsRecA 枯草芽孢杆菌(Bacillus subtilis) 125
EcRecA 大肠杆菌 126
T4SSB 埃希氏菌噬菌体P1 127
Brex27 人 128
以反式或作为与Cas12a的融合物来表达在表12中所阐明的ssDNA BP，也是在RT(5M)(反式)存在下。所测试的ssDNA BP为hRad51_s208E_A209D(SEQ ID NO:123)、hRad52(SEQ ID NO:124)、BsRecA(SEQ ID NO:125)、EcRecA(SEQ ID NO:126)、T4SSB(SEQ ID NO:127)和Brex27(SEQ ID NO:124)。结果显示在图22和图23中。当与对照(pUC19)相比较时，ssDNA BP的反式表达未改善精确编辑百分比(参见图22)。融合蛋白也未能显示出改善，除了Brex27与Cas12的N-末端和C-末端融合物外(参见图23)。Brex27为已知原位募集Rad51并且稳定其与ssDNA的相互作用的肽。
实施例7.用于在REDRAW中使用的Gam蛋白的评价
Gam蛋白通过阻止NHEJ而可以在REDRAW期间减少插入/缺失的形成方面是有帮助的。Gam与双链DNA断裂相结合，从而阻止DNA末端被加工。Gam可以用于在胞嘧啶碱基编辑期间减少插入/缺失形成。
为了评价Gam蛋白对于本发明的方法的有用性，将Gam蛋白(埃希氏菌噬菌体MuGam蛋白)(SEQ ID NO:147)融合至CRISPR-Cas效应蛋白(LbCas12a H759A)(SEQ ID NO:148)或RT(5M)(SEQ ID NO:53)。将编码LbCas12a H759A、RT(5M)和tagRNA(其编码精确突变)的质粒转染到HEK293T细胞中。三天后用高通量测序分析靶DNA。结果显示在图24和图25中。
在图24中，所述逆转录酶(RT)以反式进行表达，作为天然序列(例如，RT(5M))或者在其N-末端处融合至Gam蛋白(例如，Gam-RT(5M))。将这些构建体与LbCas12a(H759A)或者与具有融合至其N-末端的Gam蛋白的LbCas12a(H759A)(例如，Gam-LbCas12a H759A)同时进行表达。在图25中，所述Gam蛋白以反式来提供，作为与逆转录酶的融合蛋白(N-末端融合物；Gam-RT(5M))和/或作为与CRISPR-Cas效应多肽的融合蛋白(例如，Gam-LbCas12aH759A)。结果显示，在一些情况下，Gam蛋白可以用于减少插入/缺失形成，但是使用本发明的方法进行编辑的总效率并没有因包括Gam蛋白而得到改善。
实施例8.引物结合位点(PBS)长度和逆转录酶模板(RTT)长度的评价
使在本发明的tagRNA中的RTT和PBS的长度多样化，以评价长度对于编辑的效应。将LbCas12a、RT(5M)和tagRNA(其具有变化长度的RTT和PBS)转染到HEK293T细胞中，并且在转染三天后使用高通量测序(HTS)就编辑率对其进行分析。结果提供在图26中。图26的上图版和下图版显示了使用两种不同的间隔子的结果(上图版：pwsp143(GCTCAGCAGGCACCTGCCTCAGC)(SEQ ID NO:136)，下图版：pwsp139(CTGATGGTCCATGTCTGTTACTC)(SEQ ID NO:137))。虽然结果随所使用的间隔子而变化，但是对于RTT和PBS两者的许多不同的长度均显示出良好的编辑效率。一个最佳的组合的PBS长度和RTT长度可以分别为48个核苷酸和52个核苷酸。
实施例9.在tagRNA中的编辑布置的评价
REDRAW效率可以取决于所希望的编辑位于tagRNA的逆转录酶模板(RTT)内的何处而变化。在该实施例中，评价了在RTT中编辑的定位对于编辑百分比的效应。编辑定位和结果以粗体提供在图27中。上图版和下图版提供了不同的RTT序列，其中编辑定位发生变化(上图版RTT：SEQ ID NO:187；下图版RTT：SEQ ID NO:188)。在图27的上图版和下图版中的“编辑定位”栏都显示了RTT的前26个碱基(其相应于PAM序列(TTTC)和23-碱基间隔子序列)的反向互补物。我们沿着RTT铺排了所述双突变，从而当将所希望的编辑引入至DNA时，用于REDRAW的间隔子不再是互补的。用在RTT中的处于许多定位处的编辑布置确定编辑是有效的。在一些情况下，将所述编辑放置在RTT序列中的相应于核苷酸12-15(LbCas12a的TTTNPAM分别被定义为位置-4、-3、-2、-1)的位置处提供了非常高水平的编辑。
实施例10.使用不同的CRISPR-Cas效应蛋白来进行REDRAW编辑
设想REDRAW与备选的能够生成双链DNA断裂的CRISPR-Cas效应蛋白是相容的。在本实施例中，LbCas12a与cas9(SpCas9)、BhCas12b、AsCas12a(EnAsCas12a)一起显示，备选的CRISPR-Cas效应蛋白可以成功地用于本发明的方法(REDRAW)。
Cas9
将RT(5M)、编码精确编辑的tagRNA和两种形式的Cas9(Cas9(核酸酶)，nCas9(D10A)(切口酶))转化到HEK293T细胞中并且进行表达。转染后三天收获细胞，并且使用高通量测序(HTS)对靶扩增子进行测序。使PBS和RTT的长度多样化，并且将延伸添加至指导RNA的3’和5’末端(在图28中标示为“3’延伸”或“5’延伸”)。所使用的tagRNA延伸靶向四个不同的靶位点(间隔子：pwsp10：GAGTCCGAGCAGAAGAAGAA(SEQ ID NO:140)；pwsp621：GCATTTTCAGGAGGAAGCGA(SEQ ID NO:141)；pwsp15：GTCATCTTAGTCATTACCTG(SEQ ID NO:142)；pwsp11：GGAATCCCTTCTGCAGCACC(SEQ ID NO:143))。结果提供在图28中。使用多个不同的间隔子序列使用Cas9和nCas9(D10A)均观察到精确的经RT-介导的编辑，但是核酸酶版本表现最好。进一步地，虽然3’和5’tagRNA延伸在REDRAW中都是有效的，但是延伸型指导RNA的3’延伸表现最好。
BhCas12b
将RT(5M)、编码精确编辑的tagRNA和BhCas12b v4(其是经改造的高效率形式的BhCas12b)转化到HEK293T细胞中并且进行表达。转染后三天收获细胞，并且使用高通量测序(HTS)对靶扩增子进行测序。使PBS和RTT的长度多样化，并且将延伸添加至指导RNA的3’和5’末端(在图29中标示为3’或5’)。所使用的tagRNA延伸靶向三个不同的靶位点(间隔子：PWsp1099：ACGTACTGATGTTAACAGCTGA(SEQ ID NO:144)；PWsp1098：GGTCAGCTGTTAACATCAGTAC(SEQ ID NO:145)；PWsp1094：TCCAGCCCGCTGGCCCTGTAAA(SEQ IDNO:146))。结果提供在图29中。使用BhCas12b v4和多个不同的间隔子序列观察到精确的RT-介导的编辑。当使用BhLbCas12b时，RTT和PBS长度的某些组合导致比其他的更高的编辑。通常，当使用BhLbCas12b时，tagRNA的3’延伸提供比5’延伸更一致的编辑，虽然使用这两种形式的tagRNA都检测到编辑。
EnAsCas12a
AsCas12a为LbCas12a的同源物，和EnAsCas12a为经改造的形式的AsCas12a。在EnAsCas12a中的H800A突变相应于在LbCas12a中的H759A突变，其是使Cas12a的crRNA加工能力失活的突变。
将RT(5M)、编码精确编辑的tagRNA和EnAsCas12a H800A(EnAsCpf1 H800A)转化到HEK293T细胞中并且进行表达。在这种情况下，所述逆转录酶作为与EnAsCas12a的融合蛋白(C-末端融合物(EnAsCas12a-RT)和N-末端融合物(RT-EnAsCas12a))来提供。转染后三天收获细胞，并且使用高通量测序(HTS)对靶扩增子进行测序。使用多个不同的tagRNA序列使用EnAsCas12a观察到精确的RT-依赖性和tagRNA-依赖性编辑。所使用的tagRNA延伸靶向单个位点(间隔子：CCTCACTCCTGCTCGGTGAATTT(SEQ ID NO:171))。
结果提供在图30中，其显示：在各种tagRNA存在下，RT和EnAsCas12a的N-末端和C-末端融合物均导致精确编辑。没有RT融合的EnAsCas12a用作对照并且显示出没有或非常低的编辑。
实施例11.在酵母中的编辑
除了显示出人细胞可以通过使用本发明的方法来进行编辑外，还在酿酒糖酵母(酵母)(一种真核生物)中评价了本发明的方法。酿酒糖酵母对于评价本发明的方法来说是一种有吸引力的生物，因为几个原因，包括例如：(1)酿酒糖酵母利用NHEJ修复过程；在基因组中的双链断裂不是致死的，这不同于在定向进化实验中经常使用的原核生物(例如大肠杆菌)之中；(2)酵母生长相对快，从而允许快速地测试和调整对于本发明的方法(REDRAW)的许多条件；(3)数千的酵母菌株是容易可得的；和(4)可以在酵母中研究生物分子(蛋白质、RNA等)的大型文库。
对于本实施例选择酿酒糖酵母菌株W303-1a(下面，“ScW303-1a”)。ScW303-1a的基因型为：MATa ade2-1 ura3-1 his3-11trp1-1 leu2-3 leu2-112 can1-100。用于在该菌株中的编辑的靶标包括ADE2、CAN1、HIS3、LYS2、TRP1和URA3。使用桑格测序来确认关于每个PCR产物的基因座序列。所有经测序的基因座都是如所预期的，除了ADE2外。预期ADE2基因座在Gln64处具有终止密码子；然而，测序显示，代替在Gln64处的终止密码子，存在酪氨酸密码子。因此，构建了具有经修饰的ADE2基因座的定制菌株，以便在那个基因座处测试REDRAW。将该经修饰的菌株命名为ScDS21.6。表13提供了为在酵母中进行测试而选择的基因组靶标。
表14.用于REDRAW编辑的酵母基因组靶标
用于靶向这些位点的实例性的间隔子包括：
PWsp1643(ADE2靶标)：5’–GCATACGATGGAAGAGGTAACTT–3’(SEQ ID NO:159)
PWsp1894(ADE2靶标)：5’–GCATACGATTAAAGAGGTAACTT–3’(SEQ ID NO:160)
PWsp1665(URA3-1靶标)：5’–CAAATAGTCCTCTTTCAACAATA–3’(SEQ ID NO:161)
实例性的引物结合位点序列：
关于ADE2靶标的48-bp引物结合位点：
5’–CGTTGTAAAGAATAAGGAAATGATTCCGGAAGCTTTGGAAGTACTGAA–3’(SEQ ID NO:162)
关于URA3-1靶标的48-bp引物结合位点：
5’–ATAATGTCAGATCCTGTAGAGACCACATCATCCACGGTTCTATACTGT–3’(SEQ ID NO:163)
实例性的逆转录酶模板序列：
(A)ADE2靶标：
40-bp RTT：5’–TGAAGTCGAGGACTTTGGCATACGATGGAAGAGGTAACTT–3’(SEQ ID NO:164)
50-bp RTT：5’–CCATTCGTCTTGAAGTCGAGGACTTTGGCATACGATGGAAGAGGTAACTT–3’(SEQ ID NO:165)
72-bp RTT：5’–TGTTGGAAGAGATTTGGGTTTTCCATTCGTCTTGAAGTCGAGGACTTTGGCATACGATGGAAGAGGTAACTT–3’(SEQ ID NO:166)
(B)URA3-1靶标
47-bp RTT：5’–CTACCTTAGCATCCCTTCCCTTTGCAAATAGTCCTCTCTCAACAATA–3’(SEQID NO:167)
55-bp RTT：5’–TTCACCCTCTACCTTAGCATCCCTTCCCTTTGCAAATAGTCCTCTCTCAACAATA–3’(SEQ ID NO:168)
63-bp RTT：5’–CTGTAACGTTCACCCTCTACCTTAGCATCCCTTCCCTTTGCAAATAGTCCTCTCTCAACAATA–3’(SEQ ID NO:169)
实例性的LbCas12a crRNA支架：
5’–TAATTTCTACTAAGTGTAGAT–3’(SEQ ID NO:170)
使用蛋白质表达载体pESC-LEU，因为(1)它包括酵母选择标记，LEU2，其与ScW303-1a菌株是相容的，(2)在该质粒中的GAL启动子系统提供了强大的蛋白质表达控制，(3)酵母复制起点，2μ，是高拷贝的，从而允许高水平的蛋白质表达，和(4)还存在大肠杆菌复制起点(pUC起点)和选择标记AmpR，从而允许在酵母中进行工作之前在大肠杆菌中进行所有的载体操作和克隆。
使用下述的CRISPR-Cas效应蛋白和逆转录酶构造：
·LbCas12a+在C-末端处融合的MMLV-RT(5M)(SEQ ID NO:155)
·LbCas12a+在N-末端处融合的MMLV-RT(5M)(SEQ ID NO:157)。
将LbCas12a融合物置于诱导型GAL1启动子(pol II启动子)的控制之下，并且从组成性SNR52启动子(pol III启动子)表达crRNA和tagRNA。
另外，与所述两种LbCas12a和RT构造一起测试了下面的tagRNA构造：(1)不存在3’假结，(2)存在假结，(a)称为“诱饵”假结的假结(参见图7，SEQ ID NO:203)或(b)称为tEvoPreQ1假结的假结(SEQ ID NO:158)。另外，用具有48个核苷酸的固定长度的引物结合位点(PBS)测试了三种不同的逆转录酶模板(RTT)长度(47、55、64个核苷酸，或者40、50和72个核苷酸)中的每一种。这些构造阐明在表15中。
表15.实例性的在酵母中测试的REDRAW构造
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通过首先经由PEG/LiAc热休克方法将目的载体转化到酵母菌株ScDS21.6(ADE2靶位点)或酵母菌株ScW303-1a(URA3靶位点)中，在酿酒糖酵母中测试了REDRAW。将转化体铺板到缺乏亮氨酸、具有2％葡萄糖作为碳源的合成的完全培养基(SC-LEU+2％Glu)上。在大约48-72小时后，然后挑取单个菌落到3-mL的液体SC-LEU+2％棉子糖(SC-LEU+2％ Raff)中。使培养物在28℃下在以200rpm进行摇动的情况下生长大约36小时，直至OD600达到～1.8。然后，使1.5OD的细胞在室温下在离心机中旋转离心，并且带回到3mL的蛋白质表达培养基即SC-LEU+1％棉子糖+1％半乳糖(SC-LEU+1％Raff+1％ Gal)中。使表达培养物在28℃下以200rpm摇动生长4小时。然后，从摇动培养箱中取出表达培养物并且进行离心。然后，吸移去上清液，并且添加3mL的SC-LEU+2％ Glu。然后，让细胞在28℃和200rpm下另外再生长90分钟。在90分钟后，检查培养物的OD600。使每种培养物的大约1OD酵母细胞(大约1×107个酵母细胞)在室温下在离心机中形成粒状沉淀。移去上清液，并且将每种培养物重悬浮在200uL的无菌水中。将大约一半的经重悬浮的培养物(0.5OD)(大约5×106个酵母细胞)铺板到缺乏腺嘌呤(SC-ADE)或尿嘧啶(SC-URA)的合成的完全平板上以选择经编辑的菌落，并且将另一半铺板到SC-LEU平板(非选择性的，以看在大约0.5的所测量的OD中有多少细胞)上。使所述平板在28℃下生长大约3天。然后，对菌落进行计数并且记录。从SC-ADE/SC-URA平板或者SC-LEU(阴性对照)平板中选择菌落，使用菌落PCR来扩增所靶向的基因座。使用桑格测序来分析靶基因座，其确证制备了所意欲的编辑(在ADE2中的2-bp变化：AA156 TAA->GGA，和在URA3-1中的1-bp变化：AA 234GGA->GAA)。
在酵母中在两个不同的靶位点处测试了LbCas12a和RT构造/tagRNA组合中的每一个，并且将结果提供在图31和32中。图31显示了URA3-1靶基因的编辑的结果(URA3-1：1-bp变化(AA 234GGA->GAG)(编辑修复了腺嘌呤营养缺陷))，其中上图版显示了用LbCas12-RTC-末端融合物的结果，和下图版显示了关于RT-LbCas12 N-末端融合物的结果。图32显示了ADE2靶基因的编辑的结果(ADE2：2-bp变化(AA 156TAA->GGA)(编辑修复了尿嘧啶营养缺陷))，其中上图版显示了用LbCas12-RT C-末端融合物的结果，和下图版显示了关于RT-LbCas12 N-末端融合物的结果。虽然所有构造都能够编辑URA3-1基因以产生活菌落(修复了腺嘌呤营养缺陷)，但是最有效的构造包括假结和具有55个核苷酸的长度的RTT(图31)。RT,LbCas12a C-末端融合物与“诱饵”假结一起是最有效的，并且RT,LbCas12a N-末端融合物与tEvoPreQ1假结一起是最有效的(图31)。在酵母中ADE2的编辑显示出相似的结果，因为RT,LbCas12a C-末端融合物与“诱饵”假结一起是是最有效的，并且RT,LbCas12a N-末端融合物与tEvoPreQ1假结一起是最有效的(图32)。在ADE2的情况下，用具有50个核苷酸的长度的RTT，编辑是最有效的。
因此，该实施例显示，本发明的方法在所述两个靶位点处和通过使用任一种蛋白质融合物构造均能够精确地编辑酵母，其中对于这两个靶标在C-末端处融合的RT构造比在N-末端处融合的RT略微更有效。以所测试的构造中的每一种，观察到假结改善了REDRAW编辑的效率。进一步地，在tagRNA和REDRAW编辑器不存在下，在选择平板(SC-ADE或SC-URA)上未观察到生长，这表明在酵母中的这些REDRAW测定法是非常严格的，并且逃逸频率低于检测极限。
实施例12.在编辑中的ssRNA结合蛋白的评价
单链RNA结合蛋白(ssRNA BP)为与核糖核酸非特异性地相互作用的蛋白质。当用本发明的方法进行编辑时表达ssRNA结合蛋白可以稳定所暴露的tagRNA组分(延伸型指导核酸)以免于被内源蛋白质降解。为了测试这一点，我们将几种RNA结合蛋白表达为与RT(5M)-LbCas12a(H759A)的N-末端融合物。
使用防卫素(SEQ ID NO:152)和ORF5(SEQ ID NO:153)这些ssRNA结合蛋白的精确编辑结果提供在图33中。将ssRNA BP防卫素和ssRNA BP ORF5各自融合至RT-LbCas12融合蛋白(例如，RT-LbCas12a)的N-末端。相比于未在其N-末端与ssRNA结合蛋白相融合的相同的RT-Cas12a融合蛋白而言，显示出了所述编辑。对于所测试的两种tagRNA(延伸型指导核酸)之一，在使用ssRNA结合蛋白的情况下，精确编辑显示出得到改善。
实施例13.具有不同突变的逆转录酶多肽的评价
逆转录酶RT(5M)通过将五个突变引入到野生型RT序列中来进行改造(Anzalone等人,Nature 576:149-157(2019))。为了评价本发明的方法是否可以通过使用相比于RT(5M)的那种而言具有不同或另外的突变的RT结构域来进一步进行优化，将几个具有不同的突变或突变组合的逆转录酶(RT)蛋白(具有或没有RT(5M)核心突变)在N-末端处融合至LbCas12a(H759A)。所测试的RT结构域包括：RT(L139P,D200N,W388R,E607K)、RT(L139P,D200N,T306K,W313F,W388R,E607K)、RT(5M,F155Y,H638G)、RT(5M,Q221R,V223M)和RT(5M,D524N)。在RT(M)中的突变包括通过参考SEQ ID NO:172的氨基酸序列编号而言的D200N+L603W+T330P+T306K+W313F(参见，SEQ ID NO:53)。对于不包括RT(5M)突变的那些序列关于氨基酸位置编号的参考RT为SEQ ID NO:172。对于包括RT(5M)突变的那些序列关于氨基酸位置编号的参考RT为SEQ ID NO:53。在每种情况下，将RT融合至LbCas12a(H759A)的N-末端。
图34显示了结果。相比于RT(5M)(左)而言，几个其他具有不同突变组合的RT结构域能够相比于RT(5M)而言增加精确编辑。该结果受所使用的tagRNA(延伸型指导核酸)影响。
实施例14.在tagRNA的3'末端处掺入的3’结构化RNA基序的评价
进行实验以评价在tagRNA的3’末端处掺入的结构化RNA是否可以进一步稳定tagRNA并且保护其免于可能的降解。为此目的，将几种已知形成3-D结构的RNA序列(包括发夹和假结)附加至不同的tagRNA。
表16.当被转录和附加至tagRNA的3'末端时相应于RNA结构的DNA序列
表17.关于表16中的RNA结构的来源
在图35中提供了在本发明的组合物中包括各种3'结构化RNA的结果。在该实验中，在HEK293T细胞中与各种tagRNA一起表达RT(5M)-LbCas12a H759A，具有或没有3’RNA结构。在3天后，收获细胞，并且通过高通量测序来分析精确编辑效率。我们观察到，几乎所有的在tagRNA上的3’RNA结构都可以适应本发明的方法(例如，REDRAW)。通过在HEK293T细胞中使用3’RNA结构，我们未观察到REDRAW效率的增加。
实施例15.使用染色质调节肽融合物的评价
可以通过降低它们在活细胞中的活性的核小体来堵塞基因组编辑蛋白质。染色质调节蛋白质/肽可以在解决此类影响中是有帮助的，这通过促进染色质交换、组蛋白修饰和外因基因组修饰，由此增强此类可编程的DNA结合蛋白(例如Cas9或Cas12a)的接近。
为了评价这种可能性，将染色质调节肽(包括CHD1(例如，SEQ ID NO:199)、H1G(例如，SEQ ID NO:200)、HB1(例如，SEQ ID NO:201)和HN1(例如，SEQ ID NO:202)(参见例如，Ding等人,CRISPR J.2019Feb,2:51-63))以如下的各种融合方向融合至所选择的本发明的构建体：HN1-RT(5M)-LBCas12a(H759A)、HN1-RT(5M)-LBCas12a(H759A)-HB1、HN1-RT(5M)-LBCas12a(H759A)-H1G、HN1-RT(5M)-LBCas12a(H759A)-CHD1、HN1-RT(5M)-H1G-LBCas12a(H759A)和HN1-RT(5M)-CHD1-LBCas12a(H759A)。
与本发明的构建体一起使用染色质调节肽的精确编辑结果提供在图36中。相比于没有任何另外的融合物的构建体(例如，RT(5M)-LbCas12a H759A)而言，所述构建体中的许多未导致精确编辑活性的增加。用tagRNA中的两种(tagRNA5和tagRNA6)，对于HN1-RT(5M)-LbCas12a(H759A)-HB1观察到精确编辑活性的轻微增加。
实施例16.同时对于本发明的构建体的非模板链产生切口的评价
在基因组编辑事件(包括例如碱基编辑、先导编辑(Prime editing)和REDRAW)期间的中间体可以是错配的DNA双链体，其中DNA的一条链已经由酶进行了编辑(所希望的编辑)，而相对的链包含野生型序列。朝向产生所希望的编辑的此类错配的分解对于确保所希望的编辑在细胞中成为持久的来说可以是重要的。
对于包含所述编辑的链的相对的DNA链产生切口被认为促进在细胞中通过利用错配修复(MMR)来使得所述编辑成为持久的过程。在真核生物中，MMR通过鉴定包含切口的DNA链(其暗示新合成的链，因此“可能”包含“错误”)来分解碱基错配并去除那条链，并且再合成一条完全互补的DNA。这样，去除包含野生型序列的DNA，并且制备与包含所希望的编辑的DNA完全互补的新DNA。该方法已经用于改善碱基编辑器(Komor等人,Nature 533:420-424(2016))和先导编辑器(Anzalone等人,Nature 576:149-157(2019))的编辑效率。
我们寻求用本发明的构建体和方法(REDRAW)来评价它。在REDRAW中，所述编辑被包含在DNA的模板链(被crRNA杂交的DNA链)中。因此，我们想要确定在编辑过程期间在所述编辑的附近对于非模板链产生切口是否可以增加REDRAW的精确编辑效率。
Fu等人以前报道，在12-15之间的碱基位置中包含各种错配的crRNA可以导致Cas12a成为非模板链切口酶而不是充当核酸酶(TTTV PAM被标示为位置-4、-3、-2、-1)(Fu等人,Nat Microbiol.2019May,4(5):888-897)。为了确定这样的方法在增加用本发明的方法和构建体所取得的精确编辑中是否可能是有效的，制备了包含各种错配的crRNA(与延伸型指导核酸即tagRNA形成对比)并且与本发明的构建体一起进行使用以编辑靶DNA。结果显示在表18中。表18.用三种不同的tagRNA的精确编辑效率(％)。在该实验中，与tagRNA15、tagRNA16或tagRNA17一起使用RT(5M)-LbCas12a H759A融合物。在该实施例中所使用的crRNA中的间隔子的互补性/错配描述在下面。
相比于其中未使用产生切口的crRNA的处理而言，在位置12-15处包含单、双或三错配的crRNA导致编辑效率的增加。综合考虑，包含合适错配的crRNA(除了tagRNA外)的同时表达可以用于在非模板链上诱导切口，并且由此增加本发明的方法的精确编辑效率。
上述内容举例说明了本发明，而不应被解释为限制了本发明。本发明由下面的权利要求来定义，其中在那里包括这些权利要求的等价物。
序列表
<110> Pairwise Plants Services, Inc.
Kim, Yongjoo
Watts, Joseph M.
Hummel, Aaron
Lawit, Shai J.
Schwark, David
<120> 用于等位基因的RNA编码的DNA替换的组合物和方法
<130> 1499.57.WO
<150> US 63/110,386
<151> 2020-11-06
<160> 215
<170> PatentIn version 3.5
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<211> 1228
<212> PRT
<213> Unknown
<220>
<223> 毛螺菌科（Lachnospiraceae）细菌
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1145 1150 1155
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1160 1165 1170
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1190 1195 1200
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1205 1210 1215
Ile Arg Ser Val Leu Gln Met Arg Asn Ser Asn Ala Ala Thr Gly
1220 1225 1230
Glu Asp Tyr Ile Asn Ser Pro Val Arg Asp Leu Asn Gly Val Cys
1235 1240 1245
Phe Asp Ser Arg Phe Gln Asn Pro Glu Trp Pro Met Asp Ala Asp
1250 1255 1260
Ala Asn Gly Ala Tyr His Ile Ala Leu Lys Gly Gln Leu Leu Leu
1265 1270 1275
Asn His Leu Lys Glu Ser Lys Asp Leu Lys Leu Gln Asn Gly Ile
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Ser Asn Gln Asp Trp Leu Ala Tyr Ile Gln Glu Leu Arg Asn
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<211> 1241
<212> PRT
<213> 蛋白质破裂丁酸弧菌（Butyrivibrio proteoclasticus）
<400> 3
Met Leu Leu Tyr Glu Asn Tyr Thr Lys Arg Asn Gln Ile Thr Lys Ser
1 5 10 15
Leu Arg Leu Glu Leu Arg Pro Gln Gly Lys Thr Leu Arg Asn Ile Lys
20 25 30
Glu Leu Asn Leu Leu Glu Gln Asp Lys Ala Ile Tyr Ala Leu Leu Glu
35 40 45
Arg Leu Lys Pro Val Ile Asp Glu Gly Ile Lys Asp Ile Ala Arg Asp
50 55 60
Thr Leu Lys Asn Cys Glu Leu Ser Phe Glu Lys Leu Tyr Glu His Phe
65 70 75 80
Leu Ser Gly Asp Lys Lys Ala Tyr Ala Lys Glu Ser Glu Arg Leu Lys
85 90 95
Lys Glu Ile Val Lys Thr Leu Ile Lys Asn Leu Pro Glu Gly Ile Gly
100 105 110
Lys Ile Ser Glu Ile Asn Ser Ala Lys Tyr Leu Asn Gly Val Leu Tyr
115 120 125
Asp Phe Ile Asp Lys Thr His Lys Asp Ser Glu Glu Lys Gln Asn Ile
130 135 140
Leu Ser Asp Ile Leu Glu Thr Lys Gly Tyr Leu Ala Leu Phe Ser Lys
145 150 155 160
Phe Leu Thr Ser Arg Ile Thr Thr Leu Glu Gln Ser Met Pro Lys Arg
165 170 175
Val Ile Glu Asn Phe Glu Ile Tyr Ala Ala Asn Ile Pro Lys Met Gln
180 185 190
Asp Ala Leu Glu Arg Gly Ala Val Ser Phe Ala Ile Glu Tyr Glu Ser
195 200 205
Ile Cys Ser Val Asp Tyr Tyr Asn Gln Ile Leu Ser Gln Glu Asp Ile
210 215 220
Asp Ser Tyr Asn Arg Leu Ile Ser Gly Ile Met Asp Glu Asp Gly Ala
225 230 235 240
Lys Glu Lys Gly Ile Asn Gln Thr Ile Ser Glu Lys Asn Ile Lys Ile
245 250 255
Lys Ser Glu His Leu Glu Glu Lys Pro Phe Arg Ile Leu Lys Gln Leu
260 265 270
His Lys Gln Ile Leu Glu Glu Arg Glu Lys Ala Phe Thr Ile Asp His
275 280 285
Ile Asp Ser Asp Glu Glu Val Val Gln Val Thr Lys Glu Ala Phe Glu
290 295 300
Gln Thr Lys Glu Gln Trp Glu Asn Ile Lys Lys Ile Asn Gly Phe Tyr
305 310 315 320
Ala Lys Asp Pro Gly Asp Ile Thr Leu Phe Ile Val Val Gly Pro Asn
325 330 335
Gln Thr His Val Leu Ser Gln Leu Ile Tyr Gly Glu His Asp Arg Ile
340 345 350
Arg Leu Leu Leu Glu Glu Tyr Glu Lys Asn Thr Leu Glu Val Leu Pro
355 360 365
Arg Arg Thr Lys Ser Glu Asp Ala Arg Tyr Asp Lys Phe Val Asn Ala
370 375 380
Val Pro Lys Lys Val Ala Lys Glu Ser His Thr Phe Asp Gly Leu Gln
385 390 395 400
Lys Met Thr Gly Asp Asp Arg Leu Phe Ile Leu Tyr Arg Asp Glu Leu
405 410 415
Ala Arg Asn Tyr Met Arg Ile Lys Glu Ala Tyr Gly Thr Phe Glu Arg
420 425 430
Asp Ile Leu Lys Ser Arg Arg Gly Ile Lys Gly Asn Arg Asp Val Gln
435 440 445
Glu Ser Leu Val Ser Phe Tyr Asp Glu Leu Thr Lys Phe Arg Ser Ala
450 455 460
Leu Arg Ile Ile Asn Ser Gly Asn Asp Glu Lys Ala Asp Pro Ile Phe
465 470 475 480
Tyr Asn Thr Phe Asp Gly Ile Phe Glu Lys Ala Asn Arg Thr Tyr Lys
485 490 495
Ala Glu Asn Leu Cys Arg Asn Tyr Val Thr Lys Ser Pro Ala Asp Asp
500 505 510
Ala Arg Ile Met Ala Ser Cys Leu Gly Thr Pro Ala Arg Leu Arg Thr
515 520 525
His Trp Trp Asn Gly Glu Glu Asn Phe Ala Ile Asn Asp Val Ala Met
530 535 540
Ile Arg Arg Gly Asp Glu Tyr Tyr Tyr Phe Val Leu Thr Pro Asp Val
545 550 555 560
Lys Pro Val Asp Leu Lys Thr Lys Asp Glu Thr Asp Ala Gln Ile Phe
565 570 575
Val Gln Arg Lys Gly Ala Lys Ser Phe Leu Gly Leu Pro Lys Ala Leu
580 585 590
Phe Lys Cys Ile Leu Glu Pro Tyr Phe Glu Ser Pro Glu His Lys Asn
595 600 605
Asp Lys Asn Cys Val Ile Glu Glu Tyr Val Ser Lys Pro Leu Thr Ile
610 615 620
Asp Arg Arg Ala Tyr Asp Ile Phe Lys Asn Gly Thr Phe Lys Lys Thr
625 630 635 640
Asn Ile Gly Ile Asp Gly Leu Thr Glu Glu Lys Phe Lys Asp Asp Cys
645 650 655
Arg Tyr Leu Ile Asp Val Tyr Lys Glu Phe Ile Ala Val Tyr Thr Arg
660 665 670
Tyr Ser Cys Phe Asn Met Ser Gly Leu Lys Arg Ala Asp Glu Tyr Asn
675 680 685
Asp Ile Gly Glu Phe Phe Ser Asp Val Asp Thr Arg Leu Cys Thr Met
690 695 700
Glu Trp Ile Pro Val Ser Phe Glu Arg Ile Asn Asp Met Val Asp Lys
705 710 715 720
Lys Glu Gly Leu Leu Phe Leu Val Arg Ser Met Phe Leu Tyr Asn Arg
725 730 735
Pro Arg Lys Pro Tyr Glu Arg Thr Phe Ile Gln Leu Phe Ser Asp Ser
740 745 750
Asn Met Glu His Thr Ser Met Leu Leu Asn Ser Arg Ala Met Ile Gln
755 760 765
Tyr Arg Ala Ala Ser Leu Pro Arg Arg Val Thr His Lys Lys Gly Ser
770 775 780
Ile Leu Val Ala Leu Arg Asp Ser Asn Gly Glu His Ile Pro Met His
785 790 795 800
Ile Arg Glu Ala Ile Tyr Lys Met Lys Asn Asn Phe Asp Ile Ser Ser
805 810 815
Glu Asp Phe Ile Met Ala Lys Ala Tyr Leu Ala Glu His Asp Val Ala
820 825 830
Ile Lys Lys Ala Asn Glu Asp Ile Ile Arg Asn Arg Arg Tyr Thr Glu
835 840 845
Asp Lys Phe Phe Leu Ser Leu Ser Tyr Thr Lys Asn Ala Asp Ile Ser
850 855 860
Ala Arg Thr Leu Asp Tyr Ile Asn Asp Lys Val Glu Glu Asp Thr Gln
865 870 875 880
Asp Ser Arg Met Ala Val Ile Val Thr Arg Asn Leu Lys Asp Leu Thr
885 890 895
Tyr Val Ala Val Val Asp Glu Lys Asn Asn Val Leu Glu Glu Lys Ser
900 905 910
Leu Asn Glu Ile Asp Gly Val Asn Tyr Arg Glu Leu Leu Lys Glu Arg
915 920 925
Thr Lys Ile Lys Tyr His Asp Lys Thr Arg Leu Trp Gln Tyr Asp Val
930 935 940
Ser Ser Lys Gly Leu Lys Glu Ala Tyr Val Glu Leu Ala Val Thr Gln
945 950 955 960
Ile Ser Lys Leu Ala Thr Lys Tyr Asn Ala Val Val Val Val Glu Ser
965 970 975
Met Ser Ser Thr Phe Lys Asp Lys Phe Ser Phe Leu Asp Glu Gln Ile
980 985 990
Phe Lys Ala Phe Glu Ala Arg Leu Cys Ala Arg Met Ser Asp Leu Ser
995 1000 1005
Phe Asn Thr Ile Lys Glu Gly Glu Ala Gly Ser Ile Ser Asn Pro
1010 1015 1020
Ile Gln Val Ser Asn Asn Asn Gly Asn Ser Tyr Gln Asp Gly Val
1025 1030 1035
Ile Tyr Phe Leu Asn Asn Ala Tyr Thr Arg Thr Leu Cys Pro Asp
1040 1045 1050
Thr Gly Phe Val Asp Val Phe Asp Lys Thr Arg Leu Ile Thr Met
1055 1060 1065
Gln Ser Lys Arg Gln Phe Phe Ala Lys Met Lys Asp Ile Arg Ile
1070 1075 1080
Asp Asp Gly Glu Met Leu Phe Thr Phe Asn Leu Glu Glu Tyr Pro
1085 1090 1095
Thr Lys Arg Leu Leu Asp Arg Lys Glu Trp Thr Val Lys Ile Ala
1100 1105 1110
Gly Asp Gly Ser Tyr Phe Asp Lys Asp Lys Gly Glu Tyr Val Tyr
1115 1120 1125
Val Asn Asp Ile Val Arg Glu Gln Ile Ile Pro Ala Leu Leu Glu
1130 1135 1140
Asp Lys Ala Val Phe Asp Gly Asn Met Ala Glu Lys Phe Leu Asp
1145 1150 1155
Lys Thr Ala Ile Ser Gly Lys Ser Val Glu Leu Ile Tyr Lys Trp
1160 1165 1170
Phe Ala Asn Ala Leu Tyr Gly Ile Ile Thr Lys Lys Asp Gly Glu
1175 1180 1185
Lys Ile Tyr Arg Ser Pro Ile Thr Gly Thr Glu Ile Asp Val Ser
1190 1195 1200
Lys Asn Thr Thr Tyr Asn Phe Gly Lys Lys Phe Met Phe Lys Gln
1205 1210 1215
Glu Tyr Arg Gly Asp Gly Asp Phe Leu Asp Ala Phe Leu Asn Tyr
1220 1225 1230
Met Gln Ala Gln Asp Ile Ala Val
1235 1240
<210> 4
<211> 1238
<212> PRT
<213> 候选的白蚁甲烷枝原体（Candidatus Methanoplasma termitum）
<400> 4
Met Asn Asn Tyr Asp Glu Phe Thr Lys Leu Tyr Pro Ile Gln Lys Thr
1 5 10 15
Ile Arg Phe Glu Leu Lys Pro Gln Gly Arg Thr Met Glu His Leu Glu
20 25 30
Thr Phe Asn Phe Phe Glu Glu Asp Arg Asp Arg Ala Glu Lys Tyr Lys
35 40 45
Ile Leu Lys Glu Ala Ile Asp Glu Tyr His Lys Lys Phe Ile Asp Glu
50 55 60
His Leu Thr Asn Met Ser Leu Asp Trp Asn Ser Leu Lys Gln Ile Ser
65 70 75 80
Glu Lys Tyr Tyr Lys Ser Arg Glu Glu Lys Asp Lys Lys Val Phe Leu
85 90 95
Ser Glu Gln Lys Arg Met Arg Gln Glu Ile Val Ser Glu Phe Lys Lys
100 105 110
Asp Asp Arg Phe Lys Asp Leu Phe Ser Lys Lys Leu Phe Ser Glu Leu
115 120 125
Leu Lys Glu Glu Ile Tyr Lys Lys Gly Asn His Gln Glu Ile Asp Ala
130 135 140
Leu Lys Ser Phe Asp Lys Phe Ser Gly Tyr Phe Ile Gly Leu His Glu
145 150 155 160
Asn Arg Lys Asn Met Tyr Ser Asp Gly Asp Glu Ile Thr Ala Ile Ser
165 170 175
Asn Arg Ile Val Asn Glu Asn Phe Pro Lys Phe Leu Asp Asn Leu Gln
180 185 190
Lys Tyr Gln Glu Ala Arg Lys Lys Tyr Pro Glu Trp Ile Ile Lys Ala
195 200 205
Glu Ser Ala Leu Val Ala His Asn Ile Lys Met Asp Ile Val Phe Ser
210 215 220
Leu Glu Tyr Phe Asn Lys Val Leu Asn Gln Glu Gly Ile Gln Arg Tyr
225 230 235 240
Asn Leu Ala Leu Gly Gly Tyr Val Thr Lys Ser Gly Glu Lys Met Met
245 250 255
Gly Leu Asn Asp Ala Leu Asn Leu Ala His Gln Ser Glu Lys Ser Ser
260 265 270
Lys Gly Arg Ile His Met Thr Pro Leu Phe Lys Gln Ile Leu Ser Glu
275 280 285
Lys Glu Ser Phe Ser Tyr Ile Pro Asp Val Phe Thr Glu Asp Ser Gln
290 295 300
Leu Leu Pro Ser Ile Gly Gly Phe Phe Ala Gln Ile Glu Asn Asp Lys
305 310 315 320
Asp Gly Asn Ile Phe Asp Arg Ala Leu Glu Leu Ile Ser Ser Tyr Ala
325 330 335
Glu Tyr Asp Thr Glu Arg Ile Tyr Ile Arg Gln Ala Asp Ile Asn Arg
340 345 350
Val Ser Asn Val Ile Phe Gly Glu Trp Gly Thr Leu Gly Gly Leu Met
355 360 365
Arg Glu Tyr Lys Ala Asp Ser Ile Asn Asp Ile Asn Leu Glu Arg Thr
370 375 380
Cys Lys Lys Val Asp Lys Trp Leu Asp Ser Lys Glu Phe Ala Leu Ser
385 390 395 400
Asp Val Leu Glu Ala Ile Asp Arg Thr Gly Asn Asn Asp Ala Phe Asn
405 410 415
Glu Tyr Ile Ser Lys Met Arg Thr Ala Arg Glu Lys Ile Asp Ala Ala
420 425 430
Arg Lys Glu Met Lys Phe Ile Ser Glu Lys Ile Ser Gly Asp Glu Glu
435 440 445
Ser Ile His Ile Ile Lys Thr Leu Leu Asp Ser Val Gln Gln Phe Leu
450 455 460
His Phe Phe Asn Leu Phe Lys Ala Arg Gln Asp Ile Pro Leu Asp Gly
465 470 475 480
Ala Phe Tyr Ala Glu Phe Asp Glu Val His Ser Lys Leu Phe Ala Ile
485 490 495
Val Pro Leu Tyr Asn Lys Val Arg Asn Tyr Leu Thr Lys Asn Asn Leu
500 505 510
Asn Thr Lys Lys Ile Lys Leu Asn Phe Lys Asn Pro Thr Leu Ala Asn
515 520 525
Gly Trp Asp Gln Asn Lys Val Tyr Asp Tyr Ala Ser Leu Ile Phe Leu
530 535 540
Arg Asp Gly Asn Tyr Tyr Leu Gly Ile Ile Asn Pro Lys Arg Lys Lys
545 550 555 560
Asn Ile Lys Phe Glu Gln Gly Ser Gly Asn Gly Pro Phe Tyr Arg Lys
565 570 575
Met Val Tyr Lys Gln Ile Pro Gly Pro Asn Lys Asn Leu Arg Pro Val
580 585 590
Phe Leu Thr Ser Thr Lys Gly Lys Lys Glu Tyr Lys Pro Ser Lys Glu
595 600 605
Ile Ile Glu Gly Tyr Glu Ala Asp Lys His Ile Arg Gly Asp Lys Phe
610 615 620
Asp Leu Asp Phe Cys His Lys Leu Ile Asp Phe Phe Lys Glu Ser Ile
625 630 635 640
Glu Lys His Lys Asp Trp Ser Lys Phe Asn Phe Tyr Phe Ser Pro Thr
645 650 655
Glu Ser Tyr Gly Asp Ile Ser Glu Phe Tyr Leu Asp Val Glu Lys Gln
660 665 670
Gly Tyr Arg Met His Phe Glu Asn Ile Ser Ala Glu Thr Ile Asp Glu
675 680 685
Tyr Val Glu Lys Gly Asp Leu Phe Leu Phe Gln Ile Tyr Asn Lys Asp
690 695 700
Phe Val Lys Ala Ala Thr Gly Lys Lys Asp Met His Thr Ile Tyr Trp
705 710 715 720
Asn Ala Ala Phe Ser Pro Glu Asn Leu Gln Asp Val Val Val Lys Leu
725 730 735
Asn Gly Glu Ala Glu Leu Phe Tyr Arg Asp Lys Ser Asp Ile Lys Glu
740 745 750
Ile Val His Arg Glu Gly Glu Ile Leu Val Asn Arg Thr Tyr Asn Gly
755 760 765
Arg Thr Pro Val Pro Asp Lys Ile His Lys Lys Leu Thr Asp Tyr His
770 775 780
Asn Gly Arg Thr Lys Asp Leu Gly Glu Ala Lys Glu Tyr Leu Asp Lys
785 790 795 800
Val Arg Tyr Phe Lys Ala His Tyr Asp Ile Thr Lys Asp Arg Arg Tyr
805 810 815
Leu Asn Asp Lys Ile Tyr Phe His Val Pro Leu Thr Leu Asn Phe Lys
820 825 830
Ala Asn Gly Lys Lys Asn Leu Asn Lys Met Val Ile Glu Lys Phe Leu
835 840 845
Ser Asp Glu Lys Ala His Ile Ile Gly Ile Asp Arg Gly Glu Arg Asn
850 855 860
Leu Leu Tyr Tyr Ser Ile Ile Asp Arg Ser Gly Lys Ile Ile Asp Gln
865 870 875 880
Gln Ser Leu Asn Val Ile Asp Gly Phe Asp Tyr Arg Glu Lys Leu Asn
885 890 895
Gln Arg Glu Ile Glu Met Lys Asp Ala Arg Gln Ser Trp Asn Ala Ile
900 905 910
Gly Lys Ile Lys Asp Leu Lys Glu Gly Tyr Leu Ser Lys Ala Val His
915 920 925
Glu Ile Thr Lys Met Ala Ile Gln Tyr Asn Ala Ile Val Val Met Glu
930 935 940
Glu Leu Asn Tyr Gly Phe Lys Arg Gly Arg Phe Lys Val Glu Lys Gln
945 950 955 960
Ile Tyr Gln Lys Phe Glu Asn Met Leu Ile Asp Lys Met Asn Tyr Leu
965 970 975
Val Phe Lys Asp Ala Pro Asp Glu Ser Pro Gly Gly Val Leu Asn Ala
980 985 990
Tyr Gln Leu Thr Asn Pro Leu Glu Ser Phe Ala Lys Leu Gly Lys Gln
995 1000 1005
Thr Gly Ile Leu Phe Tyr Val Pro Ala Ala Tyr Thr Ser Lys Ile
1010 1015 1020
Asp Pro Thr Thr Gly Phe Val Asn Leu Phe Asn Thr Ser Ser Lys
1025 1030 1035
Thr Asn Ala Gln Glu Arg Lys Glu Phe Leu Gln Lys Phe Glu Ser
1040 1045 1050
Ile Ser Tyr Ser Ala Lys Asp Gly Gly Ile Phe Ala Phe Ala Phe
1055 1060 1065
Asp Tyr Arg Lys Phe Gly Thr Ser Lys Thr Asp His Lys Asn Val
1070 1075 1080
Trp Thr Ala Tyr Thr Asn Gly Glu Arg Met Arg Tyr Ile Lys Glu
1085 1090 1095
Lys Lys Arg Asn Glu Leu Phe Asp Pro Ser Lys Glu Ile Lys Glu
1100 1105 1110
Ala Leu Thr Ser Ser Gly Ile Lys Tyr Asp Gly Gly Gln Asn Ile
1115 1120 1125
Leu Pro Asp Ile Leu Arg Ser Asn Asn Asn Gly Leu Ile Tyr Thr
1130 1135 1140
Met Tyr Ser Ser Phe Ile Ala Ala Ile Gln Met Arg Val Tyr Asp
1145 1150 1155
Gly Lys Glu Asp Tyr Ile Ile Ser Pro Ile Lys Asn Ser Lys Gly
1160 1165 1170
Glu Phe Phe Arg Thr Asp Pro Lys Arg Arg Glu Leu Pro Ile Asp
1175 1180 1185
Ala Asp Ala Asn Gly Ala Tyr Asn Ile Ala Leu Arg Gly Glu Leu
1190 1195 1200
Thr Met Arg Ala Ile Ala Glu Lys Phe Asp Pro Asp Ser Glu Lys
1205 1210 1215
Met Ala Lys Leu Glu Leu Lys His Lys Asp Trp Phe Glu Phe Met
1220 1225 1230
Gln Thr Arg Gly Asp
1235
<210> 5
<211> 1281
<212> PRT
<213> 挑剔真杆菌（Eubacterium eligens）
<400> 5
Met Asn Gly Asn Arg Ser Ile Val Tyr Arg Glu Phe Val Gly Val Ile
1 5 10 15
Pro Val Ala Lys Thr Leu Arg Asn Glu Leu Arg Pro Val Gly His Thr
20 25 30
Gln Glu His Ile Ile Gln Asn Gly Leu Ile Gln Glu Asp Glu Leu Arg
35 40 45
Gln Glu Lys Ser Thr Glu Leu Lys Asn Ile Met Asp Asp Tyr Tyr Arg
50 55 60
Glu Tyr Ile Asp Lys Ser Leu Ser Gly Val Thr Asp Leu Asp Phe Thr
65 70 75 80
Leu Leu Phe Glu Leu Met Asn Leu Val Gln Ser Ser Pro Ser Lys Asp
85 90 95
Asn Lys Lys Ala Leu Glu Lys Glu Gln Ser Lys Met Arg Glu Gln Ile
100 105 110
Cys Thr His Leu Gln Ser Asp Ser Asn Tyr Lys Asn Ile Phe Asn Ala
115 120 125
Lys Leu Leu Lys Glu Ile Leu Pro Asp Phe Ile Lys Asn Tyr Asn Gln
130 135 140
Tyr Asp Val Lys Asp Lys Ala Gly Lys Leu Glu Thr Leu Ala Leu Phe
145 150 155 160
Asn Gly Phe Ser Thr Tyr Phe Thr Asp Phe Phe Glu Lys Arg Lys Asn
165 170 175
Val Phe Thr Lys Glu Ala Val Ser Thr Ser Ile Ala Tyr Arg Ile Val
180 185 190
His Glu Asn Ser Leu Ile Phe Leu Ala Asn Met Thr Ser Tyr Lys Lys
195 200 205
Ile Ser Glu Lys Ala Leu Asp Glu Ile Glu Val Ile Glu Lys Asn Asn
210 215 220
Gln Asp Lys Met Gly Asp Trp Glu Leu Asn Gln Ile Phe Asn Pro Asp
225 230 235 240
Phe Tyr Asn Met Val Leu Ile Gln Ser Gly Ile Asp Phe Tyr Asn Glu
245 250 255
Ile Cys Gly Val Val Asn Ala His Met Asn Leu Tyr Cys Gln Gln Thr
260 265 270
Lys Asn Asn Tyr Asn Leu Phe Lys Met Arg Lys Leu His Lys Gln Ile
275 280 285
Leu Ala Tyr Thr Ser Thr Ser Phe Glu Val Pro Lys Met Phe Glu Asp
290 295 300
Asp Met Ser Val Tyr Asn Ala Val Asn Ala Phe Ile Asp Glu Thr Glu
305 310 315 320
Lys Gly Asn Ile Ile Gly Lys Leu Lys Asp Ile Val Asn Lys Tyr Asp
325 330 335
Glu Leu Asp Glu Lys Arg Ile Tyr Ile Ser Lys Asp Phe Tyr Glu Thr
340 345 350
Leu Ser Cys Phe Met Ser Gly Asn Trp Asn Leu Ile Thr Gly Cys Val
355 360 365
Glu Asn Phe Tyr Asp Glu Asn Ile His Ala Lys Gly Lys Ser Lys Glu
370 375 380
Glu Lys Val Lys Lys Ala Val Lys Glu Asp Lys Tyr Lys Ser Ile Asn
385 390 395 400
Asp Val Asn Asp Leu Val Glu Lys Tyr Ile Asp Glu Lys Glu Arg Asn
405 410 415
Glu Phe Lys Asn Ser Asn Ala Lys Gln Tyr Ile Arg Glu Ile Ser Asn
420 425 430
Ile Ile Thr Asp Thr Glu Thr Ala His Leu Glu Tyr Asp Asp His Ile
435 440 445
Ser Leu Ile Glu Ser Glu Glu Lys Ala Asp Glu Met Lys Lys Arg Leu
450 455 460
Asp Met Tyr Met Asn Met Tyr His Trp Ala Lys Ala Phe Ile Val Asp
465 470 475 480
Glu Val Leu Asp Arg Asp Glu Met Phe Tyr Ser Asp Ile Asp Asp Ile
485 490 495
Tyr Asn Ile Leu Glu Asn Ile Val Pro Leu Tyr Asn Arg Val Arg Asn
500 505 510
Tyr Val Thr Gln Lys Pro Tyr Asn Ser Lys Lys Ile Lys Leu Asn Phe
515 520 525
Gln Ser Pro Thr Leu Ala Asn Gly Trp Ser Gln Ser Lys Glu Phe Asp
530 535 540
Asn Asn Ala Ile Ile Leu Ile Arg Asp Asn Lys Tyr Tyr Leu Ala Ile
545 550 555 560
Phe Asn Ala Lys Asn Lys Pro Asp Lys Lys Ile Ile Gln Gly Asn Ser
565 570 575
Asp Lys Lys Asn Asp Asn Asp Tyr Lys Lys Met Val Tyr Asn Leu Leu
580 585 590
Pro Gly Ala Asn Lys Met Leu Pro Lys Val Phe Leu Ser Lys Lys Gly
595 600 605
Ile Glu Thr Phe Lys Pro Ser Asp Tyr Ile Ile Ser Gly Tyr Asn Ala
610 615 620
His Lys His Ile Lys Thr Ser Glu Asn Phe Asp Ile Ser Phe Cys Arg
625 630 635 640
Asp Leu Ile Asp Tyr Phe Lys Asn Ser Ile Glu Lys His Ala Glu Trp
645 650 655
Arg Lys Tyr Glu Phe Lys Phe Ser Ala Thr Asp Ser Tyr Ser Asp Ile
660 665 670
Ser Glu Phe Tyr Arg Glu Val Glu Met Gln Gly Tyr Arg Ile Asp Trp
675 680 685
Thr Tyr Ile Ser Glu Ala Asp Ile Asn Lys Leu Asp Glu Glu Gly Lys
690 695 700
Ile Tyr Leu Phe Gln Ile Tyr Asn Lys Asp Phe Ala Glu Asn Ser Thr
705 710 715 720
Gly Lys Glu Asn Leu His Thr Met Tyr Phe Lys Asn Ile Phe Ser Glu
725 730 735
Glu Asn Leu Asp Lys Ile Ile Lys Leu Asn Gly Gln Ala Glu Leu Phe
740 745 750
Tyr Arg Arg Ala Ser Val Lys Asn Pro Val Lys His Lys Lys Asp Ser
755 760 765
Val Leu Val Asn Lys Thr Tyr Lys Asn Gln Leu Asp Asn Gly Asp Val
770 775 780
Val Arg Ile Pro Ile Pro Asp Asp Ile Tyr Asn Glu Ile Tyr Lys Met
785 790 795 800
Tyr Asn Gly Tyr Ile Lys Glu Ser Asp Leu Ser Glu Ala Ala Lys Glu
805 810 815
Tyr Leu Asp Lys Val Glu Val Arg Thr Ala Gln Lys Asp Ile Val Lys
820 825 830
Asp Tyr Arg Tyr Thr Val Asp Lys Tyr Phe Ile His Thr Pro Ile Thr
835 840 845
Ile Asn Tyr Lys Val Thr Ala Arg Asn Asn Val Asn Asp Met Val Val
850 855 860
Lys Tyr Ile Ala Gln Asn Asp Asp Ile His Val Ile Gly Ile Asp Arg
865 870 875 880
Gly Glu Arg Asn Leu Ile Tyr Ile Ser Val Ile Asp Ser His Gly Asn
885 890 895
Ile Val Lys Gln Lys Ser Tyr Asn Ile Leu Asn Asn Tyr Asp Tyr Lys
900 905 910
Lys Lys Leu Val Glu Lys Glu Lys Thr Arg Glu Tyr Ala Arg Lys Asn
915 920 925
Trp Lys Ser Ile Gly Asn Ile Lys Glu Leu Lys Glu Gly Tyr Ile Ser
930 935 940
Gly Val Val His Glu Ile Ala Met Leu Ile Val Glu Tyr Asn Ala Ile
945 950 955 960
Ile Ala Met Glu Asp Leu Asn Tyr Gly Phe Lys Arg Gly Arg Phe Lys
965 970 975
Val Glu Arg Gln Val Tyr Gln Lys Phe Glu Ser Met Leu Ile Asn Lys
980 985 990
Leu Asn Tyr Phe Ala Ser Lys Glu Lys Ser Val Asp Glu Pro Gly Gly
995 1000 1005
Leu Leu Lys Gly Tyr Gln Leu Thr Tyr Val Pro Asp Asn Ile Lys
1010 1015 1020
Asn Leu Gly Lys Gln Cys Gly Val Ile Phe Tyr Val Pro Ala Ala
1025 1030 1035
Phe Thr Ser Lys Ile Asp Pro Ser Thr Gly Phe Ile Ser Ala Phe
1040 1045 1050
Asn Phe Lys Ser Ile Ser Thr Asn Ala Ser Arg Lys Gln Phe Phe
1055 1060 1065
Met Gln Phe Asp Glu Ile Arg Tyr Cys Ala Glu Lys Asp Met Phe
1070 1075 1080
Ser Phe Gly Phe Asp Tyr Asn Asn Phe Asp Thr Tyr Asn Ile Thr
1085 1090 1095
Met Gly Lys Thr Gln Trp Thr Val Tyr Thr Asn Gly Glu Arg Leu
1100 1105 1110
Gln Ser Glu Phe Asn Asn Ala Arg Arg Thr Gly Lys Thr Lys Ser
1115 1120 1125
Ile Asn Leu Thr Glu Thr Ile Lys Leu Leu Leu Glu Asp Asn Glu
1130 1135 1140
Ile Asn Tyr Ala Asp Gly His Asp Ile Arg Ile Asp Met Glu Lys
1145 1150 1155
Met Asp Glu Asp Lys Lys Ser Glu Phe Phe Ala Gln Leu Leu Ser
1160 1165 1170
Leu Tyr Lys Leu Thr Val Gln Met Arg Asn Ser Tyr Thr Glu Ala
1175 1180 1185
Glu Glu Gln Glu Asn Gly Ile Ser Tyr Asp Lys Ile Ile Ser Pro
1190 1195 1200
Val Ile Asn Asp Glu Gly Glu Phe Phe Asp Ser Asp Asn Tyr Lys
1205 1210 1215
Glu Ser Asp Asp Lys Glu Cys Lys Met Pro Lys Asp Ala Asp Ala
1220 1225 1230
Asn Gly Ala Tyr Cys Ile Ala Leu Lys Gly Leu Tyr Glu Val Leu
1235 1240 1245
Lys Ile Lys Ser Glu Trp Thr Glu Asp Gly Phe Asp Arg Asn Cys
1250 1255 1260
Leu Lys Leu Pro His Ala Glu Trp Leu Asp Phe Ile Gln Asn Lys
1265 1270 1275
Arg Tyr Glu
1280
<210> 6
<211> 1300
<212> PRT
<213> 新凶手弗朗西丝氏菌（Francisella novicida）
<400> 6
Met Ser Ile Tyr Gln Glu Phe Val Asn Lys Tyr Ser Leu Ser Lys Thr
1 5 10 15
Leu Arg Phe Glu Leu Ile Pro Gln Gly Lys Thr Leu Glu Asn Ile Lys
20 25 30
Ala Arg Gly Leu Ile Leu Asp Asp Glu Lys Arg Ala Lys Asp Tyr Lys
35 40 45
Lys Ala Lys Gln Ile Ile Asp Lys Tyr His Gln Phe Phe Ile Glu Glu
50 55 60
Ile Leu Ser Ser Val Cys Ile Ser Glu Asp Leu Leu Gln Asn Tyr Ser
65 70 75 80
Asp Val Tyr Phe Lys Leu Lys Lys Ser Asp Asp Asp Asn Leu Gln Lys
85 90 95
Asp Phe Lys Ser Ala Lys Asp Thr Ile Lys Lys Gln Ile Ser Glu Tyr
100 105 110
Ile Lys Asp Ser Glu Lys Phe Lys Asn Leu Phe Asn Gln Asn Leu Ile
115 120 125
Asp Ala Lys Lys Gly Gln Glu Ser Asp Leu Ile Leu Trp Leu Lys Gln
130 135 140
Ser Lys Asp Asn Gly Ile Glu Leu Phe Lys Ala Asn Ser Asp Ile Thr
145 150 155 160
Asp Ile Asp Glu Ala Leu Glu Ile Ile Lys Ser Phe Lys Gly Trp Thr
165 170 175
Thr Tyr Phe Lys Gly Phe His Glu Asn Arg Lys Val Asn Tyr Ser Ser
180 185 190
Asn Asp Ile Pro Thr Ser Ile Ile Tyr Arg Ile Val Asp Asp Asn Leu
195 200 205
Pro Lys Phe Leu Glu Asn Lys Ala Lys Tyr Glu Ser Leu Lys Asp Lys
210 215 220
Ala Pro Glu Ala Ile Asn Tyr Glu Gln Ile Lys Lys Asp Leu Ala Glu
225 230 235 240
Glu Leu Thr Phe Asp Ile Asp Tyr Lys Thr Ser Glu Val Asn Gln Arg
245 250 255
Val Phe Ser Leu Asp Glu Val Phe Glu Ile Ala Asn Phe Asn Asn Tyr
260 265 270
Leu Asn Gln Ser Gly Ile Thr Lys Phe Asn Thr Ile Ile Gly Gly Lys
275 280 285
Phe Val Asn Gly Glu Asn Thr Lys Arg Lys Gly Ile Asn Glu Tyr Ile
290 295 300
Asn Leu Tyr Ser Gln Gln Ile Asn Asp Lys Thr Leu Lys Lys Tyr Lys
305 310 315 320
Met Ser Val Leu Phe Lys Gln Ile Leu Ser Asp Thr Glu Ser Lys Ser
325 330 335
Phe Val Ile Asp Lys Leu Glu Asp Asp Ser Asp Val Val Thr Thr Met
340 345 350
Gln Ser Phe Tyr Glu Gln Ile Ala Ala Phe Lys Thr Val Glu Glu Lys
355 360 365
Ser Ile Lys Glu Thr Leu Ser Leu Leu Phe Asp Asp Leu Lys Ala Gln
370 375 380
Lys Leu Asp Leu Ser Lys Ile Tyr Phe Lys Asn Asp Lys Ser Leu Thr
385 390 395 400
Asp Leu Ser Gln Gln Val Phe Asp Asp Tyr Ser Val Ile Gly Thr Ala
405 410 415
Val Leu Glu Tyr Ile Thr Gln Gln Ile Ala Pro Lys Asn Leu Asp Asn
420 425 430
Pro Ser Lys Lys Glu Gln Glu Leu Ile Ala Lys Lys Thr Glu Lys Ala
435 440 445
Lys Tyr Leu Ser Leu Glu Thr Ile Lys Leu Ala Leu Glu Glu Phe Asn
450 455 460
Lys His Arg Asp Ile Asp Lys Gln Cys Arg Phe Glu Glu Ile Leu Ala
465 470 475 480
Asn Phe Ala Ala Ile Pro Met Ile Phe Asp Glu Ile Ala Gln Asn Lys
485 490 495
Asp Asn Leu Ala Gln Ile Ser Ile Lys Tyr Gln Asn Gln Gly Lys Lys
500 505 510
Asp Leu Leu Gln Ala Ser Ala Glu Asp Asp Val Lys Ala Ile Lys Asp
515 520 525
Leu Leu Asp Gln Thr Asn Asn Leu Leu His Lys Leu Lys Ile Phe His
530 535 540
Ile Ser Gln Ser Glu Asp Lys Ala Asn Ile Leu Asp Lys Asp Glu His
545 550 555 560
Phe Tyr Leu Val Phe Glu Glu Cys Tyr Phe Glu Leu Ala Asn Ile Val
565 570 575
Pro Leu Tyr Asn Lys Ile Arg Asn Tyr Ile Thr Gln Lys Pro Tyr Ser
580 585 590
Asp Glu Lys Phe Lys Leu Asn Phe Glu Asn Ser Thr Leu Ala Asn Gly
595 600 605
Trp Asp Lys Asn Lys Glu Pro Asp Asn Thr Ala Ile Leu Phe Ile Lys
610 615 620
Asp Asp Lys Tyr Tyr Leu Gly Val Met Asn Lys Lys Asn Asn Lys Ile
625 630 635 640
Phe Asp Asp Lys Ala Ile Lys Glu Asn Lys Gly Glu Gly Tyr Lys Lys
645 650 655
Ile Val Tyr Lys Leu Leu Pro Gly Ala Asn Lys Met Leu Pro Lys Val
660 665 670
Phe Phe Ser Ala Lys Ser Ile Lys Phe Tyr Asn Pro Ser Glu Asp Ile
675 680 685
Leu Arg Ile Arg Asn His Ser Thr His Thr Lys Asn Gly Ser Pro Gln
690 695 700
Lys Gly Tyr Glu Lys Phe Glu Phe Asn Ile Glu Asp Cys Arg Lys Phe
705 710 715 720
Ile Asp Phe Tyr Lys Gln Ser Ile Ser Lys His Pro Glu Trp Lys Asp
725 730 735
Phe Gly Phe Arg Phe Ser Asp Thr Gln Arg Tyr Asn Ser Ile Asp Glu
740 745 750
Phe Tyr Arg Glu Val Glu Asn Gln Gly Tyr Lys Leu Thr Phe Glu Asn
755 760 765
Ile Ser Glu Ser Tyr Ile Asp Ser Val Val Asn Gln Gly Lys Leu Tyr
770 775 780
Leu Phe Gln Ile Tyr Asn Lys Asp Phe Ser Ala Tyr Ser Lys Gly Arg
785 790 795 800
Pro Asn Leu His Thr Leu Tyr Trp Lys Ala Leu Phe Asp Glu Arg Asn
805 810 815
Leu Gln Asp Val Val Tyr Lys Leu Asn Gly Glu Ala Glu Leu Phe Tyr
820 825 830
Arg Lys Gln Ser Ile Pro Lys Lys Ile Thr His Pro Ala Lys Glu Ala
835 840 845
Ile Ala Asn Lys Asn Lys Asp Asn Pro Lys Lys Glu Ser Val Phe Glu
850 855 860
Tyr Asp Leu Ile Lys Asp Lys Arg Phe Thr Glu Asp Lys Phe Phe Phe
865 870 875 880
His Cys Pro Ile Thr Ile Asn Phe Lys Ser Ser Gly Ala Asn Lys Phe
885 890 895
Asn Asp Glu Ile Asn Leu Leu Leu Lys Glu Lys Ala Asn Asp Val His
900 905 910
Ile Leu Ser Ile Asp Arg Gly Glu Arg His Leu Ala Tyr Tyr Thr Leu
915 920 925
Val Asp Gly Lys Gly Asn Ile Ile Lys Gln Asp Thr Phe Asn Ile Ile
930 935 940
Gly Asn Asp Arg Met Lys Thr Asn Tyr His Asp Lys Leu Ala Ala Ile
945 950 955 960
Glu Lys Asp Arg Asp Ser Ala Arg Lys Asp Trp Lys Lys Ile Asn Asn
965 970 975
Ile Lys Glu Met Lys Glu Gly Tyr Leu Ser Gln Val Val His Glu Ile
980 985 990
Ala Lys Leu Val Ile Glu Tyr Asn Ala Ile Val Val Phe Glu Asp Leu
995 1000 1005
Asn Phe Gly Phe Lys Arg Gly Arg Phe Lys Val Glu Lys Gln Val
1010 1015 1020
Tyr Gln Lys Leu Glu Lys Met Leu Ile Glu Lys Leu Asn Tyr Leu
1025 1030 1035
Val Phe Lys Asp Asn Glu Phe Asp Lys Thr Gly Gly Val Leu Arg
1040 1045 1050
Ala Tyr Gln Leu Thr Ala Pro Phe Glu Thr Phe Lys Lys Met Gly
1055 1060 1065
Lys Gln Thr Gly Ile Ile Tyr Tyr Val Pro Ala Gly Phe Thr Ser
1070 1075 1080
Lys Ile Cys Pro Val Thr Gly Phe Val Asn Gln Leu Tyr Pro Lys
1085 1090 1095
Tyr Glu Ser Val Ser Lys Ser Gln Glu Phe Phe Ser Lys Phe Asp
1100 1105 1110
Lys Ile Cys Tyr Asn Leu Asp Lys Gly Tyr Phe Glu Phe Ser Phe
1115 1120 1125
Asp Tyr Lys Asn Phe Gly Asp Lys Ala Ala Lys Gly Lys Trp Thr
1130 1135 1140
Ile Ala Ser Phe Gly Ser Arg Leu Ile Asn Phe Arg Asn Ser Asp
1145 1150 1155
Lys Asn His Asn Trp Asp Thr Arg Glu Val Tyr Pro Thr Lys Glu
1160 1165 1170
Leu Glu Lys Leu Leu Lys Asp Tyr Ser Ile Glu Tyr Gly His Gly
1175 1180 1185
Glu Cys Ile Lys Ala Ala Ile Cys Gly Glu Ser Asp Lys Lys Phe
1190 1195 1200
Phe Ala Lys Leu Thr Ser Val Leu Asn Thr Ile Leu Gln Met Arg
1205 1210 1215
Asn Ser Lys Thr Gly Thr Glu Leu Asp Tyr Leu Ile Ser Pro Val
1220 1225 1230
Ala Asp Val Asn Gly Asn Phe Phe Asp Ser Arg Gln Ala Pro Lys
1235 1240 1245
Asn Met Pro Gln Asp Ala Asp Ala Asn Gly Ala Tyr His Ile Gly
1250 1255 1260
Leu Lys Gly Leu Met Leu Leu Gly Arg Ile Lys Asn Asn Gln Glu
1265 1270 1275
Gly Lys Lys Leu Asn Leu Val Ile Lys Asn Glu Glu Tyr Phe Glu
1280 1285 1290
Phe Val Gln Asn Arg Asn Asn
1295 1300
<210> 7
<211> 1206
<212> PRT
<213> Unknown
<220>
<223> 毛螺菌科细菌
<400> 7
Met Tyr Tyr Glu Ser Leu Thr Lys Gln Tyr Pro Val Ser Lys Thr Ile
1 5 10 15
Arg Asn Glu Leu Ile Pro Ile Gly Lys Thr Leu Asp Asn Ile Arg Gln
20 25 30
Asn Asn Ile Leu Glu Ser Asp Val Lys Arg Lys Gln Asn Tyr Glu His
35 40 45
Val Lys Gly Ile Leu Asp Glu Tyr His Lys Gln Leu Ile Asn Glu Ala
50 55 60
Leu Asp Asn Cys Thr Leu Pro Ser Leu Lys Ile Ala Ala Glu Ile Tyr
65 70 75 80
Leu Lys Asn Gln Lys Glu Val Ser Asp Arg Glu Asp Phe Asn Lys Thr
85 90 95
Gln Asp Leu Leu Arg Lys Glu Val Val Glu Lys Leu Lys Ala His Glu
100 105 110
Asn Phe Thr Lys Ile Gly Lys Lys Asp Ile Leu Asp Leu Leu Glu Lys
115 120 125
Leu Pro Ser Ile Ser Glu Asp Asp Tyr Asn Ala Leu Glu Ser Phe Arg
130 135 140
Asn Phe Tyr Thr Tyr Phe Thr Ser Tyr Asn Lys Val Arg Glu Asn Leu
145 150 155 160
Tyr Ser Asp Lys Glu Lys Ser Ser Thr Val Ala Tyr Arg Leu Ile Asn
165 170 175
Glu Asn Phe Pro Lys Phe Leu Asp Asn Val Lys Ser Tyr Arg Phe Val
180 185 190
Lys Thr Ala Gly Ile Leu Ala Asp Gly Leu Gly Glu Glu Glu Gln Asp
195 200 205
Ser Leu Phe Ile Val Glu Thr Phe Asn Lys Thr Leu Thr Gln Asp Gly
210 215 220
Ile Asp Thr Tyr Asn Ser Gln Val Gly Lys Ile Asn Ser Ser Ile Asn
225 230 235 240
Leu Tyr Asn Gln Lys Asn Gln Lys Ala Asn Gly Phe Arg Lys Ile Pro
245 250 255
Lys Met Lys Met Leu Tyr Lys Gln Ile Leu Ser Asp Arg Glu Glu Ser
260 265 270
Phe Ile Asp Glu Phe Gln Ser Asp Glu Val Leu Ile Asp Asn Val Glu
275 280 285
Ser Tyr Gly Ser Val Leu Ile Glu Ser Leu Lys Ser Ser Lys Val Ser
290 295 300
Ala Phe Phe Asp Ala Leu Arg Glu Ser Lys Gly Lys Asn Val Tyr Val
305 310 315 320
Lys Asn Asp Leu Ala Lys Thr Ala Met Ser Val Ile Val Phe Glu Asn
325 330 335
Trp Arg Thr Phe Asp Asp Leu Leu Asn Gln Glu Tyr Asp Leu Ala Asn
340 345 350
Glu Asn Lys Lys Lys Asp Asp Lys Tyr Phe Glu Lys Arg Gln Lys Glu
355 360 365
Leu Lys Lys Asn Lys Ser Tyr Ser Leu Glu His Leu Cys Asn Leu Ser
370 375 380
Glu Asp Ser Cys Asn Leu Ile Glu Asn Tyr Ile His Gln Ile Ser Asp
385 390 395 400
Asp Ile Glu Asn Ile Ile Ile Asn Asn Glu Thr Phe Leu Arg Ile Val
405 410 415
Ile Asn Glu His Asp Arg Ser Arg Lys Leu Ala Lys Asn Arg Lys Ala
420 425 430
Val Lys Ala Ile Lys Asp Phe Leu Asp Ser Ile Lys Val Leu Glu Arg
435 440 445
Glu Leu Lys Leu Ile Asn Ser Ser Gly Gln Glu Leu Glu Lys Asp Leu
450 455 460
Ile Val Tyr Ser Ala His Glu Glu Leu Leu Val Glu Leu Lys Gln Val
465 470 475 480
Asp Ser Leu Tyr Asn Met Thr Arg Asn Tyr Leu Thr Lys Lys Pro Phe
485 490 495
Ser Thr Glu Lys Val Lys Leu Asn Phe Asn Arg Ser Thr Leu Leu Asn
500 505 510
Gly Trp Asp Arg Asn Lys Glu Thr Asp Asn Leu Gly Val Leu Leu Leu
515 520 525
Lys Asp Gly Lys Tyr Tyr Leu Gly Ile Met Asn Thr Ser Ala Asn Lys
530 535 540
Ala Phe Val Asn Pro Pro Val Ala Lys Thr Glu Lys Val Phe Lys Lys
545 550 555 560
Val Asp Tyr Lys Leu Leu Pro Val Pro Asn Gln Met Leu Pro Lys Val
565 570 575
Phe Phe Ala Lys Ser Asn Ile Asp Phe Tyr Asn Pro Ser Ser Glu Ile
580 585 590
Tyr Ser Asn Tyr Lys Lys Gly Thr His Lys Lys Gly Asn Met Phe Ser
595 600 605
Leu Glu Asp Cys His Asn Leu Ile Asp Phe Phe Lys Glu Ser Ile Ser
610 615 620
Lys His Glu Asp Trp Ser Lys Phe Gly Phe Lys Phe Asp Thr Gln Ala
625 630 635 640
Ser Tyr Asn Asp Ile Ser Glu Phe Tyr Arg Glu Val Glu Lys Gln Gly
645 650 655
Tyr Lys Leu Thr Tyr Thr Asp Ile Asp Glu Thr Tyr Ile Asn Asp Leu
660 665 670
Ile Glu Arg Asn Glu Leu Tyr Leu Phe Gln Ile Tyr Asn Lys Asp Phe
675 680 685
Ser Met Tyr Ser Lys Gly Lys Leu Asn Leu His Thr Leu Tyr Phe Met
690 695 700
Met Leu Phe Asp Gln Arg Asn Ile Asp Asp Val Val Tyr Lys Leu Asn
705 710 715 720
Gly Glu Ala Glu Val Phe Tyr Arg Pro Ala Ser Ile Ser Glu Asp Glu
725 730 735
Leu Ile Ile His Lys Ala Gly Glu Glu Ile Lys Asn Lys Asn Pro Asn
740 745 750
Arg Ala Arg Thr Lys Glu Thr Ser Thr Phe Ser Tyr Asp Ile Val Lys
755 760 765
Asp Lys Arg Tyr Ser Lys Asp Lys Phe Thr Leu His Ile Pro Ile Thr
770 775 780
Met Asn Phe Gly Val Asp Glu Val Lys Arg Phe Asn Asp Ala Val Asn
785 790 795 800
Ser Ala Ile Arg Ile Asp Glu Asn Val Asn Val Ile Gly Ile Asp Arg
805 810 815
Gly Glu Arg Asn Leu Leu Tyr Val Val Val Ile Asp Ser Lys Gly Asn
820 825 830
Ile Leu Glu Gln Ile Ser Leu Asn Ser Ile Ile Asn Lys Glu Tyr Asp
835 840 845
Ile Glu Thr Asp Tyr His Ala Leu Leu Asp Glu Arg Glu Gly Gly Arg
850 855 860
Asp Lys Ala Arg Lys Asp Trp Asn Thr Val Glu Asn Ile Arg Asp Leu
865 870 875 880
Lys Ala Gly Leu Tyr Leu Gln Val Val Asn Val Val Ala Lys Leu Val
885 890 895
Leu Lys Tyr Asn Ala Ile Ile Cys Leu Glu Asp Leu Asn Phe Gly Phe
900 905 910
Lys Arg Gly Arg Gln Lys Val Glu Lys Gln Val Tyr Gln Lys Phe Glu
915 920 925
Lys Met Leu Ile Asp Lys Leu Asn Tyr Leu Val Ile Asp Lys Ser Arg
930 935 940
Glu Gln Thr Ser Pro Lys Glu Leu Gly Gly Ala Leu Asn Ala Leu Gln
945 950 955 960
Leu Thr Ser Lys Phe Lys Ser Phe Lys Glu Leu Gly Lys Gln Ser Gly
965 970 975
Val Ile Tyr Tyr Val Pro Ala Tyr Leu Thr Ser Lys Ile Asp Pro Thr
980 985 990
Thr Gly Phe Ala Asn Leu Phe Tyr Met Lys Cys Glu Asn Val Glu Lys
995 1000 1005
Ser Lys Arg Phe Phe Asp Gly Phe Asp Phe Ile Arg Phe Asn Ala
1010 1015 1020
Leu Glu Asn Val Phe Glu Phe Gly Phe Asp Tyr Arg Ser Phe Thr
1025 1030 1035
Gln Arg Ala Cys Gly Ile Asn Ser Lys Trp Thr Val Cys Thr Asn
1040 1045 1050
Gly Glu Arg Ile Ile Lys Tyr Arg Asn Pro Asp Lys Asn Asn Met
1055 1060 1065
Phe Asp Glu Lys Val Val Val Val Thr Asp Glu Met Lys Asn Leu
1070 1075 1080
Phe Glu Gln Tyr Lys Ile Pro Tyr Glu Asp Gly Arg Asn Val Lys
1085 1090 1095
Asp Met Ile Ile Ser Asn Glu Glu Ala Glu Phe Tyr Arg Arg Leu
1100 1105 1110
Tyr Arg Leu Leu Gln Gln Thr Leu Gln Met Arg Asn Ser Thr Ser
1115 1120 1125
Asp Gly Thr Arg Asp Tyr Ile Ile Ser Pro Val Lys Asn Lys Arg
1130 1135 1140
Glu Ala Tyr Phe Asn Ser Glu Leu Ser Asp Gly Ser Val Pro Lys
1145 1150 1155
Asp Ala Asp Ala Asn Gly Ala Tyr Asn Ile Ala Arg Lys Gly Leu
1160 1165 1170
Trp Val Leu Glu Gln Ile Arg Gln Lys Ser Glu Gly Glu Lys Ile
1175 1180 1185
Asn Leu Ala Met Thr Asn Ala Glu Trp Leu Glu Tyr Ala Gln Thr
1190 1195 1200
His Leu Leu
1205
<210> 8
<211> 1233
<212> PRT
<213> Unknown
<220>
<223> 毛螺菌科细菌
<400> 8
Met Asp Tyr Gly Asn Gly Gln Phe Glu Arg Arg Ala Pro Leu Thr Lys
1 5 10 15
Thr Ile Thr Leu Arg Leu Lys Pro Ile Gly Glu Thr Arg Glu Thr Ile
20 25 30
Arg Glu Gln Lys Leu Leu Glu Gln Asp Ala Ala Phe Arg Lys Leu Val
35 40 45
Glu Thr Val Thr Pro Ile Val Asp Asp Cys Ile Arg Lys Ile Ala Asp
50 55 60
Asn Ala Leu Cys His Phe Gly Thr Glu Tyr Asp Phe Ser Cys Leu Gly
65 70 75 80
Asn Ala Ile Ser Lys Asn Asp Ser Lys Ala Ile Lys Lys Glu Thr Glu
85 90 95
Lys Val Glu Lys Leu Leu Ala Lys Val Leu Thr Glu Asn Leu Pro Asp
100 105 110
Gly Leu Arg Lys Val Asn Asp Ile Asn Ser Ala Ala Phe Ile Gln Asp
115 120 125
Thr Leu Thr Ser Phe Val Gln Asp Asp Ala Asp Lys Arg Val Leu Ile
130 135 140
Gln Glu Leu Lys Gly Lys Thr Val Leu Met Gln Arg Phe Leu Thr Thr
145 150 155 160
Arg Ile Thr Ala Leu Thr Val Trp Leu Pro Asp Arg Val Phe Glu Asn
165 170 175
Phe Asn Ile Phe Ile Glu Asn Ala Glu Lys Met Arg Ile Leu Leu Asp
180 185 190
Ser Pro Leu Asn Glu Lys Ile Met Lys Phe Asp Pro Asp Ala Glu Gln
195 200 205
Tyr Ala Ser Leu Glu Phe Tyr Gly Gln Cys Leu Ser Gln Lys Asp Ile
210 215 220
Asp Ser Tyr Asn Leu Ile Ile Ser Gly Ile Tyr Ala Asp Asp Glu Val
225 230 235 240
Lys Asn Pro Gly Ile Asn Glu Ile Val Lys Glu Tyr Asn Gln Gln Ile
245 250 255
Arg Gly Asp Lys Asp Glu Ser Pro Leu Pro Lys Leu Lys Lys Leu His
260 265 270
Lys Gln Ile Leu Met Pro Val Glu Lys Ala Phe Phe Val Arg Val Leu
275 280 285
Ser Asn Asp Ser Asp Ala Arg Ser Ile Leu Glu Lys Ile Leu Lys Asp
290 295 300
Thr Glu Met Leu Pro Ser Lys Ile Ile Glu Ala Met Lys Glu Ala Asp
305 310 315 320
Ala Gly Asp Ile Ala Val Tyr Gly Ser Arg Leu His Glu Leu Ser His
325 330 335
Val Ile Tyr Gly Asp His Gly Lys Leu Ser Gln Ile Ile Tyr Asp Lys
340 345 350
Glu Ser Lys Arg Ile Ser Glu Leu Met Glu Thr Leu Ser Pro Lys Glu
355 360 365
Arg Lys Glu Ser Lys Lys Arg Leu Glu Gly Leu Glu Glu His Ile Arg
370 375 380
Lys Ser Thr Tyr Thr Phe Asp Glu Leu Asn Arg Tyr Ala Glu Lys Asn
385 390 395 400
Val Met Ala Ala Tyr Ile Ala Ala Val Glu Glu Ser Cys Ala Glu Ile
405 410 415
Met Arg Lys Glu Lys Asp Leu Arg Thr Leu Leu Ser Lys Glu Asp Val
420 425 430
Lys Ile Arg Gly Asn Arg His Asn Thr Leu Ile Val Lys Asn Tyr Phe
435 440 445
Asn Ala Trp Thr Val Phe Arg Asn Leu Ile Arg Ile Leu Arg Arg Lys
450 455 460
Ser Glu Ala Glu Ile Asp Ser Asp Phe Tyr Asp Val Leu Asp Asp Ser
465 470 475 480
Val Glu Val Leu Ser Leu Thr Tyr Lys Gly Glu Asn Leu Cys Arg Ser
485 490 495
Tyr Ile Thr Lys Lys Ile Gly Ser Asp Leu Lys Pro Glu Ile Ala Thr
500 505 510
Tyr Gly Ser Ala Leu Arg Pro Asn Ser Arg Trp Trp Ser Pro Gly Glu
515 520 525
Lys Phe Asn Val Lys Phe His Thr Ile Val Arg Arg Asp Gly Arg Leu
530 535 540
Tyr Tyr Phe Ile Leu Pro Lys Gly Ala Lys Pro Val Glu Leu Glu Asp
545 550 555 560
Met Asp Gly Asp Ile Glu Cys Leu Gln Met Arg Lys Ile Pro Asn Pro
565 570 575
Thr Ile Phe Leu Pro Lys Leu Val Phe Lys Asp Pro Glu Ala Phe Phe
580 585 590
Arg Asp Asn Pro Glu Ala Asp Glu Phe Val Phe Leu Ser Gly Met Lys
595 600 605
Ala Pro Val Thr Ile Thr Arg Glu Thr Tyr Glu Ala Tyr Arg Tyr Lys
610 615 620
Leu Tyr Thr Val Gly Lys Leu Arg Asp Gly Glu Val Ser Glu Glu Glu
625 630 635 640
Tyr Lys Arg Ala Leu Leu Gln Val Leu Thr Ala Tyr Lys Glu Phe Leu
645 650 655
Glu Asn Arg Met Ile Tyr Ala Asp Leu Asn Phe Gly Phe Lys Asp Leu
660 665 670
Glu Glu Tyr Lys Asp Ser Ser Glu Phe Ile Lys Gln Val Glu Thr His
675 680 685
Asn Thr Phe Met Cys Trp Ala Lys Val Ser Ser Ser Gln Leu Asp Asp
690 695 700
Leu Val Lys Ser Gly Asn Gly Leu Leu Phe Glu Ile Trp Ser Glu Arg
705 710 715 720
Leu Glu Ser Tyr Tyr Lys Tyr Gly Asn Glu Lys Val Leu Arg Gly Tyr
725 730 735
Glu Gly Val Leu Leu Ser Ile Leu Lys Asp Glu Asn Leu Val Ser Met
740 745 750
Arg Thr Leu Leu Asn Ser Arg Pro Met Leu Val Tyr Arg Pro Lys Glu
755 760 765
Ser Ser Lys Pro Met Val Val His Arg Asp Gly Ser Arg Val Val Asp
770 775 780
Arg Phe Asp Lys Asp Gly Lys Tyr Ile Pro Pro Glu Val His Asp Glu
785 790 795 800
Leu Tyr Arg Phe Phe Asn Asn Leu Leu Ile Lys Glu Lys Leu Gly Glu
805 810 815
Lys Ala Arg Lys Ile Leu Asp Asn Lys Lys Val Lys Val Lys Val Leu
820 825 830
Glu Ser Glu Arg Val Lys Trp Ser Lys Phe Tyr Asp Glu Gln Phe Ala
835 840 845
Val Thr Phe Ser Val Lys Lys Asn Ala Asp Cys Leu Asp Thr Thr Lys
850 855 860
Asp Leu Asn Ala Glu Val Met Glu Gln Tyr Ser Glu Ser Asn Arg Leu
865 870 875 880
Ile Leu Ile Arg Asn Thr Thr Asp Ile Leu Tyr Tyr Leu Val Leu Asp
885 890 895
Lys Asn Gly Lys Val Leu Lys Gln Arg Ser Leu Asn Ile Ile Asn Asp
900 905 910
Gly Ala Arg Asp Val Asp Trp Lys Glu Arg Phe Arg Gln Val Thr Lys
915 920 925
Asp Arg Asn Glu Gly Tyr Asn Glu Trp Asp Tyr Ser Arg Thr Ser Asn
930 935 940
Asp Leu Lys Glu Val Tyr Leu Asn Tyr Ala Leu Lys Glu Ile Ala Glu
945 950 955 960
Ala Val Ile Glu Tyr Asn Ala Ile Leu Ile Ile Glu Lys Met Ser Asn
965 970 975
Ala Phe Lys Asp Lys Tyr Ser Phe Leu Asp Asp Val Thr Phe Lys Gly
980 985 990
Phe Glu Thr Lys Lys Leu Ala Lys Leu Ser Asp Leu His Phe Arg Gly
995 1000 1005
Ile Lys Asp Gly Glu Pro Cys Ser Phe Thr Asn Pro Leu Gln Leu
1010 1015 1020
Cys Gln Asn Asp Ser Asn Lys Ile Leu Gln Asp Gly Val Ile Phe
1025 1030 1035
Met Val Pro Asn Ser Met Thr Arg Ser Leu Asp Pro Asp Thr Gly
1040 1045 1050
Phe Ile Phe Ala Ile Asn Asp His Asn Ile Arg Thr Lys Lys Ala
1055 1060 1065
Lys Leu Asn Phe Leu Ser Lys Phe Asp Gln Leu Lys Val Ser Ser
1070 1075 1080
Glu Gly Cys Leu Ile Met Lys Tyr Ser Gly Asp Ser Leu Pro Thr
1085 1090 1095
His Asn Thr Asp Asn Arg Val Trp Asn Cys Cys Cys Asn His Pro
1100 1105 1110
Ile Thr Asn Tyr Asp Arg Glu Thr Lys Lys Val Glu Phe Ile Glu
1115 1120 1125
Glu Pro Val Glu Glu Leu Ser Arg Val Leu Glu Glu Asn Gly Ile
1130 1135 1140
Glu Thr Asp Thr Glu Leu Asn Lys Leu Asn Glu Arg Glu Asn Val
1145 1150 1155
Pro Gly Lys Val Val Asp Ala Ile Tyr Ser Leu Val Leu Asn Tyr
1160 1165 1170
Leu Arg Gly Thr Val Ser Gly Val Ala Gly Gln Arg Ala Val Tyr
1175 1180 1185
Tyr Ser Pro Val Thr Gly Lys Lys Tyr Asp Ile Ser Phe Ile Gln
1190 1195 1200
Ala Met Asn Leu Asn Arg Lys Cys Asp Tyr Tyr Arg Ile Gly Ser
1205 1210 1215
Lys Glu Arg Gly Glu Trp Thr Asp Phe Val Ala Gln Leu Ile Asn
1220 1225 1230
<210> 9
<211> 1227
<212> PRT
<213> Unknown
<220>
<223> 毛螺菌科细菌
<400> 9
Met Ser Lys Leu Glu Lys Phe Thr Asn Cys Tyr Ser Leu Ser Lys Thr
1 5 10 15
Leu Arg Phe Lys Ala Ile Pro Val Gly Lys Thr Gln Glu Asn Ile Asp
20 25 30
Asn Lys Arg Leu Leu Val Glu Asp Glu Lys Arg Ala Glu Asp Tyr Lys
35 40 45
Gly Val Lys Lys Leu Leu Asp Arg Tyr Tyr Leu Ser Phe Ile Asn Asp
50 55 60
Val Leu His Ser Ile Lys Leu Lys Asn Leu Asn Asn Tyr Ile Ser Leu
65 70 75 80
Phe Arg Lys Lys Thr Arg Thr Glu Lys Glu Asn Lys Glu Leu Glu Asn
85 90 95
Leu Glu Ile Asn Leu Arg Lys Glu Ile Ala Lys Ala Phe Lys Gly Asn
100 105 110
Glu Gly Tyr Lys Ser Leu Phe Lys Lys Asp Ile Ile Glu Thr Ile Leu
115 120 125
Pro Glu Phe Leu Asp Asp Lys Asp Glu Ile Ala Leu Val Asn Ser Phe
130 135 140
Asn Gly Phe Thr Thr Ala Phe Thr Gly Phe Phe Asp Asn Arg Glu Asn
145 150 155 160
Met Phe Ser Glu Glu Ala Lys Ser Thr Ser Ile Ala Phe Arg Cys Ile
165 170 175
Asn Glu Asn Leu Thr Arg Tyr Ile Ser Asn Met Asp Ile Phe Glu Lys
180 185 190
Val Asp Ala Ile Phe Asp Lys His Glu Val Gln Glu Ile Lys Glu Lys
195 200 205
Ile Leu Asn Ser Asp Tyr Asp Val Glu Asp Phe Phe Glu Gly Glu Phe
210 215 220
Phe Asn Phe Val Leu Thr Gln Glu Gly Ile Asp Val Tyr Asn Ala Ile
225 230 235 240
Ile Gly Gly Phe Val Thr Glu Ser Gly Glu Lys Ile Lys Gly Leu Asn
245 250 255
Glu Tyr Ile Asn Leu Tyr Asn Gln Lys Thr Lys Gln Lys Leu Pro Lys
260 265 270
Phe Lys Pro Leu Tyr Lys Gln Val Leu Ser Asp Arg Glu Ser Leu Ser
275 280 285
Phe Tyr Gly Glu Gly Tyr Thr Ser Asp Glu Glu Val Leu Glu Val Phe
290 295 300
Arg Asn Thr Leu Asn Lys Asn Ser Glu Ile Phe Ser Ser Ile Lys Lys
305 310 315 320
Leu Glu Lys Leu Phe Lys Asn Phe Asp Glu Tyr Ser Ser Ala Gly Ile
325 330 335
Phe Val Lys Asn Gly Pro Ala Ile Ser Thr Ile Ser Lys Asp Ile Phe
340 345 350
Gly Glu Trp Asn Val Ile Arg Asp Lys Trp Asn Ala Glu Tyr Asp Asp
355 360 365
Ile His Leu Lys Lys Lys Ala Val Val Thr Glu Lys Tyr Glu Asp Asp
370 375 380
Arg Arg Lys Ser Phe Lys Lys Ile Gly Ser Phe Ser Leu Glu Gln Leu
385 390 395 400
Gln Glu Tyr Ala Asp Ala Asp Leu Ser Val Val Glu Lys Leu Lys Glu
405 410 415
Ile Ile Ile Gln Lys Val Asp Glu Ile Tyr Lys Val Tyr Gly Ser Ser
420 425 430
Glu Lys Leu Phe Asp Ala Asp Phe Val Leu Glu Lys Ser Leu Lys Lys
435 440 445
Asn Asp Ala Val Val Ala Ile Met Lys Asp Leu Leu Asp Ser Val Lys
450 455 460
Ser Phe Glu Asn Tyr Ile Lys Ala Phe Phe Gly Glu Gly Lys Glu Thr
465 470 475 480
Asn Arg Asp Glu Ser Phe Tyr Gly Asp Phe Val Leu Ala Tyr Asp Ile
485 490 495
Leu Leu Lys Val Asp His Ile Tyr Asp Ala Ile Arg Asn Tyr Val Thr
500 505 510
Gln Lys Pro Tyr Ser Lys Asp Lys Phe Lys Leu Tyr Phe Gln Asn Pro
515 520 525
Gln Phe Met Gly Gly Trp Asp Lys Asp Lys Glu Thr Asp Tyr Arg Ala
530 535 540
Thr Ile Leu Arg Tyr Gly Ser Lys Tyr Tyr Leu Ala Ile Met Asp Lys
545 550 555 560
Lys Tyr Ala Lys Cys Leu Gln Lys Ile Asp Lys Asp Asp Val Asn Gly
565 570 575
Asn Tyr Glu Lys Ile Asn Tyr Lys Leu Leu Pro Gly Pro Asn Lys Met
580 585 590
Leu Pro Lys Val Phe Phe Ser Lys Lys Trp Met Ala Tyr Tyr Asn Pro
595 600 605
Ser Glu Asp Ile Gln Lys Ile Tyr Lys Asn Gly Thr Phe Lys Lys Gly
610 615 620
Asp Met Phe Asn Leu Asn Asp Cys His Lys Leu Ile Asp Phe Phe Lys
625 630 635 640
Asp Ser Ile Ser Arg Tyr Pro Lys Trp Ser Asn Ala Tyr Asp Phe Asn
645 650 655
Phe Ser Glu Thr Glu Lys Tyr Lys Asp Ile Ala Gly Phe Tyr Arg Glu
660 665 670
Val Glu Glu Gln Gly Tyr Lys Val Ser Phe Glu Ser Ala Ser Lys Lys
675 680 685
Glu Val Asp Lys Leu Val Glu Glu Gly Lys Leu Tyr Met Phe Gln Ile
690 695 700
Tyr Asn Lys Asp Phe Ser Asp Lys Ser His Gly Thr Pro Asn Leu His
705 710 715 720
Thr Met Tyr Phe Lys Leu Leu Phe Asp Glu Asn Asn His Gly Gln Ile
725 730 735
Arg Leu Ser Gly Gly Ala Glu Leu Phe Met Arg Arg Ala Ser Leu Lys
740 745 750
Lys Glu Glu Leu Val Val His Pro Ala Asn Ser Pro Ile Ala Asn Lys
755 760 765
Asn Pro Asp Asn Pro Lys Lys Thr Thr Thr Leu Ser Tyr Asp Val Tyr
770 775 780
Lys Asp Lys Arg Phe Ser Glu Asp Gln Tyr Glu Leu His Ile Pro Ile
785 790 795 800
Ala Asn Ile Asn Lys Cys Pro Lys Asn Ile Phe Lys Ile Asn Thr Glu
805 810 815
Val Arg Val Leu Leu Lys His Asp Asp Asn Pro Tyr Val Ile Gly Ile
820 825 830
Asp Arg Gly Glu Arg Asn Leu Leu Tyr Ile Val Val Val Asp Gly Lys
835 840 845
Gly Asn Ile Val Glu Gln Tyr Ser Leu Asn Glu Ile Ile Asn Asn Phe
850 855 860
Asn Gly Ile Arg Ile Lys Thr Asp Tyr His Ser Leu Leu Asp Lys Lys
865 870 875 880
Glu Lys Glu Arg Phe Glu Ala Arg Gln Asn Trp Thr Ser Ile Glu Asn
885 890 895
Ile Lys Glu Leu Lys Ala Gly Tyr Ile Ser Gln Val Val His Lys Ile
900 905 910
Cys Glu Leu Val Glu Lys Tyr Asp Ala Val Ile Ala Leu Glu Asp Leu
915 920 925
Asn Ser Gly Phe Lys Asn Ser Arg Val Lys Val Glu Lys Gln Val Tyr
930 935 940
Gln Lys Phe Glu Lys Met Leu Ile Asp Lys Leu Asn Tyr Met Val Asp
945 950 955 960
Lys Lys Ser Asn Pro Cys Ala Thr Gly Gly Ala Leu Lys Gly Tyr Gln
965 970 975
Ile Thr Asn Lys Phe Glu Ser Phe Lys Ser Met Ser Thr Gln Asn Gly
980 985 990
Phe Ile Phe Tyr Ile Pro Ala Trp Leu Thr Ser Lys Ile Asp Pro Ser
995 1000 1005
Thr Gly Phe Val Asn Leu Leu Lys Thr Lys Tyr Thr Ser Ile Ala
1010 1015 1020
Asp Lys Lys Phe Ile Ser Ser Phe Asp Arg Ile Met Tyr Val Pro
1025 1030 1035
Glu Glu Asp Leu Phe Glu Phe Ala Leu Asp Tyr Lys Asn Phe Ser
1040 1045 1050
Arg Thr Asp Ala Asp Tyr Ile Lys Lys Trp Lys Leu Tyr Ser Tyr
1055 1060 1065
Gly Asn Arg Ile Arg Ile Phe Arg Asn Pro Lys Lys Asn Asn Val
1070 1075 1080
Phe Asp Trp Glu Glu Val Cys Leu Thr Ser Ala Tyr Lys Glu Leu
1085 1090 1095
Phe Asn Lys Tyr Gly Ile Asn Tyr Gln Gln Gly Asp Ile Arg Ala
1100 1105 1110
Leu Leu Cys Glu Gln Ser Asp Lys Ala Phe Tyr Ser Ser Phe Met
1115 1120 1125
Ala Leu Met Ser Leu Met Leu Gln Met Arg Asn Ser Ile Thr Gly
1130 1135 1140
Arg Thr Asp Val Asp Phe Leu Ile Ser Pro Val Lys Asn Ser Asp
1145 1150 1155
Gly Ile Phe Tyr Asp Ser Arg Asn Tyr Glu Ala Gln Glu Asn Ala
1160 1165 1170
Ile Leu Pro Lys Asn Ala Asp Ala Asn Gly Ala Tyr Asn Ile Ala
1175 1180 1185
Arg Lys Val Leu Trp Ala Ile Gly Gln Phe Lys Lys Ala Glu Asp
1190 1195 1200
Glu Lys Leu Asp Lys Val Lys Ile Ala Ser Asn Lys Glu Trp Leu
1205 1210 1215
Glu Tyr Ala Gln Thr Ser Val Lys His
1220 1225
<210> 10
<211> 1264
<212> PRT
<213> 稻田氏钩端螺旋体（Leptospira inadai）
<400> 10
Met Glu Asp Tyr Ser Gly Phe Val Asn Ile Tyr Ser Ile Gln Lys Thr
1 5 10 15
Leu Arg Phe Glu Leu Lys Pro Val Gly Lys Thr Leu Glu His Ile Glu
20 25 30
Lys Lys Gly Phe Leu Lys Lys Asp Lys Ile Arg Ala Glu Asp Tyr Lys
35 40 45
Ala Val Lys Lys Ile Ile Asp Lys Tyr His Arg Ala Tyr Ile Glu Glu
50 55 60
Val Phe Asp Ser Val Leu His Gln Lys Lys Lys Lys Asp Lys Thr Arg
65 70 75 80
Phe Ser Thr Gln Phe Ile Lys Glu Ile Lys Glu Phe Ser Glu Leu Tyr
85 90 95
Tyr Lys Thr Glu Lys Asn Ile Pro Asp Lys Glu Arg Leu Glu Ala Leu
100 105 110
Ser Glu Lys Leu Arg Lys Met Leu Val Gly Ala Phe Lys Gly Glu Phe
115 120 125
Ser Glu Glu Val Ala Glu Lys Tyr Asn Lys Asn Leu Phe Ser Lys Glu
130 135 140
Leu Ile Arg Asn Glu Ile Glu Lys Phe Cys Glu Thr Asp Glu Glu Arg
145 150 155 160
Lys Gln Val Ser Asn Phe Lys Ser Phe Thr Thr Tyr Phe Thr Gly Phe
165 170 175
His Ser Asn Arg Gln Asn Ile Tyr Ser Asp Glu Lys Lys Ser Thr Ala
180 185 190
Ile Gly Tyr Arg Ile Ile His Gln Asn Leu Pro Lys Phe Leu Asp Asn
195 200 205
Leu Lys Ile Ile Glu Ser Ile Gln Arg Arg Phe Lys Asp Phe Pro Trp
210 215 220
Ser Asp Leu Lys Lys Asn Leu Lys Lys Ile Asp Lys Asn Ile Lys Leu
225 230 235 240
Thr Glu Tyr Phe Ser Ile Asp Gly Phe Val Asn Val Leu Asn Gln Lys
245 250 255
Gly Ile Asp Ala Tyr Asn Thr Ile Leu Gly Gly Lys Ser Glu Glu Ser
260 265 270
Gly Glu Lys Ile Gln Gly Leu Asn Glu Tyr Ile Asn Leu Tyr Arg Gln
275 280 285
Lys Asn Asn Ile Asp Arg Lys Asn Pro Leu Asn Val Lys Ile Leu Phe
290 295 300
Lys Gln Ile Leu Gly Asp Arg Glu Thr Lys Ser Phe Ile Pro Glu Ala
305 310 315 320
Phe Pro Asp Asp Gln Ser Val Leu Asn Ser Ile Thr Glu Phe Ala Lys
325 330 335
Tyr Leu Lys Leu Asp Lys Lys Lys Lys Ser Ile Ile Ala Glu Leu Lys
340 345 350
Lys Phe Leu Ser Ser Phe Asn Arg Tyr Glu Leu Asp Gly Ile Tyr Leu
355 360 365
Ala Asn Asp Asn Ser Leu Ala Ser Ile Ser Thr Phe Leu Phe Asp Asp
370 375 380
Trp Ser Phe Ile Lys Lys Ser Val Ser Phe Lys Tyr Asp Glu Ser Val
385 390 395 400
Gly Asp Pro Lys Lys Lys Ile Lys Ser Pro Leu Lys Tyr Glu Lys Glu
405 410 415
Lys Glu Lys Trp Leu Lys Gln Lys Tyr Tyr Thr Ile Ser Phe Leu Asn
420 425 430
Asp Ala Ile Glu Ser Tyr Ser Lys Ser Gln Asp Glu Lys Arg Val Lys
435 440 445
Ile Arg Leu Glu Ala Tyr Phe Ala Glu Phe Lys Ser Lys Asp Asp Ala
450 455 460
Lys Lys Gln Phe Asp Leu Leu Glu Arg Ile Glu Glu Ala Tyr Ala Ile
465 470 475 480
Val Glu Pro Leu Leu Gly Ala Glu Tyr Pro Arg Asp Arg Asn Leu Lys
485 490 495
Ala Asp Lys Lys Glu Val Gly Lys Ile Lys Asp Phe Leu Asp Ser Ile
500 505 510
Lys Ser Leu Gln Phe Phe Leu Lys Pro Leu Leu Ser Ala Glu Ile Phe
515 520 525
Asp Glu Lys Asp Leu Gly Phe Tyr Asn Gln Leu Glu Gly Tyr Tyr Glu
530 535 540
Glu Ile Asp Ile Ser Gly His Leu Tyr Asn Lys Val Arg Asn Tyr Leu
545 550 555 560
Thr Gly Lys Ile Tyr Ser Lys Glu Lys Phe Lys Leu Asn Phe Glu Asn
565 570 575
Ser Thr Leu Leu Lys Gly Trp Asp Glu Asn Arg Glu Val Ala Asn Leu
580 585 590
Cys Val Ile Phe Arg Glu Asp Gln Lys Tyr Tyr Leu Gly Val Met Asp
595 600 605
Lys Glu Asn Asn Thr Ile Leu Ser Asp Ile Pro Lys Val Lys Pro Asn
610 615 620
Glu Leu Phe Tyr Glu Lys Met Val Tyr Lys Leu Ile Pro Thr Pro His
625 630 635 640
Met Gln Leu Pro Arg Ile Ile Phe Ser Ser Asp Asn Leu Ser Ile Tyr
645 650 655
Asn Pro Ser Lys Ser Ile Leu Lys Ile Arg Glu Ala Lys Ser Phe Lys
660 665 670
Glu Gly Lys Asn Phe Lys Leu Lys Asp Cys His Lys Phe Ile Asp Phe
675 680 685
Tyr Lys Glu Ser Ile Ser Lys Asn Glu Asp Trp Ser Arg Phe Asp Phe
690 695 700
Lys Phe Ser Lys Thr Ser Ser Tyr Glu Asn Ile Ser Glu Phe Tyr Arg
705 710 715 720
Glu Val Glu Arg Gln Gly Tyr Asn Leu Asp Phe Lys Lys Val Ser Lys
725 730 735
Phe Tyr Ile Asp Ser Leu Val Glu Asp Gly Lys Leu Tyr Leu Phe Gln
740 745 750
Ile Tyr Asn Lys Asp Phe Ser Ile Phe Ser Lys Gly Lys Pro Asn Leu
755 760 765
His Thr Ile Tyr Phe Arg Ser Leu Phe Ser Lys Glu Asn Leu Lys Asp
770 775 780
Val Cys Leu Lys Leu Asn Gly Glu Ala Glu Met Phe Phe Arg Lys Lys
785 790 795 800
Ser Ile Asn Tyr Asp Glu Lys Lys Lys Arg Glu Gly His His Pro Glu
805 810 815
Leu Phe Glu Lys Leu Lys Tyr Pro Ile Leu Lys Asp Lys Arg Tyr Ser
820 825 830
Glu Asp Lys Phe Gln Phe His Leu Pro Ile Ser Leu Asn Phe Lys Ser
835 840 845
Lys Glu Arg Leu Asn Phe Asn Leu Lys Val Asn Glu Phe Leu Lys Arg
850 855 860
Asn Lys Asp Ile Asn Ile Ile Gly Ile Asp Arg Gly Glu Arg Asn Leu
865 870 875 880
Leu Tyr Leu Val Met Ile Asn Gln Lys Gly Glu Ile Leu Lys Gln Thr
885 890 895
Leu Leu Asp Ser Met Gln Ser Gly Lys Gly Arg Pro Glu Ile Asn Tyr
900 905 910
Lys Glu Lys Leu Gln Glu Lys Glu Ile Glu Arg Asp Lys Ala Arg Lys
915 920 925
Ser Trp Gly Thr Val Glu Asn Ile Lys Glu Leu Lys Glu Gly Tyr Leu
930 935 940
Ser Ile Val Ile His Gln Ile Ser Lys Leu Met Val Glu Asn Asn Ala
945 950 955 960
Ile Val Val Leu Glu Asp Leu Asn Ile Gly Phe Lys Arg Gly Arg Gln
965 970 975
Lys Val Glu Arg Gln Val Tyr Gln Lys Phe Glu Lys Met Leu Ile Asp
980 985 990
Lys Leu Asn Phe Leu Val Phe Lys Glu Asn Lys Pro Thr Glu Pro Gly
995 1000 1005
Gly Val Leu Lys Ala Tyr Gln Leu Thr Asp Glu Phe Gln Ser Phe
1010 1015 1020
Glu Lys Leu Ser Lys Gln Thr Gly Phe Leu Phe Tyr Val Pro Ser
1025 1030 1035
Trp Asn Thr Ser Lys Ile Asp Pro Arg Thr Gly Phe Ile Asp Phe
1040 1045 1050
Leu His Pro Ala Tyr Glu Asn Ile Glu Lys Ala Lys Gln Trp Ile
1055 1060 1065
Asn Lys Phe Asp Ser Ile Arg Phe Asn Ser Lys Met Asp Trp Phe
1070 1075 1080
Glu Phe Thr Ala Asp Thr Arg Lys Phe Ser Glu Asn Leu Met Leu
1085 1090 1095
Gly Lys Asn Arg Val Trp Val Ile Cys Thr Thr Asn Val Glu Arg
1100 1105 1110
Tyr Phe Thr Ser Lys Thr Ala Asn Ser Ser Ile Gln Tyr Asn Ser
1115 1120 1125
Ile Gln Ile Thr Glu Lys Leu Lys Glu Leu Phe Val Asp Ile Pro
1130 1135 1140
Phe Ser Asn Gly Gln Asp Leu Lys Pro Glu Ile Leu Arg Lys Asn
1145 1150 1155
Asp Ala Val Phe Phe Lys Ser Leu Leu Phe Tyr Ile Lys Thr Thr
1160 1165 1170
Leu Ser Leu Arg Gln Asn Asn Gly Lys Lys Gly Glu Glu Glu Lys
1175 1180 1185
Asp Phe Ile Leu Ser Pro Val Val Asp Ser Lys Gly Arg Phe Phe
1190 1195 1200
Asn Ser Leu Glu Ala Ser Asp Asp Glu Pro Lys Asp Ala Asp Ala
1205 1210 1215
Asn Gly Ala Tyr His Ile Ala Leu Lys Gly Leu Met Asn Leu Leu
1220 1225 1230
Val Leu Asn Glu Thr Lys Glu Glu Asn Leu Ser Arg Pro Lys Trp
1235 1240 1245
Lys Ile Lys Asn Lys Asp Trp Leu Glu Phe Val Trp Glu Arg Asn
1250 1255 1260
Arg
<210> 11
<211> 1373
<212> PRT
<213> 牛眼莫拉氏菌（Moraxella bovoculi）
<400> 11
Met Leu Phe Gln Asp Phe Thr His Leu Tyr Pro Leu Ser Lys Thr Val
1 5 10 15
Arg Phe Glu Leu Phe Ile Asp Arg Thr Leu Glu His Ile His Ala Lys
20 25 30
Asn Phe Leu Ser Gln Asp Glu Thr Met Ala Asp Met His Gln Lys Val
35 40 45
Lys Val Ile Leu Asp Asp Tyr His Arg Asp Phe Ile Ala Asp Met Met
50 55 60
Gly Glu Val Lys Leu Thr Lys Leu Ala Glu Phe Tyr Asp Val Tyr Leu
65 70 75 80
Lys Phe Arg Lys Asn Pro Lys Asp Asp Glu Leu Gln Lys Ala Gln Leu
85 90 95
Lys Asp Leu Gln Ala Val Leu Arg Lys Glu Ile Val Lys Pro Ile Gly
100 105 110
Asn Gly Gly Lys Tyr Lys Ala Gly Tyr Asp Arg Leu Phe Gly Ala Lys
115 120 125
Leu Phe Lys Asp Gly Lys Glu Leu Gly Asp Leu Ala Lys Phe Val Ile
130 135 140
Ala Gln Glu Gly Glu Ser Ser Pro Lys Leu Ala His Leu Ala His Phe
145 150 155 160
Glu Lys Phe Ser Thr Tyr Phe Thr Gly Phe His Asp Asn Arg Lys Asn
165 170 175
Met Tyr Ser Asp Glu Asp Lys His Thr Ala Ile Ala Tyr Arg Leu Ile
180 185 190
His Glu Asn Leu Pro Arg Phe Ile Asp Asn Leu Gln Ile Leu Thr Thr
195 200 205
Ile Lys Gln Lys His Ser Ala Leu Tyr Asp Gln Ile Ile Asn Glu Leu
210 215 220
Thr Ala Ser Gly Leu Asp Val Ser Leu Ala Ser His Leu Asp Gly Tyr
225 230 235 240
His Lys Leu Leu Thr Gln Glu Gly Ile Thr Ala Tyr Asn Thr Leu Leu
245 250 255
Gly Gly Ile Ser Gly Glu Ala Gly Ser Pro Lys Ile Gln Gly Ile Asn
260 265 270
Glu Leu Ile Asn Ser His His Asn Gln His Cys His Lys Ser Glu Arg
275 280 285
Ile Ala Lys Leu Arg Pro Leu His Lys Gln Ile Leu Ser Asp Gly Met
290 295 300
Ser Val Ser Phe Leu Pro Ser Lys Phe Ala Asp Asp Ser Glu Met Cys
305 310 315 320
Gln Ala Val Asn Glu Phe Tyr Arg His Tyr Ala Asp Val Phe Ala Lys
325 330 335
Val Gln Ser Leu Phe Asp Gly Phe Asp Asp His Gln Lys Asp Gly Ile
340 345 350
Tyr Val Glu His Lys Asn Leu Asn Glu Leu Ser Lys Gln Ala Phe Gly
355 360 365
Asp Phe Ala Leu Leu Gly Arg Val Leu Asp Gly Tyr Tyr Val Asp Val
370 375 380
Val Asn Pro Glu Phe Asn Glu Arg Phe Ala Lys Ala Lys Thr Asp Asn
385 390 395 400
Ala Lys Ala Lys Leu Thr Lys Glu Lys Asp Lys Phe Ile Lys Gly Val
405 410 415
His Ser Leu Ala Ser Leu Glu Gln Ala Ile Glu His Tyr Thr Ala Arg
420 425 430
His Asp Asp Glu Ser Val Gln Ala Gly Lys Leu Gly Gln Tyr Phe Lys
435 440 445
His Gly Leu Ala Gly Val Asp Asn Pro Ile Gln Lys Ile His Asn Asn
450 455 460
His Ser Thr Ile Lys Gly Phe Leu Glu Arg Glu Arg Pro Ala Gly Glu
465 470 475 480
Arg Ala Leu Pro Lys Ile Lys Ser Gly Lys Asn Pro Glu Met Thr Gln
485 490 495
Leu Arg Gln Leu Lys Glu Leu Leu Asp Asn Ala Leu Asn Val Ala His
500 505 510
Phe Ala Lys Leu Leu Thr Thr Lys Thr Thr Leu Asp Asn Gln Asp Gly
515 520 525
Asn Phe Tyr Gly Glu Phe Gly Val Leu Tyr Asp Glu Leu Ala Lys Ile
530 535 540
Pro Thr Leu Tyr Asn Lys Val Arg Asp Tyr Leu Ser Gln Lys Pro Phe
545 550 555 560
Ser Thr Glu Lys Tyr Lys Leu Asn Phe Gly Asn Pro Thr Leu Leu Asn
565 570 575
Gly Trp Asp Leu Asn Lys Glu Lys Asp Asn Phe Gly Val Ile Leu Gln
580 585 590
Lys Asp Gly Cys Tyr Tyr Leu Ala Leu Leu Asp Lys Ala His Lys Lys
595 600 605
Val Phe Asp Asn Ala Pro Asn Thr Gly Lys Ser Ile Tyr Gln Lys Met
610 615 620
Ile Tyr Lys Tyr Leu Glu Val Arg Lys Gln Phe Pro Lys Val Phe Phe
625 630 635 640
Ser Lys Glu Ala Ile Ala Ile Asn Tyr His Pro Ser Lys Glu Leu Val
645 650 655
Glu Ile Lys Asp Lys Gly Arg Gln Arg Ser Asp Asp Glu Arg Leu Lys
660 665 670
Leu Tyr Arg Phe Ile Leu Glu Cys Leu Lys Ile His Pro Lys Tyr Asp
675 680 685
Lys Lys Phe Glu Gly Ala Ile Gly Asp Ile Gln Leu Phe Lys Lys Asp
690 695 700
Lys Lys Gly Arg Glu Val Pro Ile Ser Glu Lys Asp Leu Phe Lys Asp
705 710 715 720
Ile Asn Gly Ile Phe Ser Ser Lys Pro Lys Leu Glu Met Glu Asp Phe
725 730 735
Phe Ile Gly Glu Phe Lys Arg Tyr Asn Pro Ser Gln Asp Leu Val Asp
740 745 750
Gln Tyr Asn Ile Tyr Lys Lys Ile Asp Ser Asn Asp Asn Arg Lys Lys
755 760 765
Glu Asn Phe Tyr Asn Asn His Pro Lys Phe Lys Lys Asp Leu Val Arg
770 775 780
Tyr Tyr Tyr Glu Ser Met Cys Lys His Glu Glu Trp Glu Glu Ser Phe
785 790 795 800
Glu Phe Ser Lys Lys Leu Gln Asp Ile Gly Cys Tyr Val Asp Val Asn
805 810 815
Glu Leu Phe Thr Glu Ile Glu Thr Arg Arg Leu Asn Tyr Lys Ile Ser
820 825 830
Phe Cys Asn Ile Asn Ala Asp Tyr Ile Asp Glu Leu Val Glu Gln Gly
835 840 845
Gln Leu Tyr Leu Phe Gln Ile Tyr Asn Lys Asp Phe Ser Pro Lys Ala
850 855 860
His Gly Lys Pro Asn Leu His Thr Leu Tyr Phe Lys Ala Leu Phe Ser
865 870 875 880
Glu Asp Asn Leu Ala Asp Pro Ile Tyr Lys Leu Asn Gly Glu Ala Gln
885 890 895
Ile Phe Tyr Arg Lys Ala Ser Leu Asp Met Asn Glu Thr Thr Ile His
900 905 910
Arg Ala Gly Glu Val Leu Glu Asn Lys Asn Pro Asp Asn Pro Lys Lys
915 920 925
Arg Gln Phe Val Tyr Asp Ile Ile Lys Asp Lys Arg Tyr Thr Gln Lys
930 935 940
Asp Phe Met Leu His Val Pro Ile Thr Met Asn Phe Gly Val Gln Gly
945 950 955 960
Met Thr Ile Lys Glu Phe Asn Lys Lys Val Asn Gln Ser Ile Gln Gln
965 970 975
Tyr Asp Glu Val Asn Val Ile Gly Ile Asp Arg Gly Glu Arg His Leu
980 985 990
Leu Tyr Leu Thr Val Ile Asn Ser Lys Gly Glu Ile Leu Glu Gln Cys
995 1000 1005
Ser Leu Asn Asp Ile Thr Thr Ala Ser Ala Asn Gly Thr Gln Met
1010 1015 1020
Thr Thr Pro Tyr His Lys Ile Leu Asp Lys Arg Glu Ile Glu Arg
1025 1030 1035
Leu Asn Ala Arg Val Gly Trp Gly Glu Ile Glu Thr Ile Lys Glu
1040 1045 1050
Leu Lys Ser Gly Tyr Leu Ser His Val Val His Gln Ile Ser Gln
1055 1060 1065
Leu Met Leu Lys Tyr Asn Ala Ile Val Val Leu Glu Asp Leu Asn
1070 1075 1080
Phe Gly Phe Lys Arg Gly Arg Phe Lys Val Glu Lys Gln Ile Tyr
1085 1090 1095
Gln Asn Phe Glu Asn Ala Leu Ile Lys Lys Leu Asn His Leu Val
1100 1105 1110
Leu Lys Asp Lys Ala Asp Asp Glu Ile Gly Ser Tyr Lys Asn Ala
1115 1120 1125
Leu Gln Leu Thr Asn Asn Phe Thr Asp Leu Lys Ser Ile Gly Lys
1130 1135 1140
Gln Thr Gly Phe Leu Phe Tyr Val Pro Ala Trp Asn Thr Ser Lys
1145 1150 1155
Ile Asp Pro Glu Thr Gly Phe Val Asp Leu Leu Lys Pro Arg Tyr
1160 1165 1170
Glu Asn Ile Gln Ala Ser Gln Ala Phe Phe Gly Lys Phe Asp Lys
1175 1180 1185
Ile Cys Tyr Asn Ala Asp Lys Asp Tyr Phe Glu Phe His Ile Asp
1190 1195 1200
Tyr Ala Lys Phe Thr Asp Lys Ala Lys Asn Ser Arg Gln Ile Trp
1205 1210 1215
Thr Ile Cys Ser His Gly Asp Lys Arg Tyr Val Tyr Asp Lys Thr
1220 1225 1230
Ala Asn Gln Asn Lys Gly Ala Ala Lys Gly Ile Asn Val Asn Asp
1235 1240 1245
Ile Leu Lys Ser Leu Phe Ala Arg His His Ile Asn Glu Lys Gln
1250 1255 1260
Pro Asn Leu Val Met Asp Ile Cys Gln Asn Asn Asp Lys Glu Phe
1265 1270 1275
His Lys Ser Leu Met Tyr Leu Leu Lys Thr Leu Leu Ala Leu Arg
1280 1285 1290
Tyr Ser Asn Ala Ser Ser Asp Glu Asp Phe Ile Leu Ser Pro Val
1295 1300 1305
Ala Asn Asp Glu Gly Val Phe Phe Asn Ser Ala Leu Ala Asp Asp
1310 1315 1320
Thr Gln Pro Gln Asn Ala Asp Ala Asn Gly Ala Tyr His Ile Ala
1325 1330 1335
Leu Lys Gly Leu Trp Leu Leu Asn Glu Leu Lys Asn Ser Asp Asp
1340 1345 1350
Leu Asn Lys Val Lys Leu Ala Ile Asp Asn Gln Thr Trp Leu Asn
1355 1360 1365
Phe Ala Gln Asn Arg
1370
<210> 12
<211> 1352
<212> PRT
<213> Unknown
<220>
<223> Parcubacteria细菌
<400> 12
Met Glu Asn Ile Phe Asp Gln Phe Ile Gly Lys Tyr Ser Leu Ser Lys
1 5 10 15
Thr Leu Arg Phe Glu Leu Lys Pro Val Gly Lys Thr Glu Asp Phe Leu
20 25 30
Lys Ile Asn Lys Val Phe Glu Lys Asp Gln Thr Ile Asp Asp Ser Tyr
35 40 45
Asn Gln Ala Lys Phe Tyr Phe Asp Ser Leu His Gln Lys Phe Ile Asp
50 55 60
Ala Ala Leu Ala Ser Asp Lys Thr Ser Glu Leu Ser Phe Gln Asn Phe
65 70 75 80
Ala Asp Val Leu Glu Lys Gln Asn Lys Ile Ile Leu Asp Lys Lys Arg
85 90 95
Glu Met Gly Ala Leu Arg Lys Arg Asp Lys Asn Ala Val Gly Ile Asp
100 105 110
Arg Leu Gln Lys Glu Ile Asn Asp Ala Glu Asp Ile Ile Gln Lys Glu
115 120 125
Lys Glu Lys Ile Tyr Lys Asp Val Arg Thr Leu Phe Asp Asn Glu Ala
130 135 140
Glu Ser Trp Lys Thr Tyr Tyr Gln Glu Arg Glu Val Asp Gly Lys Lys
145 150 155 160
Ile Thr Glu Ser Lys Ala Asp Leu Lys Gln Lys Gly Ala Asp Phe Leu
165 170 175
Thr Ala Ala Gly Ile Leu Lys Val Leu Lys Tyr Glu Phe Pro Glu Glu
180 185 190
Lys Glu Lys Glu Phe Gln Ala Lys Asn Gln Pro Ser Leu Phe Val Glu
195 200 205
Glu Lys Glu Asn Pro Gly Gln Lys Arg Tyr Ile Phe Asp Ser Phe Asp
210 215 220
Lys Phe Ala Gly Tyr Leu Thr Lys Phe Gln Gln Thr Lys Lys Asn Leu
225 230 235 240
Tyr Ala Ala Asp Gly Thr Ser Thr Ala Val Ala Thr Arg Ile Ala Asp
245 250 255
Asn Phe Ile Ile Phe His Gln Asn Thr Lys Val Phe Arg Asp Lys Tyr
260 265 270
Lys Asn Asn His Thr Asp Leu Gly Phe Asp Glu Glu Asn Ile Phe Glu
275 280 285
Ile Glu Arg Tyr Lys Asn Cys Leu Leu Gln Arg Glu Ile Glu His Ile
290 295 300
Lys Asn Glu Asn Ser Tyr Asn Lys Ile Ile Gly Arg Ile Asn Lys Lys
305 310 315 320
Ile Lys Glu Tyr Arg Asp Gln Lys Ala Lys Asp Thr Lys Leu Thr Lys
325 330 335
Ser Asp Phe Pro Phe Phe Lys Asn Leu Asp Lys Gln Ile Leu Gly Glu
340 345 350
Val Glu Lys Glu Lys Gln Leu Ile Glu Lys Thr Arg Glu Lys Thr Glu
355 360 365
Glu Asp Val Leu Ile Glu Arg Phe Lys Glu Phe Ile Glu Asn Asn Glu
370 375 380
Glu Arg Phe Thr Ala Ala Lys Lys Leu Met Asn Ala Phe Cys Asn Gly
385 390 395 400
Glu Phe Glu Ser Glu Tyr Glu Gly Ile Tyr Leu Lys Asn Lys Ala Ile
405 410 415
Asn Thr Ile Ser Arg Arg Trp Phe Val Ser Asp Arg Asp Phe Glu Leu
420 425 430
Lys Leu Pro Gln Gln Lys Ser Lys Asn Lys Ser Glu Lys Asn Glu Pro
435 440 445
Lys Val Lys Lys Phe Ile Ser Ile Ala Glu Ile Lys Asn Ala Val Glu
450 455 460
Glu Leu Asp Gly Asp Ile Phe Lys Ala Val Phe Tyr Asp Lys Lys Ile
465 470 475 480
Ile Ala Gln Gly Gly Ser Lys Leu Glu Gln Phe Leu Val Ile Trp Lys
485 490 495
Tyr Glu Phe Glu Tyr Leu Phe Arg Asp Ile Glu Arg Glu Asn Gly Glu
500 505 510
Lys Leu Leu Gly Tyr Asp Ser Cys Leu Lys Ile Ala Lys Gln Leu Gly
515 520 525
Ile Phe Pro Gln Glu Lys Glu Ala Arg Glu Lys Ala Thr Ala Val Ile
530 535 540
Lys Asn Tyr Ala Asp Ala Gly Leu Gly Ile Phe Gln Met Met Lys Tyr
545 550 555 560
Phe Ser Leu Asp Asp Lys Asp Arg Lys Asn Thr Pro Gly Gln Leu Ser
565 570 575
Thr Asn Phe Tyr Ala Glu Tyr Asp Gly Tyr Tyr Lys Asp Phe Glu Phe
580 585 590
Ile Lys Tyr Tyr Asn Glu Phe Arg Asn Phe Ile Thr Lys Lys Pro Phe
595 600 605
Asp Glu Asp Lys Ile Lys Leu Asn Phe Glu Asn Gly Ala Leu Leu Lys
610 615 620
Gly Trp Asp Glu Asn Lys Glu Tyr Asp Phe Met Gly Val Ile Leu Lys
625 630 635 640
Lys Glu Gly Arg Leu Tyr Leu Gly Ile Met His Lys Asn His Arg Lys
645 650 655
Leu Phe Gln Ser Met Gly Asn Ala Lys Gly Asp Asn Ala Asn Arg Tyr
660 665 670
Gln Lys Met Ile Tyr Lys Gln Ile Ala Asp Ala Ser Lys Asp Val Pro
675 680 685
Arg Leu Leu Leu Thr Ser Lys Lys Ala Met Glu Lys Phe Lys Pro Ser
690 695 700
Gln Glu Ile Leu Arg Ile Lys Lys Glu Lys Thr Phe Lys Arg Glu Ser
705 710 715 720
Lys Asn Phe Ser Leu Arg Asp Leu His Ala Leu Ile Glu Tyr Tyr Arg
725 730 735
Asn Cys Ile Pro Gln Tyr Ser Asn Trp Ser Phe Tyr Asp Phe Gln Phe
740 745 750
Gln Asp Thr Gly Lys Tyr Gln Asn Ile Lys Glu Phe Thr Asp Asp Val
755 760 765
Gln Lys Tyr Gly Tyr Lys Ile Ser Phe Arg Asp Ile Asp Asp Glu Tyr
770 775 780
Ile Asn Gln Ala Leu Asn Glu Gly Lys Met Tyr Leu Phe Glu Val Val
785 790 795 800
Asn Lys Asp Ile Tyr Asn Thr Lys Asn Gly Ser Lys Asn Leu His Thr
805 810 815
Leu Tyr Phe Glu His Ile Leu Ser Ala Glu Asn Leu Asn Asp Pro Val
820 825 830
Phe Lys Leu Ser Gly Met Ala Glu Ile Phe Gln Arg Gln Pro Ser Val
835 840 845
Asn Glu Arg Glu Lys Ile Thr Thr Gln Lys Asn Gln Cys Ile Leu Asp
850 855 860
Lys Gly Asp Arg Ala Tyr Lys Tyr Arg Arg Tyr Thr Glu Lys Lys Ile
865 870 875 880
Met Phe His Met Ser Leu Val Leu Asn Thr Gly Lys Gly Glu Ile Lys
885 890 895
Gln Val Gln Phe Asn Lys Ile Ile Asn Gln Arg Ile Ser Ser Ser Asp
900 905 910
Asn Glu Met Arg Val Asn Val Ile Gly Ile Asp Arg Gly Glu Lys Asn
915 920 925
Leu Leu Tyr Tyr Ser Val Val Lys Gln Asn Gly Glu Ile Ile Glu Gln
930 935 940
Ala Ser Leu Asn Glu Ile Asn Gly Val Asn Tyr Arg Asp Lys Leu Ile
945 950 955 960
Glu Arg Glu Lys Glu Arg Leu Lys Asn Arg Gln Ser Trp Lys Pro Val
965 970 975
Val Lys Ile Lys Asp Leu Lys Lys Gly Tyr Ile Ser His Val Ile His
980 985 990
Lys Ile Cys Gln Leu Ile Glu Lys Tyr Ser Ala Ile Val Val Leu Glu
995 1000 1005
Asp Leu Asn Met Arg Phe Lys Gln Ile Arg Gly Gly Ile Glu Arg
1010 1015 1020
Ser Val Tyr Gln Gln Phe Glu Lys Ala Leu Ile Asp Lys Leu Gly
1025 1030 1035
Tyr Leu Val Phe Lys Asp Asn Arg Asp Leu Arg Ala Pro Gly Gly
1040 1045 1050
Val Leu Asn Gly Tyr Gln Leu Ser Ala Pro Phe Val Ser Phe Glu
1055 1060 1065
Lys Met Arg Lys Gln Thr Gly Ile Leu Phe Tyr Thr Gln Ala Glu
1070 1075 1080
Tyr Thr Ser Lys Thr Asp Pro Ile Thr Gly Phe Arg Lys Asn Val
1085 1090 1095
Tyr Ile Ser Asn Ser Ala Ser Leu Asp Lys Ile Lys Glu Ala Val
1100 1105 1110
Lys Lys Phe Asp Ala Ile Gly Trp Asp Gly Lys Glu Gln Ser Tyr
1115 1120 1125
Phe Phe Lys Tyr Asn Pro Tyr Asn Leu Ala Asp Glu Lys Tyr Lys
1130 1135 1140
Asn Ser Thr Val Ser Lys Glu Trp Ala Ile Phe Ala Ser Ala Pro
1145 1150 1155
Arg Ile Arg Arg Gln Lys Gly Glu Asp Gly Tyr Trp Lys Tyr Asp
1160 1165 1170
Arg Val Lys Val Asn Glu Glu Phe Glu Lys Leu Leu Lys Val Trp
1175 1180 1185
Asn Phe Val Asn Pro Lys Ala Thr Asp Ile Lys Gln Glu Ile Ile
1190 1195 1200
Lys Lys Ile Lys Ala Gly Asp Leu Gln Gly Glu Lys Glu Leu Asp
1205 1210 1215
Gly Arg Leu Arg Asn Phe Trp His Ser Phe Ile Tyr Leu Phe Asn
1220 1225 1230
Leu Val Leu Glu Leu Arg Asn Ser Phe Ser Leu Gln Ile Lys Ile
1235 1240 1245
Lys Ala Gly Glu Val Ile Ala Val Asp Glu Gly Val Asp Phe Ile
1250 1255 1260
Ala Ser Pro Val Lys Pro Phe Phe Thr Thr Pro Asn Pro Tyr Ile
1265 1270 1275
Pro Ser Asn Leu Cys Trp Leu Ala Val Glu Asn Ala Asp Ala Asn
1280 1285 1290
Gly Ala Tyr Asn Ile Ala Arg Lys Gly Val Met Ile Leu Lys Lys
1295 1300 1305
Ile Arg Glu His Ala Lys Lys Asp Pro Glu Phe Lys Lys Leu Pro
1310 1315 1320
Asn Leu Phe Ile Ser Asn Ala Glu Trp Asp Glu Ala Ala Arg Asp
1325 1330 1335
Trp Gly Lys Tyr Ala Gly Thr Thr Ala Leu Asn Leu Asp His
1340 1345 1350
<210> 13
<211> 1260
<212> PRT
<213> 狗口腔卟啉单胞菌（Porphyromonas crevioricanis）
<400> 13
Met Asp Ser Leu Lys Asp Phe Thr Asn Leu Tyr Pro Val Ser Lys Thr
1 5 10 15
Leu Arg Phe Glu Leu Lys Pro Val Gly Lys Thr Leu Glu Asn Ile Glu
20 25 30
Lys Ala Gly Ile Leu Lys Glu Asp Glu His Arg Ala Glu Ser Tyr Arg
35 40 45
Arg Val Lys Lys Ile Ile Asp Thr Tyr His Lys Val Phe Ile Asp Ser
50 55 60
Ser Leu Glu Asn Met Ala Lys Met Gly Ile Glu Asn Glu Ile Lys Ala
65 70 75 80
Met Leu Gln Ser Phe Cys Glu Leu Tyr Lys Lys Asp His Arg Thr Glu
85 90 95
Gly Glu Asp Lys Ala Leu Asp Lys Ile Arg Ala Val Leu Arg Gly Leu
100 105 110
Ile Val Gly Ala Phe Thr Gly Val Cys Gly Arg Arg Glu Asn Thr Val
115 120 125
Gln Asn Glu Lys Tyr Glu Ser Leu Phe Lys Glu Lys Leu Ile Lys Glu
130 135 140
Ile Leu Pro Asp Phe Val Leu Ser Thr Glu Ala Glu Ser Leu Pro Phe
145 150 155 160
Ser Val Glu Glu Ala Thr Arg Ser Leu Lys Glu Phe Asp Ser Phe Thr
165 170 175
Ser Tyr Phe Ala Gly Phe Tyr Glu Asn Arg Lys Asn Ile Tyr Ser Thr
180 185 190
Lys Pro Gln Ser Thr Ala Ile Ala Tyr Arg Leu Ile His Glu Asn Leu
195 200 205
Pro Lys Phe Ile Asp Asn Ile Leu Val Phe Gln Lys Ile Lys Glu Pro
210 215 220
Ile Ala Lys Glu Leu Glu His Ile Arg Ala Asp Phe Ser Ala Gly Gly
225 230 235 240
Tyr Ile Lys Lys Asp Glu Arg Leu Glu Asp Ile Phe Ser Leu Asn Tyr
245 250 255
Tyr Ile His Val Leu Ser Gln Ala Gly Ile Glu Lys Tyr Asn Ala Leu
260 265 270
Ile Gly Lys Ile Val Thr Glu Gly Asp Gly Glu Met Lys Gly Leu Asn
275 280 285
Glu His Ile Asn Leu Tyr Asn Gln Gln Arg Gly Arg Glu Asp Arg Leu
290 295 300
Pro Leu Phe Arg Pro Leu Tyr Lys Gln Ile Leu Ser Asp Arg Glu Gln
305 310 315 320
Leu Ser Tyr Leu Pro Glu Ser Phe Glu Lys Asp Glu Glu Leu Leu Arg
325 330 335
Ala Leu Lys Glu Phe Tyr Asp His Ile Ala Glu Asp Ile Leu Gly Arg
340 345 350
Thr Gln Gln Leu Met Thr Ser Ile Ser Glu Tyr Asp Leu Ser Arg Ile
355 360 365
Tyr Val Arg Asn Asp Ser Gln Leu Thr Asp Ile Ser Lys Lys Met Leu
370 375 380
Gly Asp Trp Asn Ala Ile Tyr Met Ala Arg Glu Arg Ala Tyr Asp His
385 390 395 400
Glu Gln Ala Pro Lys Arg Ile Thr Ala Lys Tyr Glu Arg Asp Arg Ile
405 410 415
Lys Ala Leu Lys Gly Glu Glu Ser Ile Ser Leu Ala Asn Leu Asn Ser
420 425 430
Cys Ile Ala Phe Leu Asp Asn Val Arg Asp Cys Arg Val Asp Thr Tyr
435 440 445
Leu Ser Thr Leu Gly Gln Lys Glu Gly Pro His Gly Leu Ser Asn Leu
450 455 460
Val Glu Asn Val Phe Ala Ser Tyr His Glu Ala Glu Gln Leu Leu Ser
465 470 475 480
Phe Pro Tyr Pro Glu Glu Asn Asn Leu Ile Gln Asp Lys Asp Asn Val
485 490 495
Val Leu Ile Lys Asn Leu Leu Asp Asn Ile Ser Asp Leu Gln Arg Phe
500 505 510
Leu Lys Pro Leu Trp Gly Met Gly Asp Glu Pro Asp Lys Asp Glu Arg
515 520 525
Phe Tyr Gly Glu Tyr Asn Tyr Ile Arg Gly Ala Leu Asp Gln Val Ile
530 535 540
Pro Leu Tyr Asn Lys Val Arg Asn Tyr Leu Thr Arg Lys Pro Tyr Ser
545 550 555 560
Thr Arg Lys Val Lys Leu Asn Phe Gly Asn Ser Gln Leu Leu Ser Gly
565 570 575
Trp Asp Arg Asn Lys Glu Lys Asp Asn Ser Cys Val Ile Leu Arg Lys
580 585 590
Gly Gln Asn Phe Tyr Leu Ala Ile Met Asn Asn Arg His Lys Arg Ser
595 600 605
Phe Glu Asn Lys Met Leu Pro Glu Tyr Lys Glu Gly Glu Pro Tyr Phe
610 615 620
Glu Lys Met Asp Tyr Lys Phe Leu Pro Asp Pro Asn Lys Met Leu Pro
625 630 635 640
Lys Val Phe Leu Ser Lys Lys Gly Ile Glu Ile Tyr Lys Pro Ser Pro
645 650 655
Lys Leu Leu Glu Gln Tyr Gly His Gly Thr His Lys Lys Gly Asp Thr
660 665 670
Phe Ser Met Asp Asp Leu His Glu Leu Ile Asp Phe Phe Lys His Ser
675 680 685
Ile Glu Ala His Glu Asp Trp Lys Gln Phe Gly Phe Lys Phe Ser Asp
690 695 700
Thr Ala Thr Tyr Glu Asn Val Ser Ser Phe Tyr Arg Glu Val Glu Asp
705 710 715 720
Gln Gly Tyr Lys Leu Ser Phe Arg Lys Val Ser Glu Ser Tyr Val Tyr
725 730 735
Ser Leu Ile Asp Gln Gly Lys Leu Tyr Leu Phe Gln Ile Tyr Asn Lys
740 745 750
Asp Phe Ser Pro Cys Ser Lys Gly Thr Pro Asn Leu His Thr Leu Tyr
755 760 765
Trp Arg Met Leu Phe Asp Glu Arg Asn Leu Ala Asp Val Ile Tyr Lys
770 775 780
Leu Asp Gly Lys Ala Glu Ile Phe Phe Arg Glu Lys Ser Leu Lys Asn
785 790 795 800
Asp His Pro Thr His Pro Ala Gly Lys Pro Ile Lys Lys Lys Ser Arg
805 810 815
Gln Lys Lys Gly Glu Glu Ser Leu Phe Glu Tyr Asp Leu Val Lys Asp
820 825 830
Arg Arg Tyr Thr Met Asp Lys Phe Gln Phe His Val Pro Ile Thr Met
835 840 845
Asn Phe Lys Cys Ser Ala Gly Ser Lys Val Asn Asp Met Val Asn Ala
850 855 860
His Ile Arg Glu Ala Lys Asp Met His Val Ile Gly Ile Asp Arg Gly
865 870 875 880
Glu Arg Asn Leu Leu Tyr Ile Cys Val Ile Asp Ser Arg Gly Thr Ile
885 890 895
Leu Asp Gln Ile Ser Leu Asn Thr Ile Asn Asp Ile Asp Tyr His Asp
900 905 910
Leu Leu Glu Ser Arg Asp Lys Asp Arg Gln Gln Glu His Arg Asn Trp
915 920 925
Gln Thr Ile Glu Gly Ile Lys Glu Leu Lys Gln Gly Tyr Leu Ser Gln
930 935 940
Ala Val His Arg Ile Ala Glu Leu Met Val Ala Tyr Lys Ala Val Val
945 950 955 960
Ala Leu Glu Asp Leu Asn Met Gly Phe Lys Arg Gly Arg Gln Lys Val
965 970 975
Glu Ser Ser Val Tyr Gln Gln Phe Glu Lys Gln Leu Ile Asp Lys Leu
980 985 990
Asn Tyr Leu Val Asp Lys Lys Lys Arg Pro Glu Asp Ile Gly Gly Leu
995 1000 1005
Leu Arg Ala Tyr Gln Phe Thr Ala Pro Phe Lys Ser Phe Lys Glu
1010 1015 1020
Met Gly Lys Gln Asn Gly Phe Leu Phe Tyr Ile Pro Ala Trp Asn
1025 1030 1035
Thr Ser Asn Ile Asp Pro Thr Thr Gly Phe Val Asn Leu Phe His
1040 1045 1050
Val Gln Tyr Glu Asn Val Asp Lys Ala Lys Ser Phe Phe Gln Lys
1055 1060 1065
Phe Asp Ser Ile Ser Tyr Asn Pro Lys Lys Asp Trp Phe Glu Phe
1070 1075 1080
Ala Phe Asp Tyr Lys Asn Phe Thr Lys Lys Ala Glu Gly Ser Arg
1085 1090 1095
Ser Met Trp Ile Leu Cys Thr His Gly Ser Arg Ile Lys Asn Phe
1100 1105 1110
Arg Asn Ser Gln Lys Asn Gly Gln Trp Asp Ser Glu Glu Phe Ala
1115 1120 1125
Leu Thr Glu Ala Phe Lys Ser Leu Phe Val Arg Tyr Glu Ile Asp
1130 1135 1140
Tyr Thr Ala Asp Leu Lys Thr Ala Ile Val Asp Glu Lys Gln Lys
1145 1150 1155
Asp Phe Phe Val Asp Leu Leu Lys Leu Phe Lys Leu Thr Val Gln
1160 1165 1170
Met Arg Asn Ser Trp Lys Glu Lys Asp Leu Asp Tyr Leu Ile Ser
1175 1180 1185
Pro Val Ala Gly Ala Asp Gly Arg Phe Phe Asp Thr Arg Glu Gly
1190 1195 1200
Asn Lys Ser Leu Pro Lys Asp Ala Asp Ala Asn Gly Ala Tyr Asn
1205 1210 1215
Ile Ala Leu Lys Gly Leu Trp Ala Leu Arg Gln Ile Arg Gln Thr
1220 1225 1230
Ser Glu Gly Gly Lys Leu Lys Leu Ala Ile Ser Asn Lys Glu Trp
1235 1240 1245
Leu Gln Phe Val Gln Glu Arg Ser Tyr Glu Lys Asp
1250 1255 1260
<210> 14
<211> 1324
<212> PRT
<213> 解糖胨普雷沃氏菌（Prevotella disiens）
<400> 14
Met Glu Asn Tyr Gln Glu Phe Thr Asn Leu Phe Gln Leu Asn Lys Thr
1 5 10 15
Leu Arg Phe Glu Leu Lys Pro Ile Gly Lys Thr Cys Glu Leu Leu Glu
20 25 30
Glu Gly Lys Ile Phe Ala Ser Gly Ser Phe Leu Glu Lys Asp Lys Val
35 40 45
Arg Ala Asp Asn Val Ser Tyr Val Lys Lys Glu Ile Asp Lys Lys His
50 55 60
Lys Ile Phe Ile Glu Glu Thr Leu Ser Ser Phe Ser Ile Ser Asn Asp
65 70 75 80
Leu Leu Lys Gln Tyr Phe Asp Cys Tyr Asn Glu Leu Lys Ala Phe Lys
85 90 95
Lys Asp Cys Lys Ser Asp Glu Glu Glu Val Lys Lys Thr Ala Leu Arg
100 105 110
Asn Lys Cys Thr Ser Ile Gln Arg Ala Met Arg Glu Ala Ile Ser Gln
115 120 125
Ala Phe Leu Lys Ser Pro Gln Lys Lys Leu Leu Ala Ile Lys Asn Leu
130 135 140
Ile Glu Asn Val Phe Lys Ala Asp Glu Asn Val Gln His Phe Ser Glu
145 150 155 160
Phe Thr Ser Tyr Phe Ser Gly Phe Glu Thr Asn Arg Glu Asn Phe Tyr
165 170 175
Ser Asp Glu Glu Lys Ser Thr Ser Ile Ala Tyr Arg Leu Val His Asp
180 185 190
Asn Leu Pro Ile Phe Ile Lys Asn Ile Tyr Ile Phe Glu Lys Leu Lys
195 200 205
Glu Gln Phe Asp Ala Lys Thr Leu Ser Glu Ile Phe Glu Asn Tyr Lys
210 215 220
Leu Tyr Val Ala Gly Ser Ser Leu Asp Glu Val Phe Ser Leu Glu Tyr
225 230 235 240
Phe Asn Asn Thr Leu Thr Gln Lys Gly Ile Asp Asn Tyr Asn Ala Val
245 250 255
Ile Gly Lys Ile Val Lys Glu Asp Lys Gln Glu Ile Gln Gly Leu Asn
260 265 270
Glu His Ile Asn Leu Tyr Asn Gln Lys His Lys Asp Arg Arg Leu Pro
275 280 285
Phe Phe Ile Ser Leu Lys Lys Gln Ile Leu Ser Asp Arg Glu Ala Leu
290 295 300
Ser Trp Leu Pro Asp Met Phe Lys Asn Asp Ser Glu Val Ile Asp Ala
305 310 315 320
Leu Lys Gly Phe Tyr Ile Glu Asp Gly Phe Glu Asn Asn Val Leu Thr
325 330 335
Pro Leu Ala Thr Leu Leu Ser Ser Leu Asp Lys Tyr Asn Leu Asn Gly
340 345 350
Ile Phe Ile Arg Asn Asn Glu Ala Leu Ser Ser Leu Ser Gln Asn Val
355 360 365
Tyr Arg Asn Phe Ser Ile Asp Glu Ala Ile Asp Ala Gln Asn Ala Glu
370 375 380
Leu Gln Thr Phe Asn Asn Tyr Glu Leu Ile Ala Asn Ala Leu Arg Ala
385 390 395 400
Lys Ile Lys Lys Glu Thr Lys Gln Gly Arg Lys Ser Phe Glu Lys Tyr
405 410 415
Glu Glu Tyr Ile Asp Lys Lys Val Lys Ala Ile Asp Ser Leu Ser Ile
420 425 430
Gln Glu Ile Asn Glu Leu Val Glu Asn Tyr Val Ser Glu Phe Asn Ser
435 440 445
Asn Ser Gly Asn Met Pro Arg Lys Val Glu Asp Tyr Phe Ser Leu Met
450 455 460
Arg Lys Gly Asp Phe Gly Ser Asn Asp Leu Ile Glu Asn Ile Lys Thr
465 470 475 480
Lys Leu Ser Ala Ala Glu Lys Leu Leu Gly Thr Lys Tyr Gln Glu Thr
485 490 495
Ala Lys Asp Ile Phe Lys Lys Asp Glu Asn Ser Lys Leu Ile Lys Glu
500 505 510
Leu Leu Asp Ala Thr Lys Gln Phe Gln His Phe Ile Lys Pro Leu Leu
515 520 525
Gly Thr Gly Glu Glu Ala Asp Arg Asp Leu Val Phe Tyr Gly Asp Phe
530 535 540
Leu Pro Leu Tyr Glu Lys Phe Glu Glu Leu Thr Leu Leu Tyr Asn Lys
545 550 555 560
Val Arg Asn Arg Leu Thr Gln Lys Pro Tyr Ser Lys Asp Lys Ile Arg
565 570 575
Leu Cys Phe Asn Lys Pro Lys Leu Met Thr Gly Trp Val Asp Ser Lys
580 585 590
Thr Glu Lys Ser Asp Asn Gly Thr Gln Tyr Gly Gly Tyr Leu Phe Arg
595 600 605
Lys Lys Asn Glu Ile Gly Glu Tyr Asp Tyr Phe Leu Gly Ile Ser Ser
610 615 620
Lys Ala Gln Leu Phe Arg Lys Asn Glu Ala Val Ile Gly Asp Tyr Glu
625 630 635 640
Arg Leu Asp Tyr Tyr Gln Pro Lys Ala Asn Thr Ile Tyr Gly Ser Ala
645 650 655
Tyr Glu Gly Glu Asn Ser Tyr Lys Glu Asp Lys Lys Arg Leu Asn Lys
660 665 670
Val Ile Ile Ala Tyr Ile Glu Gln Ile Lys Gln Thr Asn Ile Lys Lys
675 680 685
Ser Ile Ile Glu Ser Ile Ser Lys Tyr Pro Asn Ile Ser Asp Asp Asp
690 695 700
Lys Val Thr Pro Ser Ser Leu Leu Glu Lys Ile Lys Lys Val Ser Ile
705 710 715 720
Asp Ser Tyr Asn Gly Ile Leu Ser Phe Lys Ser Phe Gln Ser Val Asn
725 730 735
Lys Glu Val Ile Asp Asn Leu Leu Lys Thr Ile Ser Pro Leu Lys Asn
740 745 750
Lys Ala Glu Phe Leu Asp Leu Ile Asn Lys Asp Tyr Gln Ile Phe Thr
755 760 765
Glu Val Gln Ala Val Ile Asp Glu Ile Cys Lys Gln Lys Thr Phe Ile
770 775 780
Tyr Phe Pro Ile Ser Asn Val Glu Leu Glu Lys Glu Met Gly Asp Lys
785 790 795 800
Asp Lys Pro Leu Cys Leu Phe Gln Ile Ser Asn Lys Asp Leu Ser Phe
805 810 815
Ala Lys Thr Phe Ser Ala Asn Leu Arg Lys Lys Arg Gly Ala Glu Asn
820 825 830
Leu His Thr Met Leu Phe Lys Ala Leu Met Glu Gly Asn Gln Asp Asn
835 840 845
Leu Asp Leu Gly Ser Gly Ala Ile Phe Tyr Arg Ala Lys Ser Leu Asp
850 855 860
Gly Asn Lys Pro Thr His Pro Ala Asn Glu Ala Ile Lys Cys Arg Asn
865 870 875 880
Val Ala Asn Lys Asp Lys Val Ser Leu Phe Thr Tyr Asp Ile Tyr Lys
885 890 895
Asn Arg Arg Tyr Met Glu Asn Lys Phe Leu Phe His Leu Ser Ile Val
900 905 910
Gln Asn Tyr Lys Ala Ala Asn Asp Ser Ala Gln Leu Asn Ser Ser Ala
915 920 925
Thr Glu Tyr Ile Arg Lys Ala Asp Asp Leu His Ile Ile Gly Ile Asp
930 935 940
Arg Gly Glu Arg Asn Leu Leu Tyr Tyr Ser Val Ile Asp Met Lys Gly
945 950 955 960
Asn Ile Val Glu Gln Asp Ser Leu Asn Ile Ile Arg Asn Asn Asp Leu
965 970 975
Glu Thr Asp Tyr His Asp Leu Leu Asp Lys Arg Glu Lys Glu Arg Lys
980 985 990
Ala Asn Arg Gln Asn Trp Glu Ala Val Glu Gly Ile Lys Asp Leu Lys
995 1000 1005
Lys Gly Tyr Leu Ser Gln Ala Val His Gln Ile Ala Gln Leu Met
1010 1015 1020
Leu Lys Tyr Asn Ala Ile Ile Ala Leu Glu Asp Leu Gly Gln Met
1025 1030 1035
Phe Val Thr Arg Gly Gln Lys Ile Glu Lys Ala Val Tyr Gln Gln
1040 1045 1050
Phe Glu Lys Ser Leu Val Asp Lys Leu Ser Tyr Leu Val Asp Lys
1055 1060 1065
Lys Arg Pro Tyr Asn Glu Leu Gly Gly Ile Leu Lys Ala Tyr Gln
1070 1075 1080
Leu Ala Ser Ser Ile Thr Lys Asn Asn Ser Asp Lys Gln Asn Gly
1085 1090 1095
Phe Leu Phe Tyr Val Pro Ala Trp Asn Thr Ser Lys Ile Asp Pro
1100 1105 1110
Val Thr Gly Phe Thr Asp Leu Leu Arg Pro Lys Ala Met Thr Ile
1115 1120 1125
Lys Glu Ala Gln Asp Phe Phe Gly Ala Phe Asp Asn Ile Ser Tyr
1130 1135 1140
Asn Asp Lys Gly Tyr Phe Glu Phe Glu Thr Asn Tyr Asp Lys Phe
1145 1150 1155
Lys Ile Arg Met Lys Ser Ala Gln Thr Arg Trp Thr Ile Cys Thr
1160 1165 1170
Phe Gly Asn Arg Ile Lys Arg Lys Lys Asp Lys Asn Tyr Trp Asn
1175 1180 1185
Tyr Glu Glu Val Glu Leu Thr Glu Glu Phe Lys Lys Leu Phe Lys
1190 1195 1200
Asp Ser Asn Ile Asp Tyr Glu Asn Cys Asn Leu Lys Glu Glu Ile
1205 1210 1215
Gln Asn Lys Asp Asn Arg Lys Phe Phe Asp Asp Leu Ile Lys Leu
1220 1225 1230
Leu Gln Leu Thr Leu Gln Met Arg Asn Ser Asp Asp Lys Gly Asn
1235 1240 1245
Asp Tyr Ile Ile Ser Pro Val Ala Asn Ala Glu Gly Gln Phe Phe
1250 1255 1260
Asp Ser Arg Asn Gly Asp Lys Lys Leu Pro Leu Asp Ala Asp Ala
1265 1270 1275
Asn Gly Ala Tyr Asn Ile Ala Arg Lys Gly Leu Trp Asn Ile Arg
1280 1285 1290
Gln Ile Lys Gln Thr Lys Asn Lys Asp Asp Leu Asn Leu Ser Ile
1295 1300 1305
Ser Ser Thr Glu Trp Leu Asp Phe Val Arg Glu Lys Pro Tyr Leu
1310 1315 1320
Lys
<210> 15
<211> 1484
<212> PRT
<213> Unknown
<220>
<223> Peregrinibacteria细菌
<220>
<221> misc_feature
<222> (1073)..(1073)
<223> Xaa可以为任何天然出现的氨基酸
<400> 15
Met Ser Asn Phe Phe Lys Asn Phe Thr Asn Leu Tyr Glu Leu Ser Lys
1 5 10 15
Thr Leu Arg Phe Glu Leu Lys Pro Val Gly Asp Thr Leu Thr Asn Met
20 25 30
Lys Asp His Leu Glu Tyr Asp Glu Lys Leu Gln Thr Phe Leu Lys Asp
35 40 45
Gln Asn Ile Asp Asp Ala Tyr Gln Ala Leu Lys Pro Gln Phe Asp Glu
50 55 60
Ile His Glu Glu Phe Ile Thr Asp Ser Leu Glu Ser Lys Lys Ala Lys
65 70 75 80
Glu Ile Asp Phe Ser Glu Tyr Leu Asp Leu Phe Gln Glu Lys Lys Glu
85 90 95
Leu Asn Asp Ser Glu Lys Lys Leu Arg Asn Lys Ile Gly Glu Thr Phe
100 105 110
Asn Lys Ala Gly Glu Lys Trp Lys Lys Glu Lys Tyr Pro Gln Tyr Glu
115 120 125
Trp Lys Lys Gly Ser Lys Ile Ala Asn Gly Ala Asp Ile Leu Ser Cys
130 135 140
Gln Asp Met Leu Gln Phe Ile Lys Tyr Lys Asn Pro Glu Asp Glu Lys
145 150 155 160
Ile Lys Asn Tyr Ile Asp Asp Thr Leu Lys Gly Phe Phe Thr Tyr Phe
165 170 175
Gly Gly Phe Asn Gln Asn Arg Ala Asn Tyr Tyr Glu Thr Lys Lys Glu
180 185 190
Ala Ser Thr Ala Val Ala Thr Arg Ile Val His Glu Asn Leu Pro Lys
195 200 205
Phe Cys Asp Asn Val Ile Gln Phe Lys His Ile Ile Lys Arg Lys Lys
210 215 220
Asp Gly Thr Val Glu Lys Thr Glu Arg Lys Thr Glu Tyr Leu Asn Ala
225 230 235 240
Tyr Gln Tyr Leu Lys Asn Asn Asn Lys Ile Thr Gln Ile Lys Asp Ala
245 250 255
Glu Thr Glu Lys Met Ile Glu Ser Thr Pro Ile Ala Glu Lys Ile Phe
260 265 270
Asp Val Tyr Tyr Phe Ser Ser Cys Leu Ser Gln Lys Gln Ile Glu Glu
275 280 285
Tyr Asn Arg Ile Ile Gly His Tyr Asn Leu Leu Ile Asn Leu Tyr Asn
290 295 300
Gln Ala Lys Arg Ser Glu Gly Lys His Leu Ser Ala Asn Glu Lys Lys
305 310 315 320
Tyr Lys Asp Leu Pro Lys Phe Lys Thr Leu Tyr Lys Gln Ile Gly Cys
325 330 335
Gly Lys Lys Lys Asp Leu Phe Tyr Thr Ile Lys Cys Asp Thr Glu Glu
340 345 350
Glu Ala Asn Lys Ser Arg Asn Glu Gly Lys Glu Ser His Ser Val Glu
355 360 365
Glu Ile Ile Asn Lys Ala Gln Glu Ala Ile Asn Lys Tyr Phe Lys Ser
370 375 380
Asn Asn Asp Cys Glu Asn Ile Asn Thr Val Pro Asp Phe Ile Asn Tyr
385 390 395 400
Ile Leu Thr Lys Glu Asn Tyr Glu Gly Val Tyr Trp Ser Lys Ala Ala
405 410 415
Met Asn Thr Ile Ser Asp Lys Tyr Phe Ala Asn Tyr His Asp Leu Gln
420 425 430
Asp Arg Leu Lys Glu Ala Lys Val Phe Gln Lys Ala Asp Lys Lys Ser
435 440 445
Glu Asp Asp Ile Lys Ile Pro Glu Ala Ile Glu Leu Ser Gly Leu Phe
450 455 460
Gly Val Leu Asp Ser Leu Ala Asp Trp Gln Thr Thr Leu Phe Lys Ser
465 470 475 480
Ser Ile Leu Ser Asn Glu Lys Leu Lys Ile Ile Thr Asp Ser Gln Thr
485 490 495
Pro Ser Glu Ala Leu Leu Lys Met Ile Phe Asn Asp Ile Glu Lys Asn
500 505 510
Met Glu Ser Phe Leu Lys Glu Thr Asn Asp Ile Ile Thr Leu Lys Lys
515 520 525
Tyr Lys Gly Asn Lys Glu Gly Thr Glu Lys Ile Lys Gln Trp Phe Asp
530 535 540
Tyr Thr Leu Ala Ile Asn Arg Met Leu Lys Tyr Phe Leu Val Lys Glu
545 550 555 560
Asn Lys Ile Lys Gly Asn Ser Leu Asp Thr Asn Ile Ser Glu Ala Leu
565 570 575
Lys Thr Leu Ile Tyr Ser Asp Asp Ala Glu Trp Phe Lys Trp Tyr Asp
580 585 590
Ala Leu Arg Asn Tyr Leu Thr Gln Lys Pro Gln Asp Glu Ala Lys Glu
595 600 605
Asn Lys Leu Lys Leu Asn Phe Asp Asn Pro Ser Leu Ala Gly Gly Trp
610 615 620
Asp Val Asn Lys Glu Cys Ser Asn Phe Cys Val Ile Leu Lys Asp Lys
625 630 635 640
Asn Glu Lys Lys Tyr Leu Ala Met Ile Lys Lys Gly Glu Asn Thr Leu
645 650 655
Phe Gln Lys Glu Trp Thr Glu Gly Arg Gly Lys Asn Leu Thr Lys Lys
660 665 670
Ser Asn Pro Leu Phe Glu Ile Asn Asn Cys Glu Ile Leu Ser Lys Met
675 680 685
Glu Tyr Asp Phe Trp Ala Asp Val Ser Lys Met Ile Pro Lys Cys Ser
690 695 700
Thr Gln Leu Lys Ala Val Val Asn His Phe Lys Gln Ser Asp Asn Glu
705 710 715 720
Phe Ile Phe Pro Ile Gly Tyr Lys Val Thr Ser Gly Glu Lys Phe Arg
725 730 735
Glu Glu Cys Lys Ile Ser Lys Gln Asp Phe Glu Leu Asn Asn Lys Val
740 745 750
Phe Asn Lys Asn Glu Leu Ser Val Thr Ala Met Arg Tyr Asp Leu Ser
755 760 765
Ser Thr Gln Glu Lys Gln Tyr Ile Lys Ala Phe Gln Lys Glu Tyr Trp
770 775 780
Glu Leu Leu Phe Lys Gln Glu Lys Arg Asp Thr Lys Leu Thr Asn Asn
785 790 795 800
Glu Ile Phe Asn Glu Trp Ile Asn Phe Cys Asn Lys Lys Tyr Ser Glu
805 810 815
Leu Leu Ser Trp Glu Arg Lys Tyr Lys Asp Ala Leu Thr Asn Trp Ile
820 825 830
Asn Phe Cys Lys Tyr Phe Leu Ser Lys Tyr Pro Lys Thr Thr Leu Phe
835 840 845
Asn Tyr Ser Phe Lys Glu Ser Glu Asn Tyr Asn Ser Leu Asp Glu Phe
850 855 860
Tyr Arg Asp Val Asp Ile Cys Ser Tyr Lys Leu Asn Ile Asn Thr Thr
865 870 875 880
Ile Asn Lys Ser Ile Leu Asp Arg Leu Val Glu Glu Gly Lys Leu Tyr
885 890 895
Leu Phe Glu Ile Lys Asn Gln Asp Ser Asn Asp Gly Lys Ser Ile Gly
900 905 910
His Lys Asn Asn Leu His Thr Ile Tyr Trp Asn Ala Ile Phe Glu Asn
915 920 925
Phe Asp Asn Arg Pro Lys Leu Asn Gly Glu Ala Glu Ile Phe Tyr Arg
930 935 940
Lys Ala Ile Ser Lys Asp Lys Leu Gly Ile Val Lys Gly Lys Lys Thr
945 950 955 960
Lys Asn Gly Thr Trp Ile Ile Lys Asn Tyr Arg Phe Ser Lys Glu Lys
965 970 975
Phe Ile Leu His Val Pro Ile Thr Leu Asn Phe Cys Ser Asn Asn Glu
980 985 990
Tyr Val Asn Asp Ile Val Asn Thr Lys Phe Tyr Asn Phe Ser Asn Leu
995 1000 1005
His Phe Leu Gly Ile Asp Arg Gly Glu Lys His Leu Ala Tyr Tyr
1010 1015 1020
Ser Leu Val Asn Lys Asn Gly Glu Ile Val Asp Gln Gly Thr Leu
1025 1030 1035
Asn Leu Pro Phe Thr Asp Lys Asp Gly Asn Gln Arg Ser Ile Lys
1040 1045 1050
Lys Glu Lys Tyr Phe Tyr Asn Lys Gln Glu Asp Lys Trp Glu Ala
1055 1060 1065
Lys Glu Val Asp Xaa Trp Asn Tyr Asn Asp Leu Leu Asp Ala Met
1070 1075 1080
Ala Ser Asn Arg Asp Met Ala Arg Lys Asn Trp Gln Arg Ile Gly
1085 1090 1095
Thr Ile Lys Glu Ala Lys Asn Gly Tyr Val Ser Leu Val Ile Arg
1100 1105 1110
Lys Ile Ala Asp Leu Ala Val Asn Asn Glu Arg Pro Ala Phe Ile
1115 1120 1125
Val Leu Glu Asp Leu Asn Thr Gly Phe Lys Arg Ser Arg Gln Lys
1130 1135 1140
Ile Asp Lys Ser Val Tyr Gln Lys Phe Glu Leu Ala Leu Ala Lys
1145 1150 1155
Lys Leu Asn Phe Leu Val Asp Lys Asn Ala Lys Arg Asp Glu Ile
1160 1165 1170
Gly Ser Pro Thr Lys Ala Leu Gln Leu Thr Pro Pro Val Asn Asn
1175 1180 1185
Tyr Gly Asp Ile Glu Asn Lys Lys Gln Ala Gly Ile Met Leu Tyr
1190 1195 1200
Thr Arg Ala Asn Tyr Thr Ser Gln Thr Asp Pro Ala Thr Gly Trp
1205 1210 1215
Arg Lys Thr Ile Tyr Leu Lys Ala Gly Pro Glu Glu Thr Thr Tyr
1220 1225 1230
Lys Lys Asp Gly Lys Ile Lys Asn Lys Ser Val Lys Asp Gln Ile
1235 1240 1245
Ile Glu Thr Phe Thr Asp Ile Gly Phe Asp Gly Lys Asp Tyr Tyr
1250 1255 1260
Phe Glu Tyr Asp Lys Gly Glu Phe Val Asp Glu Lys Thr Gly Glu
1265 1270 1275
Ile Lys Pro Lys Lys Trp Arg Leu Tyr Ser Gly Glu Asn Gly Lys
1280 1285 1290
Ser Leu Asp Arg Phe Arg Gly Glu Arg Glu Lys Asp Lys Tyr Glu
1295 1300 1305
Trp Lys Ile Asp Lys Ile Asp Ile Val Lys Ile Leu Asp Asp Leu
1310 1315 1320
Phe Val Asn Phe Asp Lys Asn Ile Ser Leu Leu Lys Gln Leu Lys
1325 1330 1335
Glu Gly Val Glu Leu Thr Arg Asn Asn Glu His Gly Thr Gly Glu
1340 1345 1350
Ser Leu Arg Phe Ala Ile Asn Leu Ile Gln Gln Ile Arg Asn Thr
1355 1360 1365
Gly Asn Asn Glu Arg Asp Asn Asp Phe Ile Leu Ser Pro Val Arg
1370 1375 1380
Asp Glu Asn Gly Lys His Phe Asp Ser Arg Glu Tyr Trp Asp Lys
1385 1390 1395
Glu Thr Lys Gly Glu Lys Ile Ser Met Pro Ser Ser Gly Asp Ala
1400 1405 1410
Asn Gly Ala Phe Asn Ile Ala Arg Lys Gly Ile Ile Met Asn Ala
1415 1420 1425
His Ile Leu Ala Asn Ser Asp Ser Lys Asp Leu Ser Leu Phe Val
1430 1435 1440
Ser Asp Glu Glu Trp Asp Leu His Leu Asn Asn Lys Thr Glu Trp
1445 1450 1455
Lys Lys Gln Leu Asn Ile Phe Ser Ser Arg Lys Ala Met Ala Lys
1460 1465 1470
Arg Lys Lys Lys Arg Pro Ala Ala Thr Lys Lys
1475 1480
<210> 16
<211> 1245
<212> PRT
<213> 猕猴卟啉单胞菌（Porphyromonas macacae）
<400> 16
Met Lys Thr Gln His Phe Phe Glu Asp Phe Thr Ser Leu Tyr Ser Leu
1 5 10 15
Ser Lys Thr Ile Arg Phe Glu Leu Lys Pro Ile Gly Lys Thr Leu Glu
20 25 30
Asn Ile Lys Lys Asn Gly Leu Ile Arg Arg Asp Glu Gln Arg Leu Asp
35 40 45
Asp Tyr Glu Lys Leu Lys Lys Val Ile Asp Glu Tyr His Glu Asp Phe
50 55 60
Ile Ala Asn Ile Leu Ser Ser Phe Ser Phe Ser Glu Glu Ile Leu Gln
65 70 75 80
Ser Tyr Ile Gln Asn Leu Ser Ile Ser Glu Ala Arg Ala Lys Ile Glu
85 90 95
Lys Thr Met Arg Asp Thr Leu Ala Lys Ala Phe Ser Glu Asp Glu Arg
100 105 110
Tyr Lys Ser Ile Phe Lys Lys Glu Leu Val Lys Lys Asp Ile Pro Val
115 120 125
Trp Cys Pro Ala Tyr Lys Ser Leu Cys Lys Lys Phe Asp Asn Phe Thr
130 135 140
Thr Ser Leu Val Pro Phe His Glu Asn Arg Lys Asn Leu Tyr Thr Ser
145 150 155 160
Asn Glu Ile Thr Ala Ser Ile Pro Tyr Arg Ile Val His Val Asn Leu
165 170 175
Pro Lys Phe Ile Gln Asn Ile Glu Ala Leu Cys Glu Leu Gln Lys Lys
180 185 190
Met Gly Ala Asp Leu Tyr Leu Glu Met Met Glu Asn Leu Arg Asn Val
195 200 205
Trp Pro Ser Phe Val Lys Thr Pro Asp Asp Leu Cys Asn Leu Lys Thr
210 215 220
Tyr Asn His Leu Met Val Gln Ser Ser Ile Ser Glu Tyr Asn Arg Phe
225 230 235 240
Val Gly Gly Tyr Ser Thr Glu Asp Gly Thr Lys His Gln Gly Ile Asn
245 250 255
Glu Trp Ile Asn Ile Tyr Arg Gln Arg Asn Lys Glu Met Arg Leu Pro
260 265 270
Gly Leu Val Phe Leu His Lys Gln Ile Leu Ala Lys Val Asp Ser Ser
275 280 285
Ser Phe Ile Ser Asp Thr Leu Glu Asn Asp Asp Gln Val Phe Cys Val
290 295 300
Leu Arg Gln Phe Arg Lys Leu Phe Trp Asn Thr Val Ser Ser Lys Glu
305 310 315 320
Asp Asp Ala Ala Ser Leu Lys Asp Leu Phe Cys Gly Leu Ser Gly Tyr
325 330 335
Asp Pro Glu Ala Ile Tyr Val Ser Asp Ala His Leu Ala Thr Ile Ser
340 345 350
Lys Asn Ile Phe Asp Arg Trp Asn Tyr Ile Ser Asp Ala Ile Arg Arg
355 360 365
Lys Thr Glu Val Leu Met Pro Arg Lys Lys Glu Ser Val Glu Arg Tyr
370 375 380
Ala Glu Lys Ile Ser Lys Gln Ile Lys Lys Arg Gln Ser Tyr Ser Leu
385 390 395 400
Ala Glu Leu Asp Asp Leu Leu Ala His Tyr Ser Glu Glu Ser Leu Pro
405 410 415
Ala Gly Phe Ser Leu Leu Ser Tyr Phe Thr Ser Leu Gly Gly Gln Lys
420 425 430
Tyr Leu Val Ser Asp Gly Glu Val Ile Leu Tyr Glu Glu Gly Ser Asn
435 440 445
Ile Trp Asp Glu Val Leu Ile Ala Phe Arg Asp Leu Gln Val Ile Leu
450 455 460
Asp Lys Asp Phe Thr Glu Lys Lys Leu Gly Lys Asp Glu Glu Ala Val
465 470 475 480
Ser Val Ile Lys Lys Ala Leu Asp Ser Ala Leu Arg Leu Arg Lys Phe
485 490 495
Phe Asp Leu Leu Ser Gly Thr Gly Ala Glu Ile Arg Arg Asp Ser Ser
500 505 510
Phe Tyr Ala Leu Tyr Thr Asp Arg Met Asp Lys Leu Lys Gly Leu Leu
515 520 525
Lys Met Tyr Asp Lys Val Arg Asn Tyr Leu Thr Lys Lys Pro Tyr Ser
530 535 540
Ile Glu Lys Phe Lys Leu His Phe Asp Asn Pro Ser Leu Leu Ser Gly
545 550 555 560
Trp Asp Lys Asn Lys Glu Leu Asn Asn Leu Ser Val Ile Phe Arg Gln
565 570 575
Asn Gly Tyr Tyr Tyr Leu Gly Ile Met Thr Pro Lys Gly Lys Asn Leu
580 585 590
Phe Lys Thr Leu Pro Lys Leu Gly Ala Glu Glu Met Phe Tyr Glu Lys
595 600 605
Met Glu Tyr Lys Gln Ile Ala Glu Pro Met Leu Met Leu Pro Lys Val
610 615 620
Phe Phe Pro Lys Lys Thr Lys Pro Ala Phe Ala Pro Asp Gln Ser Val
625 630 635 640
Val Asp Ile Tyr Asn Lys Lys Thr Phe Lys Thr Gly Gln Lys Gly Phe
645 650 655
Asn Lys Lys Asp Leu Tyr Arg Leu Ile Asp Phe Tyr Lys Glu Ala Leu
660 665 670
Thr Val His Glu Trp Lys Leu Phe Asn Phe Ser Phe Ser Pro Thr Glu
675 680 685
Gln Tyr Arg Asn Ile Gly Glu Phe Phe Asp Glu Val Arg Glu Gln Ala
690 695 700
Tyr Lys Val Ser Met Val Asn Val Pro Ala Ser Tyr Ile Asp Glu Ala
705 710 715 720
Val Glu Asn Gly Lys Leu Tyr Leu Phe Gln Ile Tyr Asn Lys Asp Phe
725 730 735
Ser Pro Tyr Ser Lys Gly Ile Pro Asn Leu His Thr Leu Tyr Trp Lys
740 745 750
Ala Leu Phe Ser Glu Gln Asn Gln Ser Arg Val Tyr Lys Leu Cys Gly
755 760 765
Gly Gly Glu Leu Phe Tyr Arg Lys Ala Ser Leu His Met Gln Asp Thr
770 775 780
Thr Val His Pro Lys Gly Ile Ser Ile His Lys Lys Asn Leu Asn Lys
785 790 795 800
Lys Gly Glu Thr Ser Leu Phe Asn Tyr Asp Leu Val Lys Asp Lys Arg
805 810 815
Phe Thr Glu Asp Lys Phe Phe Phe His Val Pro Ile Ser Ile Asn Tyr
820 825 830
Lys Asn Lys Lys Ile Thr Asn Val Asn Gln Met Val Arg Asp Tyr Ile
835 840 845
Ala Gln Asn Asp Asp Leu Gln His Gly Ile Asp Arg Gly Glu Arg Asn
850 855 860
Leu Leu Tyr Ile Ser Arg Ile Asp Thr Arg Gly Asn Leu Leu Glu Gln
865 870 875 880
Phe Ser Leu Asn Val Ile Glu Ser Asp Lys Gly Asp Leu Arg Thr Asp
885 890 895
Tyr Gln Lys Ile Leu Gly Asp Arg Glu Gln Glu Arg Leu Arg Arg Arg
900 905 910
Gln Glu Trp Lys Ser Ile Glu Ser Ile Lys Asp Leu Lys Asp Gly Tyr
915 920 925
Met Ser Gln Val Val His Lys Ile Cys Asn Met Val Val Glu His Lys
930 935 940
Ala Ile Val Val Leu Glu Asn Leu Asn Leu Ser Phe Met Lys Gly Arg
945 950 955 960
Lys Lys Val Glu Lys Ser Val Tyr Glu Lys Phe Glu Arg Met Leu Val
965 970 975
Asp Lys Leu Asn Tyr Leu Val Val Asp Lys Lys Asn Leu Ser Asn Glu
980 985 990
Pro Gly Gly Leu Tyr Ala Ala Tyr Gln Leu Thr Asn Pro Leu Phe Ser
995 1000 1005
Phe Glu Glu Leu His Arg Tyr Pro Gln Ser Gly Ile Leu Phe Phe
1010 1015 1020
Val Asp Pro Trp Asn Thr Ser Leu Thr Asp Pro Ser Thr Gly Phe
1025 1030 1035
Val Asn Leu Leu Gly Arg Ile Asn Tyr Thr Asn Val Gly Asp Ala
1040 1045 1050
Arg Lys Phe Phe Asp Arg Phe Asn Ala Ile Arg Tyr Asp Gly Lys
1055 1060 1065
Gly Asn Ile Leu Phe Asp Leu Asp Leu Ser Arg Phe Asp Val Arg
1070 1075 1080
Val Glu Thr Gln Arg Lys Leu Trp Thr Leu Thr Thr Phe Gly Ser
1085 1090 1095
Arg Ile Ala Lys Ser Lys Lys Ser Gly Lys Trp Met Val Glu Arg
1100 1105 1110
Ile Glu Asn Leu Ser Leu Cys Phe Leu Glu Leu Phe Glu Gln Phe
1115 1120 1125
Asn Ile Gly Tyr Arg Val Glu Lys Asp Leu Lys Lys Ala Ile Leu
1130 1135 1140
Ser Gln Asp Arg Lys Glu Phe Tyr Val Arg Leu Ile Tyr Leu Phe
1145 1150 1155
Asn Leu Met Met Gln Ile Arg Asn Ser Asp Gly Glu Glu Asp Tyr
1160 1165 1170
Ile Leu Ser Pro Ala Leu Asn Glu Lys Asn Leu Gln Phe Asp Ser
1175 1180 1185
Arg Leu Ile Glu Ala Lys Asp Leu Pro Val Asp Ala Asp Ala Asn
1190 1195 1200
Gly Ala Tyr Asn Val Ala Arg Lys Gly Leu Met Val Val Gln Arg
1205 1210 1215
Ile Lys Arg Gly Asp His Glu Ser Ile His Arg Ile Gly Arg Ala
1220 1225 1230
Gln Trp Leu Arg Tyr Val Gln Glu Gly Ile Val Glu
1235 1240 1245
<210> 17
<211> 1250
<212> PRT
<213> 史密斯氏菌属物种（Smithella sp.）
<400> 17
Met Gln Thr Leu Phe Glu Asn Phe Thr Asn Gln Tyr Pro Val Ser Lys
1 5 10 15
Thr Leu Arg Phe Glu Leu Ile Pro Gln Gly Lys Thr Lys Asp Phe Ile
20 25 30
Glu Gln Lys Gly Leu Leu Lys Lys Asp Glu Asp Arg Ala Glu Lys Tyr
35 40 45
Lys Lys Val Lys Asn Ile Ile Asp Glu Tyr His Lys Asp Phe Ile Glu
50 55 60
Lys Ser Leu Asn Gly Leu Lys Leu Asp Gly Leu Glu Lys Tyr Lys Thr
65 70 75 80
Leu Tyr Leu Lys Gln Glu Lys Asp Asp Lys Asp Lys Lys Ala Phe Asp
85 90 95
Lys Glu Lys Glu Asn Leu Arg Lys Gln Ile Ala Asn Ala Phe Arg Asn
100 105 110
Asn Glu Lys Phe Lys Thr Leu Phe Ala Lys Glu Leu Ile Lys Asn Asp
115 120 125
Leu Met Ser Phe Ala Cys Glu Glu Asp Lys Lys Asn Val Lys Glu Phe
130 135 140
Glu Ala Phe Thr Thr Tyr Phe Thr Gly Phe His Gln Asn Arg Ala Asn
145 150 155 160
Met Tyr Val Ala Asp Glu Lys Arg Thr Ala Ile Ala Ser Arg Leu Ile
165 170 175
His Glu Asn Leu Pro Lys Phe Ile Asp Asn Ile Lys Ile Phe Glu Lys
180 185 190
Met Lys Lys Glu Ala Pro Glu Leu Leu Ser Pro Phe Asn Gln Thr Leu
195 200 205
Lys Asp Met Lys Asp Val Ile Lys Gly Thr Thr Leu Glu Glu Ile Phe
210 215 220
Ser Leu Asp Tyr Phe Asn Lys Thr Leu Thr Gln Ser Gly Ile Asp Ile
225 230 235 240
Tyr Asn Ser Val Ile Gly Gly Arg Thr Pro Glu Glu Gly Lys Thr Lys
245 250 255
Ile Lys Gly Leu Asn Glu Tyr Ile Asn Thr Asp Phe Asn Gln Lys Gln
260 265 270
Thr Asp Lys Lys Lys Arg Gln Pro Lys Phe Lys Gln Leu Tyr Lys Gln
275 280 285
Ile Leu Ser Asp Arg Gln Ser Leu Ser Phe Ile Ala Glu Ala Phe Lys
290 295 300
Asn Asp Thr Glu Ile Leu Glu Ala Ile Glu Lys Phe Tyr Val Asn Glu
305 310 315 320
Leu Leu His Phe Ser Asn Glu Gly Lys Ser Thr Asn Val Leu Asp Ala
325 330 335
Ile Lys Asn Ala Val Ser Asn Leu Glu Ser Phe Asn Leu Thr Lys Met
340 345 350
Tyr Phe Arg Ser Gly Ala Ser Leu Thr Asp Val Ser Arg Lys Val Phe
355 360 365
Gly Glu Trp Ser Ile Ile Asn Arg Ala Leu Asp Asn Tyr Tyr Ala Thr
370 375 380
Thr Tyr Pro Ile Lys Pro Arg Glu Lys Ser Glu Lys Tyr Glu Glu Arg
385 390 395 400
Lys Glu Lys Trp Leu Lys Gln Asp Phe Asn Val Ser Leu Ile Gln Thr
405 410 415
Ala Ile Asp Glu Tyr Asp Asn Glu Thr Val Lys Gly Lys Asn Ser Gly
420 425 430
Lys Val Ile Ala Asp Tyr Phe Ala Lys Phe Cys Asp Asp Lys Glu Thr
435 440 445
Asp Leu Ile Gln Lys Val Asn Glu Gly Tyr Ile Ala Val Lys Asp Leu
450 455 460
Leu Asn Thr Pro Cys Pro Glu Asn Glu Lys Leu Gly Ser Asn Lys Asp
465 470 475 480
Gln Val Lys Gln Ile Lys Ala Phe Met Asp Ser Ile Met Asp Ile Met
485 490 495
His Phe Val Arg Pro Leu Ser Leu Lys Asp Thr Asp Lys Glu Lys Asp
500 505 510
Glu Thr Phe Tyr Ser Leu Phe Thr Pro Leu Tyr Asp His Leu Thr Gln
515 520 525
Thr Ile Ala Leu Tyr Asn Lys Val Arg Asn Tyr Leu Thr Gln Lys Pro
530 535 540
Tyr Ser Thr Glu Lys Ile Lys Leu Asn Phe Glu Asn Ser Thr Leu Leu
545 550 555 560
Gly Gly Trp Asp Leu Asn Lys Glu Thr Asp Asn Thr Ala Ile Ile Leu
565 570 575
Arg Lys Asp Asn Leu Tyr Tyr Leu Gly Ile Met Asp Lys Arg His Asn
580 585 590
Arg Ile Phe Arg Asn Val Pro Lys Ala Asp Lys Lys Asp Phe Cys Tyr
595 600 605
Glu Lys Met Val Tyr Lys Leu Leu Pro Gly Ala Asn Lys Met Leu Pro
610 615 620
Lys Val Phe Phe Ser Gln Ser Arg Ile Gln Glu Phe Thr Pro Ser Ala
625 630 635 640
Lys Leu Leu Glu Asn Tyr Ala Asn Glu Thr His Lys Lys Gly Asp Asn
645 650 655
Phe Asn Leu Asn His Cys His Lys Leu Ile Asp Phe Phe Lys Asp Ser
660 665 670
Ile Asn Lys His Glu Asp Trp Lys Asn Phe Asp Phe Arg Phe Ser Ala
675 680 685
Thr Ser Thr Tyr Ala Asp Leu Ser Gly Phe Tyr His Glu Val Glu His
690 695 700
Gln Gly Tyr Lys Ile Ser Phe Gln Ser Val Ala Asp Ser Phe Ile Asp
705 710 715 720
Asp Leu Val Asn Glu Gly Lys Leu Tyr Leu Phe Gln Ile Tyr Asn Lys
725 730 735
Asp Phe Ser Pro Phe Ser Lys Gly Lys Pro Asn Leu His Thr Leu Tyr
740 745 750
Trp Lys Met Leu Phe Asp Glu Asn Asn Leu Lys Asp Val Val Tyr Lys
755 760 765
Leu Asn Gly Glu Ala Glu Val Phe Tyr Arg Lys Lys Ser Ile Ala Glu
770 775 780
Lys Asn Thr Thr Ile His Lys Ala Asn Glu Ser Ile Ile Asn Lys Asn
785 790 795 800
Pro Asp Asn Pro Lys Ala Thr Ser Thr Phe Asn Tyr Asp Ile Val Lys
805 810 815
Asp Lys Arg Tyr Thr Ile Asp Lys Phe Gln Phe His Ile Pro Ile Thr
820 825 830
Met Asn Phe Lys Ala Glu Gly Ile Phe Asn Met Asn Gln Arg Val Asn
835 840 845
Gln Phe Leu Lys Ala Asn Pro Asp Ile Asn Ile Ile Gly Ile Asp Arg
850 855 860
Gly Glu Arg His Leu Leu Tyr Tyr Ala Leu Ile Asn Gln Lys Gly Lys
865 870 875 880
Ile Leu Lys Gln Asp Thr Leu Asn Val Ile Ala Asn Glu Lys Gln Lys
885 890 895
Val Asp Tyr His Asn Leu Leu Asp Lys Lys Glu Gly Asp Arg Ala Thr
900 905 910
Ala Arg Gln Glu Trp Gly Val Ile Glu Thr Ile Lys Glu Leu Lys Glu
915 920 925
Gly Tyr Leu Ser Gln Val Ile His Lys Leu Thr Asp Leu Met Ile Glu
930 935 940
Asn Asn Ala Ile Ile Val Met Glu Asp Leu Asn Phe Gly Phe Lys Arg
945 950 955 960
Gly Arg Gln Lys Val Glu Lys Gln Val Tyr Gln Lys Phe Glu Lys Met
965 970 975
Leu Ile Asp Lys Leu Asn Tyr Leu Val Asp Lys Asn Lys Lys Ala Asn
980 985 990
Glu Leu Gly Gly Leu Leu Asn Ala Phe Gln Leu Ala Asn Lys Phe Glu
995 1000 1005
Ser Phe Gln Lys Met Gly Lys Gln Asn Gly Phe Ile Phe Tyr Val
1010 1015 1020
Pro Ala Trp Asn Thr Ser Lys Thr Asp Pro Ala Thr Gly Phe Ile
1025 1030 1035
Asp Phe Leu Lys Pro Arg Tyr Glu Asn Leu Asn Gln Ala Lys Asp
1040 1045 1050
Phe Phe Glu Lys Phe Asp Ser Ile Arg Leu Asn Ser Lys Ala Asp
1055 1060 1065
Tyr Phe Glu Phe Ala Phe Asp Phe Lys Asn Phe Thr Glu Lys Ala
1070 1075 1080
Asp Gly Gly Arg Thr Lys Trp Thr Val Cys Thr Thr Asn Glu Asp
1085 1090 1095
Arg Tyr Gln Trp Asn Arg Ala Leu Asn Asn Asn Arg Gly Ser Gln
1100 1105 1110
Glu Lys Tyr Asp Ile Thr Ala Glu Leu Lys Ser Leu Phe Asp Gly
1115 1120 1125
Lys Val Asp Tyr Lys Ser Gly Lys Asp Leu Lys Gln Gln Ile Ala
1130 1135 1140
Ser Gln Glu Ser Ala Asp Phe Phe Lys Ala Leu Met Lys Asn Leu
1145 1150 1155
Ser Ile Thr Leu Ser Leu Arg His Asn Asn Gly Glu Lys Gly Asp
1160 1165 1170
Asn Glu Gln Asp Tyr Ile Leu Ser Pro Val Ala Asp Ser Lys Gly
1175 1180 1185
Arg Phe Phe Asp Ser Arg Lys Ala Asp Asp Asp Met Pro Lys Asn
1190 1195 1200
Ala Asp Ala Asn Gly Ala Tyr His Ile Ala Leu Lys Gly Leu Trp
1205 1210 1215
Cys Leu Glu Gln Ile Ser Lys Thr Asp Asp Leu Lys Lys Val Lys
1220 1225 1230
Leu Ala Ile Ser Asn Lys Glu Trp Leu Glu Phe Val Gln Thr Leu
1235 1240 1245
Lys Gly
1250
<210> 18
<211> 3987
<212> DNA
<213> 人工的
<220>
<223> 合成的Cas12a多核苷酸
<400> 18
atggccggga gcaagaagcg ccggataaag caggacacgc agttcgaggg cttcaccaac 60
ctgtaccaag tctccaagac gctccggttc gagcttatcc cgcaagggaa gaccctgaaa 120
cacatccagg aacaaggttt catcgaggag gacaaggccc gcaacgacca ctacaaggag 180
ctcaagccca taatcgatcg gatctacaag acgtacgccg accagtgcct ccaactggtg 240
cagctcgact gggagaacct gagcgccgcc attgacagct accgcaagga aaagacggag 300
gagacgcgca acgcccttat tgaggagcaa gccacctacc gcaacgccat ccacgactac 360
ttcatcgggc gcaccgacaa cctgacggac gcgatcaaca agcgccacgc ggaaatctac 420
aagggccttt tcaaggccga gctcttcaac gggaaggtcc taaaacagct cgggactgtc 480
acgacaaccg agcatgagaa cgccctcctt cgcagcttcg acaagttcac cacatacttc 540
tcgggcttct accggaaccg caagaacgtt ttcagcgccg aggacatctc caccgccatc 600
ccgcacagga tcgtccagga caacttcccc aagttcaagg agaactgcca catcttcacg 660
cgcctgatta cagccgtacc ttcacttcgt gagcacttcg agaacgtcaa aaaggccatc 720
gggatcttcg tctccacgtc catcgaggag gtattctctt tcccgttcta taaccagctc 780
ctgacccaga cgcagatcga cctctacaac cagctactgg gcggcatcag ccgggaggcc 840
gggaccgaga aaataaaggg cctcaacgaa gttctcaacc tggccatcca gaagaacgac 900
gagaccgcgc atatcatcgc atccctgccg catcgcttca ttcctttgtt caagcagata 960
ttgagcgacc ggaacaccct ctcgttcatc ctcgaagaat tcaagagcga cgaggaggtc 1020
attcagtctt tctgcaagta caagacgctc ctacggaatg agaatgtgct ggagaccgcg 1080
gaggcactct tcaatgagct gaactccatt gacctgaccc acatcttcat tagccacaag 1140
aaactggaga cgatctccag cgccctgtgc gaccactggg acactctccg caacgccctc 1200
tacgaacgcc ggatctccga acttaccggc aagataacta agtcggctaa ggagaaggtg 1260
caacggagcc tcaagcacga ggacatcaac cttcaggaaa tcatctcagc cgcgggcaag 1320
gagctgagcg aggcgtttaa gcagaaaaca tcggagatac tgagccacgc gcacgcggcc 1380
ctggatcaac cgctgccgac gactctcaag aagcaagagg agaaggaaat ccttaagtcc 1440
cagctcgact cgctgctcgg cctctatcac ttgctcgact ggttcgcggt tgatgagtcc 1500
aacgaggtgg acccggagtt ctccgcgcgc ctcacgggta ttaagctgga gatggagcca 1560
agcttaagct tctacaacaa ggcccgcaac tacgcgacca aaaaaccgta ctcagtcgag 1620
aaattcaagc tgaatttcca gatgcctaca ttggcgaggg ggtgggacgt gaaccgcgag 1680
aagaacaatg gagccatcct gttcgtcaaa aatgggttgt actacctggg catcatgccc 1740
aagcagaagg gccgttacaa ggccctgtca ttcgagccta ccgagaagac ctcggagggc 1800
ttcgacaaga tgtactacga ctatttcccg gacgccgcca agatgatccc gaagtgctcc 1860
acgcagctca aagccgtcac ggcccacttc cagacgcata ccacgccgat acttctgagc 1920
aacaacttca ttgagccgct agagatcacg aaggagatat acgacctaaa caaccccgaa 1980
aaggagccca agaagttcca gacagcctac gctaagaaga caggtgatca gaagggatat 2040
agggaggcac tctgcaagtg gatcgacttc acgcgcgact tcctgtcgaa atatacaaag 2100
acgaccagca ttgacctaag ttctctccgc ccatcctccc agtacaagga tctgggcgag 2160
tattatgcgg agctgaaccc attgctgtac cacatcagct tccagaggat cgccgagaag 2220
gagattatgg acgcggtgga gacggggaaa ctatacctgt tccaaatata taacaaggac 2280
ttcgctaaag ggcaccacgg gaagcccaac ctgcacacac tctactggac gggcttgttt 2340
tcgccagaaa atttggccaa gacttcgatc aagctcaacg gccaggcgga gttgttttac 2400
cgtcccaagt ctcgcatgaa gcgcatggcg catcgcctcg gagagaaaat gcttaacaag 2460
aagctcaagg atcagaagac gcccatacct gatacgttgt accaggaatt gtacgactac 2520
gtgaaccacc gcctatcgca cgacctctca gacgaggccc gcgccctcct cccaaacgtg 2580
attactaagg aggtttccca tgaaataatc aaggaccgac ggttcaccag cgacaaattt 2640
tttttccacg tgcctatcac gctcaattac caggcggcca actccccatc gaagttcaac 2700
cagcgcgtga acgcctacct taaggagcac ccggagaccc caatcatcgg gatcgaccgt 2760
ggcgagcgga acctgatcta tattacggtg atcgatagca ccgggaagat cctggagcag 2820
cgctccctga acacaatcca gcagtttgac taccagaaga aactcgacaa ccgggagaag 2880
gagcgcgtcg cagcccggca agcatggagt gtggtcggca ccataaagga cctgaaacag 2940
ggttacctaa gtcaagttat ccacgagatc gttgacctga tgatacacta tcaagccgta 3000
gtcgtgctgg agaacctcaa cttcgggttt aagtccaagc gcaccggcat cgcggagaag 3060
gcggtgtacc agcagttcga gaagatgctg atcgacaagc tgaactgcct ggtgctcaag 3120
gactaccctg cggagaaggt cggcggggtc ttgaacccgt accagctaac cgaccagttc 3180
acgagcttcg ccaaaatggg cacgcagtcc ggattcttgt tttatgtccc ggctccatat 3240
acaagtaaga tcgacccgct gacagggttt gttgacccat tcgtgtggaa gaccatcaag 3300
aaccacgaga gcaggaaaca cttcttagag ggcttcgact tcctgcatta cgacgttaag 3360
acaggcgact tcatcctgca cttcaagatg aaccgcaacc tgtcgttcca gaggggcctg 3420
cccggcttca tgcccgcctg ggatatcgtc tttgagaaga atgagacgca gttcgacgcg 3480
aaggggacgc cgttcatcgc tggaaagcgg atcgtgccgg tcatcgagaa ccaccgcttc 3540
acgggtcgct accgagattt ataccccgcc aacgaactaa ttgcgctgct ggaggagaag 3600
gggatcgtgt tccgagatgg cagcaacatt ctcccgaagc tgctggagaa cgacgactcg 3660
cacgctattg acacgatggt cgccctcata cggagcgtgc ttcagatgcg gaacagtaac 3720
gctgccacgg gcgaggacta cattaactcc cccgtccgcg acctcaacgg ggtctgcttc 3780
gatagccgct tccagaaccc ggagtggcct atggatgcgg acgcgaacgg ggcctaccac 3840
atcgccctca agggccaact cctgctcaac cacttgaagg aaagcaaaga cctcaaattg 3900
cagaatggca tcagtaacca ggactggctc gcgtacatcc aggaactgag aaacgggtcc 3960
aagaagcggc gtatcaagca agattga 3987
<210> 19
<211> 3987
<212> DNA
<213> 人工的
<220>
<223> 合成的Cas12a多核苷酸
<400> 19
atggcgggaa gcaaaaagcg ccggattaag caagacacgc agttcgaggg cttcacgaac 60
ctctaccaag tcagcaagac cctccggttc gagctgatac cacagggaaa gacgctcaag 120
cacatccagg aacagggctt catcgaggag gacaaggcgc gcaacgacca ctacaaggag 180
ttgaaaccga tcatcgaccg catctacaag acgtacgccg accagtgcct ccagctcgtg 240
cagctcgact gggagaacct ctccgccgcc attgactcgt accggaagga gaagactgag 300
gagacccgca acgccctgat cgaggagcaa gcaacctacc ggaacgccat ccacgactac 360
ttcatcggcc gcaccgacaa cctcaccgac gcgatcaaca agcggcacgc ggagatatac 420
aaagggctgt tcaaggcgga gctgttcaac ggcaaggtgc tcaagcagct agggacggtg 480
accacgaccg agcacgagaa cgcgctcctc cgcagcttcg acaagttcac cacctacttc 540
agcggcttct accggaaccg caagaatgtg ttcagcgcgg aggacatcag cacggccatc 600
ccgcaccgca tcgtccagga caacttcccg aagttcaagg agaactgcca catcttcacc 660
cgcctgataa ccgccgtccc ctccctgcgg gagcacttcg agaacgtcaa aaaggcaatt 720
gggatcttcg tctcgaccag cattgaggag gtgttcagct tccccttcta caaccagctc 780
ctcacccaga cgcagatcga cctgtacaat cagttgctcg gcgggataag ccgcgaggcg 840
ggaaccgaaa aaatcaaggg gctgaacgaa gtgttgaacc tcgccatcca gaagaacgac 900
gagaccgcgc acatcatcgc ctccctgccc caccggttca tcccgctgtt caagcagatc 960
ctctctgacc ggaacaccct gtccttcatt cttgaggagt tcaagtcgga cgaggaggtc 1020
atccagagct tctgcaagta caagacgctg ctacggaacg agaacgtgct ggagacggcg 1080
gaggcactgt tcaacgagct aaacagcatc gacctcacgc acatcttcat cagtcacaag 1140
aaactggaga ccatctcctc cgcgctgtgc gaccactggg acacgctcag gaacgcgctc 1200
tacgagcgcc gaatcagtga gctgacgggc aagatcacga agtccgcgaa ggagaaggtg 1260
cagcggtccc tcaagcacga ggacatcaac ctccaggaga tcatctcagc ggctgggaaa 1320
gagctgtccg aggcgttcaa gcagaaaacg agcgaaatcc tgtcccacgc gcacgcggcc 1380
ctggatcagc ctctgccgac gaccctcaag aaacaagaag aaaaggaaat cctcaagtcg 1440
cagctcgact cgctgctggg cctgtaccat ctcctcgact ggttcgccgt ggacgagagc 1500
aacgaggtgg accccgagtt ctccgcgcgg cttacgggga tcaagctgga gatggagccc 1560
agcctgtcct tctacaacaa ggcgcgcaac tacgccacca agaagcccta cagcgtggag 1620
aagttcaagc tcaacttcca gatgcccact ctcgcacgtg ggtgggacgt caaccgcgaa 1680
aaaaataatg gggcgatcct gttcgtcaag aacggcctgt actacttggg catcatgccg 1740
aaacagaagg gccgctacaa ggccctgagc ttcgaaccga ccgagaaaac gagcgagggg 1800
ttcgacaaga tgtactacga ctacttcccc gacgccgcga agatgattcc aaagtgctcc 1860
acgcagctta aggccgtgac ggcccacttc cagacgcaca cgaccccgat cctcctcagc 1920
aacaacttca tcgagcccct ggagatcacg aaggagatat acgacctgaa caacccggag 1980
aaggagccca agaaattcca gaccgcctac gccaagaaga caggcgacca aaagggttac 2040
agggaggccc tctgcaagtg gatcgacttc actagggact tcctgtccaa gtacaccaag 2100
actacctcta tcgacctgtc cagcctccgc ccgtcgtccc agtacaaaga tttgggcgag 2160
tattacgcgg agctgaaccc actgctctac cacatcagct tccagcgcat cgcggagaag 2220
gagatcatgg acgcagtgga gacgggcaag ctatacctat ttcagatata caacaaagac 2280
ttcgctaagg gacaccacgg caagcctaac ctgcacaccc tctactggac ggggctcttc 2340
agcccggaga acctcgccaa gacctcgatc aagctcaacg gccaggccga gctgttctac 2400
cggcccaagt cccgcatgaa gcggatggcc caccggctcg gggagaaaat gctcaacaag 2460
aaattgaagg accaaaaaac gccgataccc gacaccctat accaggagct gtacgactat 2520
gtgaaccacc gcctgagcca cgacctcagc gacgaggcgc gggccctcct gccgaacgtc 2580
atcacaaagg aggtcagcca cgagatcatc aaggaccggc gcttcacctc cgacaagttt 2640
ttctttcacg tgcccatcac gctcaactac caggccgcca actcgccgtc caagttcaac 2700
cagcgcgtga acgcctacct caaggagcac cccgagaccc cgatcatcgg gattgaccga 2760
ggggagcgga acctcatcta catcaccgtc atcgacagca ccgggaagat ccttgaacag 2820
cggtcgctca acaccatcca gcagttcgac taccagaaga aactcgacaa ccgggagaag 2880
gagagagtgg cggcccgcca ggcttggtcc gtcgtcggga cgattaagga cttgaaacaa 2940
ggttacctgt cgcaagtgat ccacgagatc gttgacctga tgatccacta ccaagccgtc 3000
gtggtcctgg agaacctcaa cttcggcttc aagagcaaac gaaccggcat cgcggagaag 3060
gccgtgtacc agcagttcga aaaaatgctg atcgacaagc tgaactgcct cgtgctcaag 3120
gactaccccg ctgagaaggt cggcggggtg ctgaacccgt accagctcac tgaccagttc 3180
accagcttcg caaagatggg cacccagtcc ggcttcctgt tctacgtgcc tgcgccatac 3240
acctcgaaga tcgacccgct caccgggttc gtggacccct tcgtctggaa gaccatcaag 3300
aaccacgaga gccgcaagca cttcctggag ggcttcgact tcctccacta cgacgtcaag 3360
accggggact tcatcctgca cttcaagatg aaccgcaacc tcagtttcca gcgcggcctg 3420
ccggggttca tgcccgcttg ggatatagtc ttcgagaaga atgagacgca gttcgacgcg 3480
aagggcaccc cgttcatcgc cgggaagcgc atcgtgccgg tcatcgagaa ccaccggttc 3540
accgggcgct accgcgacct atacccggcg aacgagttga tcgccctcct ggaggagaag 3600
ggcatcgtgt tccgcgacgg ctccaacatc ctcccgaagc tgctcgaaaa cgacgactcc 3660
cacgccatcg acacgatggt cgcgctgatc cggtcggtgc tccagatgcg gaactccaac 3720
gccgcgacgg gcgaggacta catcaacagt ccggtccgcg atctgaacgg cgtctgcttc 3780
gactcccggt tccagaaccc cgagtggccg atggacgcgg acgcgaacgg cgcataccac 3840
atcgccctaa aagggcaatt gctgctcaac cacctcaagg aatccaaaga cctaaagctc 3900
cagaacggca tctccaacca ggactggctg gcgtacatcc aggaactgcg gaacgggagc 3960
aaaaaacgtc ggatcaagca agattga 3987
<210> 20
<211> 3987
<212> DNA
<213> 人工的
<220>
<223> 合成的Cas12a多核苷酸
<400> 20
atggcgggct ccaagaaacg ccggattaag caagataccc agttcgaggg gttcacgaac 60
ctctaccaag tgagcaagac cctccgattc gaactgattc ctcaggggaa gaccctcaag 120
cacatccagg agcaagggtt catcgaggag gacaaggcgc ggaacgacca ctacaaggaa 180
ctcaaaccca tcatcgaccg catctacaag acctacgccg atcagtgcct ccagctcgtg 240
cagttggact gggagaacct cagcgcggcc attgactcct accggaagga gaaaacggag 300
gagacgcgca acgcgctcat cgaggaacag gcaacctatc gcaacgccat ccacgactac 360
ttcatcggga ggactgacaa cctcactgac gcgattaaca agcgccacgc ggagatatac 420
aagggactct tcaaagcgga gctgtttaac ggcaaggttc tcaagcaact cggcactgtg 480
accacgaccg agcatgagaa cgccctgctc cgctccttcg acaagttcac cacctacttc 540
tccgggttct accgcaaccg caagaatgtc ttcagcgcgg aggacatcag cacggccatt 600
ccacatcgaa tcgtccaaga taacttcccg aagttcaagg agaactgcca catcttcacc 660
cgactcatta ctgctgtacc gtcgttacgc gaacacttcg agaacgtcaa gaaggcaatt 720
ggaatcttcg tctctacgtc aatagaggag gtgttcagct tccctttcta caaccagctc 780
cttacgcaga cccagataga cctgtacaat cagctcctcg gtgggatcag ccgggaggcg 840
gggactgaga agattaaagg gctcaacgag gtcttgaacc tggccatcca aaaaaacgat 900
gagacggcgc acatcatcgc ctcgctgccc caccggttca tcccgctgtt caagcagatc 960
ctcagtgaca ggaacacctt gagctttatc ctagaggagt tcaagagcga cgaggaggtg 1020
atccagagct tctgcaagta caaaaccctg ctgaggaacg agaacgtcct ggagacggcg 1080
gaggcgctgt tcaacgagct gaactctatc gacttaactc acatattcat ctcgcacaag 1140
aagctggaga ctattagctc tgcactctgc gaccactggg acaccctccg caacgcgctc 1200
tacgagcgcc gcatctcgga gctgaccggg aagatcacca aatccgcgaa ggaaaaggtc 1260
cagcgttccc tcaaacacga ggatattaac ttacaggaga ttatctcagc ggctgggaag 1320
gagttgtcag aggcgttcaa gcagaaaact tccgagatcc tgagccacgc gcacgcagcg 1380
ctcgaccagc ctctgcccac caccctcaaa aagcaggaag aaaaagagat cctcaagagc 1440
cagttggact ccctgctggg gctctatcac cttctcgact ggttcgccgt cgatgagtcg 1500
aacgaggtgg accccgagtt ctccgcccgg ctgaccggca tcaagctaga gatggagccg 1560
tccctcagct tctacaataa ggcccgcaac tacgcgacca aaaaacccta cagcgtggag 1620
aagttcaagc tgaacttcca gatgccgacc ttagcacgcg gttgggacgt aaacagggag 1680
aagaacaatg gagccatcct gttcgtcaag aacgggcttt actacctcgg gataatgccc 1740
aagcagaagg gccgctacaa ggccctttcc ttcgagccga cggagaaaac ctccgagggg 1800
ttcgacaaga tgtactacga ctacttcccc gacgccgcca agatgatccc gaagtgctca 1860
acgcagctaa aagccgtgac cgcccacttc cagacccaca cgacgccgat cctgctgagc 1920
aacaacttca tcgagcccct tgagatcact aaggagatat acgacctgaa caaccccgag 1980
aaggagccca agaagtttca aaccgcctac gccaaaaaaa ctggcgacca aaagggctac 2040
agggaggcgc tgtgtaagtg gatcgacttc acacgcgact tcctttcgaa gtatacgaag 2100
acaacctcta ttgacctgag cagcctgcgt cctagctccc agtacaaaga tttgggcgag 2160
tactacgcgg agcttaatcc actactctac cacatctcat tccagcgcat cgctgagaag 2220
gaaatcatgg acgcggtgga gacaggcaaa ctgtacctct tccagatata caacaaagac 2280
ttcgctaagg ggcaccacgg gaagcccaac cttcatacgc tctactggac gggcctattc 2340
agccccgaaa atctggccaa gacctccatc aagctgaacg gccaagcgga gctgttctac 2400
agacccaaga gccggatgaa gcggatggcc cacaggctcg gcgagaaaat gcttaacaaa 2460
aagttgaagg accagaaaac ccctatcccc gacaccctct accaggaact gtacgactac 2520
gtgaaccaca ggctctcgca cgacctttcc gacgaggccc gtgccctact cccgaacgtc 2580
attaccaaag aggtttcgca cgagatcatc aaggaccggc ggttcacgag cgacaagttt 2640
ttctttcacg tccccatcac ccttaactac caggcggcca actccccatc caagttcaac 2700
cagcgtgtga atgcctacct caaggagcac ccagagaccc cgatcattgg gatcgaccgg 2760
ggcgagcgga acctgatcta catcaccgtc atcgactcga cgggcaagat tcttgagcag 2820
agatcgttga ataccataca gcagttcgac taccagaaga aactcgacaa ccgcgagaag 2880
gagcgcgtgg cggcccgcca ggcgtggtcc gtcgttggga cgattaagga cttgaaacaa 2940
ggttatctgt cccaagtcat ccacgagatc gttgatctga tgatccacta tcaggcagtg 3000
gtggtgctgg agaatctcaa cttcggcttc aagagtaagc ggacgggaat cgccgagaag 3060
gccgtgtacc agcagttcga gaagatgctg atcgacaagc tcaactgcct tgtgctgaaa 3120
gactacccgg ccgagaaggt cggcggcgtc ctcaacccgt accaacttac cgaccagttc 3180
acctccttcg ccaagatggg cactcagtcc gggttcttgt tctacgtccc cgcaccttac 3240
acctctaaga tcgaccctct gactggcttc gtagatccat tcgtgtggaa gaccattaag 3300
aaccacgaga gccgcaagca cttcctggag ggcttcgact tcctgcacta cgacgtgaag 3360
accggggact tcatccttca cttcaagatg aaccggaacc tcagcttcca gcggggcctg 3420
ccggggttca tgcccgcctg ggacatcgtg ttcgagaaga acgagaccca gttcgacgcg 3480
aagggcacgc ccttcatcgc cgggaagcgt atcgtgccgg tgatcgagaa ccatcgtttc 3540
acgggtcgct accgtgacct ctacccggcg aacgagctta tcgcactcct ggaggagaag 3600
ggcatcgtct tccgggacgg ctccaacatc ctcccgaaac tgctggaaaa cgacgactct 3660
cacgccatcg acacgatggt ggccctcatc cggtccgtgc tccaaatgcg gaacagcaac 3720
gccgccaccg gtgaggacta catcaacagc ccggtccggg atctgaacgg ggtgtgcttc 3780
gattcgcggt tccagaatcc tgagtggccg atggacgcgg atgcaaacgg ggcgtaccac 3840
atcgcgctca agggccagtt acttctgaac caccttaagg agtctaaaga tttgaaactc 3900
cagaacggga tctcgaacca ggactggctg gcctacatcc aagagttgcg gaacggcagc 3960
aagaagcggc ggattaagca agattag 3987
<210> 21
<211> 1592
<212> DNA
<213> 蒺藜苜蓿（Medicago truncatula）
<400> 21
actgttaata atttttaaac gtcagcgcac taaaaaaacg aaaagacgga cacgtgaaaa 60
taaaaaacac acactagttt atgacgcaat actattttac ttatgatttg ggtacattag 120
acaaaaccgt gaaagagatg tatcagctat gaaacctgta tacttcaata cagagactta 180
ctcatatcgg atacgtacgc acgaagtatc atattaatta ttttaatttt taataaatat 240
tttatcggat acttatgtga tactctacat atacacaagg atatttctaa gatactttat 300
agatacgtat cctagaaaaa catgaagagt aaaaaagtga gacaatgttg taaaaattca 360
ttataaatgt atatgattca attttagata tgcatcagta taattgattc tcgatgaaac 420
acttaaaatt atatttcttg tggaagaacg tagcgagaga ggtgattcag ttagacaaca 480
ttaaataaaa ttaatgttaa gttcttttaa tgatgtttct ctcaatatca catcatatga 540
aaatgtaata tgatttataa gaaaattttt aaaaaattta ttttaataat cacatgtact 600
attttttaaa aattgtatct tttataataa tacaataata aagagtaatc agtgttaatt 660
tttcttcaaa tataagtttt attataaatc attgttaacg tatcataagt cattaccgta 720
tcgtatctta attttttttt aaaaaccgct aattcacgta cccgtattgt attgtacccg 780
cacctgtatc acaatcgatc ttagttagaa gaattgtctc gaggcggtgc aagacagcat 840
ataatagacg tggactctct tataccaaac gttgtcgtat cacaaagggt taggtaacaa 900
gtcacagttt gtccacgtgt cacgttttaa ttggaagagg tgccgttggc gtaatataac 960
agccaatcga tttttgctat aaaagcaaat caggtaaact aaacttcttc attcttttct 1020
tccccatcgc tacaaaaccg gttcctttgg aaaagagatt cattcaaacc tagcacccaa 1080
ttccgtttca aggtataatc tactttctat tcttcgatta ttttattatt attagctact 1140
atcgtttaat cgatcttttc ttttgatccg tcaaatttaa attcaattag ggttttgttc 1200
ttttctttca tctgattgaa atccttctga attgaaccgt ttacttgatt ttactgttta 1260
ttgtatgatt taatcctttg tttttcaaag acagtcttta gattgtgatt aggggttcat 1320
ataaattttt agatttggat ttttgtattg tatgattcaa aaaatacgtc ctttaattag 1380
attagtacat ggatattttt tacccgattt attgattgtc agggagaatt tgatgagcaa 1440
gtttttttga tgtctgttgt aaattgaatt gattataatt gctgatctgc tgcttccagt 1500
tttcataacc catattcttt taaccttgtt gtacacacaa tgaaaaattg gtgattgatt 1560
catttgtttt tctttgtttt ggattataca gg 1592
<210> 22
<211> 2000
<212> DNA
<213> 玉蜀黍（Zea mays）
<400> 22
gtcgtgcccc tctctagaga taaagagcat tgcatgtcta aagtataaaa aattaccaca 60
tatttttttg tcacacttat ttgaagtgta gtttatctat ctctatacat atatttaaac 120
ttcactctac aaataatata gtctataata ctaaaataat attagtgttt tagaggatca 180
tataaataaa ctgctagaca tggtctaaag gataattgaa tattttgaca atctacagtt 240
ttatcttttt agtgtgcatg tgatctctct gttttttttg caaatagctt gacctatata 300
atacttcatc cattttatta gtacatccat ttaggattta gggttgatgg tttctataga 360
ctaattttta gtacatccat tttattcttt ttagtctcta aattttttaa aactaaaact 420
ctattttagt tttttattta ataatttaga tataaaatga aataaaataa attgactaca 480
aataaaacaa atacccttta agaaataaaa aaactaagca aacatttttc ttgtttcgag 540
tagataatga caggctgttc aacgccgtcg acgagtctaa cggacaccaa ccagcgaacc 600
agcagcgtcg cgtcgggcca agcgaagcag acggcacggc atctctgtag ctgcctctgg 660
acccctctcg agagttccgc tccaccgttg gacttgctcc gctgtcggca tccagaaatt 720
gcgtggcgga gcggcagacg tgaggcggca cggcaggcgg cctcttcctc ctctcacggc 780
accggcagct acgggggatt cctttcccac cgctccttcg ctttcccttc ctcgcccgcc 840
gtaataaata gacaccccct ccacaccctc tttccccaac ctcgtgttcg ttcggagcgc 900
acacacacgc aaccagatct cccccaaatc cagccgtcgg cacctccgct tcaaggtacg 960
ccgctcatcc tccccccccc cctctctcta ccttctctag atcggcgatc cggtccatgg 1020
ttagggcccg gtagttctac ttctgttcat gtttgtgtta gagcaaacat gttcatgttc 1080
atgtttgtga tgatgtggtc tggttgggcg gtcgttctag atcggagtag gatactgttt 1140
caagctacct ggtggattta ttaattttgt atctgtatgt gtgtgccata catcttcata 1200
gttacgagtt taagatgatg gatggaaata tcgatctagg ataggtatac atgttgatgc 1260
gggttttact gatgcatata cagagatgct ttttttctcg cttggttgtg atgatatggt 1320
ctggttgggc ggtcgttcta gatcggagta gaatactgtt tcaaactacc tggtggattt 1380
attaaaggat aaagggtcgt tctagatcgg agtagaatac tgtttcaaac tacctggtgg 1440
atttattaaa ggatctgtat gtatgtgcct acatcttcat agttacgagt ttaagatgat 1500
ggatggaaat atcgatctag gataggtata catgttgatg cgggttttac tgatgcatat 1560
acagagatgc tttttttcgc ttggttgtga tgatgtggtc tggttgggcg gtcgttctag 1620
atcggagtag aatactgttt caaactacct ggtggattta ttaattttgt atctttatgt 1680
gtgtgccata catcttcata gttacgagtt taagatgatg gatggaaata ttgatctagg 1740
ataggtatac atgttgatgt gggttttact gatgcatata catgatggca tatgcggcat 1800
ctattcatat gctctaacct tgagtaccta tctattataa taaacaagta tgttttataa 1860
ttattttgat cttgatatac ttggatgatg gcatatgcag cagctatatg tggatttttt 1920
agccctgcct tcatacgcta tttatttgct tggtactgtt tcttttgtcc gatgctcacc 1980
ctgttgtttg gtgatacttc 2000
<210> 23
<211> 24
<212> PRT
<213> 人工的
<220>
<223> 合成的GCN4序列
<400> 23
Glu Glu Leu Leu Ser Lys Asn Tyr His Leu Glu Asn Glu Val Ala Arg
1 5 10 15
Leu Lys Lys Gly Ser Gly Ser Gly
20
<210> 24
<211> 241
<212> PRT
<213> 人工的
<220>
<223> 合成的GCN4序列
<400> 24
Glu Glu Glu Leu Leu Ser Lys Asn Tyr His Leu Glu Asn Glu Val Ala
1 5 10 15
Arg Leu Lys Lys Gly Ser Gly Ser Gly Glu Glu Leu Leu Ser Lys Asn
20 25 30
Tyr His Leu Glu Asn Glu Val Ala Arg Leu Lys Lys Gly Ser Gly Ser
35 40 45
Gly Glu Glu Leu Leu Ser Lys Asn Tyr His Leu Glu Asn Glu Val Ala
50 55 60
Arg Leu Lys Lys Gly Ser Gly Ser Gly Glu Glu Leu Leu Ser Lys Asn
65 70 75 80
Tyr His Leu Glu Asn Glu Val Ala Arg Leu Lys Lys Gly Ser Gly Ser
85 90 95
Gly Glu Glu Leu Leu Ser Lys Asn Tyr His Leu Glu Asn Glu Val Ala
100 105 110
Arg Leu Lys Lys Gly Ser Gly Ser Gly Glu Glu Leu Leu Ser Lys Asn
115 120 125
Tyr His Leu Glu Asn Glu Val Ala Arg Leu Lys Lys Gly Ser Gly Ser
130 135 140
Gly Glu Glu Leu Leu Ser Lys Asn Tyr His Leu Glu Asn Glu Val Ala
145 150 155 160
Arg Leu Lys Lys Gly Ser Gly Ser Gly Glu Glu Leu Leu Ser Lys Asn
165 170 175
Tyr His Leu Glu Asn Glu Val Ala Arg Leu Lys Lys Gly Ser Gly Ser
180 185 190
Gly Glu Glu Leu Leu Ser Lys Asn Tyr His Leu Glu Asn Glu Val Ala
195 200 205
Arg Leu Lys Lys Gly Ser Gly Ser Gly Glu Glu Leu Leu Ser Lys Asn
210 215 220
Tyr His Leu Glu Asn Glu Val Ala Arg Leu Lys Lys Gly Ser Gly Ser
225 230 235 240
Gly
<210> 25
<211> 277
<212> PRT
<213> 人工的
<220>
<223> 合成的ScFv抗体
<400> 25
Met Gly Pro Asp Ile Val Met Thr Gln Ser Pro Ser Ser Leu Ser Ala
1 5 10 15
Ser Val Gly Asp Arg Val Thr Ile Thr Cys Arg Ser Ser Thr Gly Ala
20 25 30
Val Thr Thr Ser Asn Tyr Ala Ser Trp Val Gln Glu Lys Pro Gly Lys
35 40 45
Leu Phe Lys Gly Leu Ile Gly Gly Thr Asn Asn Arg Ala Pro Gly Val
50 55 60
Pro Ser Arg Phe Ser Gly Ser Leu Ile Gly Asp Lys Ala Thr Leu Thr
65 70 75 80
Ile Ser Ser Leu Gln Pro Glu Asp Phe Ala Thr Tyr Phe Cys Ala Leu
85 90 95
Trp Tyr Ser Asn His Trp Val Phe Gly Gln Gly Thr Lys Val Glu Leu
100 105 110
Lys Arg Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly
115 120 125
Ser Ser Gly Gly Gly Ser Glu Val Lys Leu Leu Glu Ser Gly Gly Gly
130 135 140
Leu Val Gln Pro Gly Gly Ser Leu Lys Leu Ser Cys Ala Val Ser Gly
145 150 155 160
Phe Ser Leu Thr Asp Tyr Gly Val Asn Trp Val Arg Gln Ala Pro Gly
165 170 175
Arg Gly Leu Glu Trp Ile Gly Val Ile Trp Gly Asp Gly Ile Thr Asp
180 185 190
Tyr Asn Ser Ala Leu Lys Asp Arg Phe Ile Ile Ser Lys Asp Asn Gly
195 200 205
Lys Asn Thr Val Tyr Leu Gln Met Ser Lys Val Arg Ser Asp Asp Thr
210 215 220
Ala Leu Tyr Tyr Cys Val Thr Gly Leu Phe Asp Tyr Trp Gly Gln Gly
225 230 235 240
Thr Leu Val Thr Val Ser Ser Tyr Pro Tyr Asp Val Pro Asp Tyr Ala
245 250 255
Gly Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser
260 265 270
Gly Gly Gly Gly Ser
275
<210> 26
<211> 66
<212> DNA
<213> 贝糖酵母（Saccharomyces bayanus）
<400> 26
ttcttgtcgt acttatagat cgctacgtta tttcaatttt gaaaatctga gtcctgggag 60
tgcgga 66
<210> 27
<211> 609
<212> PRT
<213> 智人（Homo sapiens）
<400> 27
Met Ser Gly Trp Glu Ser Tyr Tyr Lys Thr Glu Gly Asp Glu Glu Ala
1 5 10 15
Glu Glu Glu Gln Glu Glu Asn Leu Glu Ala Ser Gly Asp Tyr Lys Tyr
20 25 30
Ser Gly Arg Asp Ser Leu Ile Phe Leu Val Asp Ala Ser Lys Ala Met
35 40 45
Phe Glu Ser Gln Ser Glu Asp Glu Leu Thr Pro Phe Asp Met Ser Ile
50 55 60
Gln Cys Ile Gln Ser Val Tyr Ile Ser Lys Ile Ile Ser Ser Asp Arg
65 70 75 80
Asp Leu Leu Ala Val Val Phe Tyr Gly Thr Glu Lys Asp Lys Asn Ser
85 90 95
Val Asn Phe Lys Asn Ile Tyr Val Leu Gln Glu Leu Asp Asn Pro Gly
100 105 110
Ala Lys Arg Ile Leu Glu Leu Asp Gln Phe Lys Gly Gln Gln Gly Gln
115 120 125
Lys Arg Phe Gln Asp Met Met Gly His Gly Ser Asp Tyr Ser Leu Ser
130 135 140
Glu Val Leu Trp Val Cys Ala Asn Leu Phe Ser Asp Val Gln Phe Lys
145 150 155 160
Met Ser His Lys Arg Ile Met Leu Phe Thr Asn Glu Asp Asn Pro His
165 170 175
Gly Asn Asp Ser Thr Lys Ala Ser Arg Ala Arg Thr Lys Ala Gly Asp
180 185 190
Leu Arg Asp Thr Gly Ile Phe Leu Asp Leu Met His Leu Lys Lys Pro
195 200 205
Gly Gly Phe Asp Ile Ser Leu Phe Tyr Arg Asp Ile Ile Ser Ile Ala
210 215 220
Glu Asp Glu Asp Leu Arg Val His Phe Glu Glu Ser Ser Lys Leu Glu
225 230 235 240
Asp Leu Leu Arg Lys Val Arg Ala Lys Glu Thr Arg Lys Arg Ala Leu
245 250 255
Ser Arg Leu Lys Leu Lys Leu Asn Lys Asp Ile Val Ile Ser Val Gly
260 265 270
Ile Tyr Asn Leu Val Gln Lys Ala Leu Lys Pro Pro Pro Ile Lys Leu
275 280 285
Tyr Arg Glu Thr Asn Glu Pro Val Lys Thr Lys Thr Arg Thr Phe Asn
290 295 300
Thr Ser Thr Gly Gly Leu Leu Leu Pro Ser Asp Thr Lys Arg Ser Gln
305 310 315 320
Ile Tyr Gly Ser Arg Gln Ile Ile Leu Glu Lys Glu Glu Thr Glu Glu
325 330 335
Leu Lys Arg Phe Asp Asp Pro Gly Leu Met Leu Met Gly Phe Lys Pro
340 345 350
Leu Val Leu Leu Lys Lys His His Tyr Leu Arg Pro Ser Leu Phe Val
355 360 365
Tyr Pro Glu Glu Ser Leu Val Ile Gly Ser Ser Thr Leu Phe Ser Ala
370 375 380
Leu Leu Ile Lys Cys Leu Glu Lys Glu Val Ala Ala Leu Cys Arg Tyr
385 390 395 400
Thr Pro Arg Arg Asn Ile Pro Pro Tyr Phe Val Ala Leu Val Pro Gln
405 410 415
Glu Glu Glu Leu Asp Asp Gln Lys Ile Gln Val Thr Pro Pro Gly Phe
420 425 430
Gln Leu Val Phe Leu Pro Phe Ala Asp Asp Lys Arg Lys Met Pro Phe
435 440 445
Thr Glu Lys Ile Met Ala Thr Pro Glu Gln Val Gly Lys Met Lys Ala
450 455 460
Ile Val Glu Lys Leu Arg Phe Thr Tyr Arg Ser Asp Ser Phe Glu Asn
465 470 475 480
Pro Val Leu Gln Gln His Phe Arg Asn Leu Glu Ala Leu Ala Leu Asp
485 490 495
Leu Met Glu Pro Glu Gln Ala Val Asp Leu Thr Leu Pro Lys Val Glu
500 505 510
Ala Met Asn Lys Arg Leu Gly Ser Leu Val Asp Glu Phe Lys Glu Leu
515 520 525
Val Tyr Pro Pro Asp Tyr Asn Pro Glu Gly Lys Val Thr Lys Arg Lys
530 535 540
His Asp Asn Glu Gly Ser Gly Ser Lys Arg Pro Lys Val Glu Tyr Ser
545 550 555 560
Glu Glu Glu Leu Lys Thr His Ile Ser Lys Gly Thr Leu Gly Lys Phe
565 570 575
Thr Val Pro Met Leu Lys Glu Ala Cys Arg Ala Tyr Gly Leu Lys Ser
580 585 590
Gly Leu Lys Lys Gln Glu Leu Leu Glu Ala Leu Thr Lys His Phe Gln
595 600 605
Asp
<210> 28
<211> 482
<212> PRT
<213> 人工的
<220>
<223> 合成的多肽
<400> 28
Met Val Arg Ser Gly Asn Lys Ala Ala Trp Leu Cys Met Asp Val Gly
1 5 10 15
Phe Thr Met Ser Asn Ser Ile Pro Gly Ile Glu Ser Pro Phe Glu Gln
20 25 30
Ala Lys Lys Val Ile Thr Met Phe Val Gln Arg Gln Val Phe Ala Glu
35 40 45
Asn Lys Asp Glu Ile Ala Leu Val Leu Phe Gly Thr Asp Gly Thr Asp
50 55 60
Asn Pro Leu Ser Gly Gly Asp Gln Tyr Gln Asn Ile Thr Val His Arg
65 70 75 80
His Leu Met Leu Pro Asp Phe Asp Leu Leu Glu Asp Ile Glu Ser Lys
85 90 95
Ile Gln Pro Gly Ser Gln Gln Ala Asp Phe Leu Asp Ala Leu Ile Val
100 105 110
Ser Met Asp Val Ile Gln His Glu Thr Ile Gly Lys Lys Phe Glu Lys
115 120 125
Arg His Ile Glu Ile Phe Thr Asp Leu Ser Ser Arg Phe Ser Lys Ser
130 135 140
Gln Leu Asp Ile Ile Ile His Ser Leu Lys Lys Cys Asp Ile Ser Glu
145 150 155 160
Arg His Ser Ile His Trp Pro Cys Arg Leu Thr Ile Gly Ser Asn Leu
165 170 175
Ser Ile Arg Ile Ala Ala Tyr Lys Ser Ile Leu Gln Glu Arg Val Lys
180 185 190
Lys Thr Thr Trp Asp Ala Lys Thr Leu Lys Lys Glu Asp Ile Gln Lys
195 200 205
Glu Thr Val Tyr Cys Leu Asn Asp Asp Asp Glu Thr Glu Val Leu Lys
210 215 220
Glu Asp Ile Ile Gln Gly Phe Arg Tyr Gly Ser Asp Ile Val Pro Phe
225 230 235 240
Ser Lys Val Asp Glu Glu Gln Met Lys Tyr Lys Ser Glu Gly Lys Cys
245 250 255
Phe Ser Val Leu Gly Phe Cys Lys Ser Ser Gln Val Gln Arg Arg Phe
260 265 270
Phe Met Gly Asn Gln Val Leu Lys Val Phe Ala Ala Arg Asp Asp Glu
275 280 285
Ala Ala Ala Val Ala Leu Ser Ser Leu Ile His Ala Leu Asp Asp Leu
290 295 300
Asp Ile Trp Ala Ile Val Arg Tyr Ala Tyr Asp Lys Arg Ala Asn Pro
305 310 315 320
Gln Val Gly Val Ala Phe Pro His Ile Lys His Asn Tyr Glu Cys Leu
325 330 335
Val Tyr Val Gln Leu Pro Phe Met Glu Asp Leu Arg Gln Tyr Met Phe
340 345 350
Ser Ser Leu Lys Asn Ser Lys Lys Tyr Ala Pro Thr Glu Ala Gln Leu
355 360 365
Asn Ala Val Asp Ala Leu Ile Asp Ser Met Ser Leu Ala Lys Lys Asp
370 375 380
Glu Lys Thr Asp Thr Leu Glu Asp Leu Phe Pro Thr Thr Lys Ile Pro
385 390 395 400
Asn Pro Arg Phe Gln Arg Leu Phe Gln Cys Leu Leu His Arg Ala Leu
405 410 415
His Pro Arg Glu Pro Leu Pro Pro Ile Gln Gln His Ile Trp Asn Met
420 425 430
Leu Asn Pro Pro Ala Glu Val Thr Thr Lys Ser Gln Ile Pro Leu Ser
435 440 445
Lys Ile Lys Thr Leu Phe Pro Leu Ile Glu Ala Lys Lys Lys Asp Gln
450 455 460
Val Thr Ala Gln Glu Ile Phe Gln Asp Asn His Glu Asp Gly Pro Thr
465 470 475 480
Ala Lys
<210> 29
<211> 10
<212> DNA
<213> 热自养甲烷杆菌（Methanobacterium thermoautotrophicum）
<400> 29
aatttttgga 10
<210> 30
<211> 83
<212> PRT
<213> 热自养甲烷杆菌
<400> 30
Gly Ser Val Ile Asp Val Ser Ser Gln Arg Val Asn Val Gln Arg Pro
1 5 10 15
Leu Asp Ala Leu Gly Asn Ser Leu Asn Ser Pro Val Ile Ile Lys Leu
20 25 30
Lys Gly Asp Arg Glu Phe Arg Gly Val Leu Lys Ser Phe Asp Leu His
35 40 45
Met Asn Leu Val Leu Asn Asp Ala Glu Glu Leu Glu Asp Gly Glu Val
50 55 60
Thr Arg Arg Leu Gly Thr Val Leu Ile Arg Gly Asp Asn Ile Val Tyr
65 70 75 80
Ile Ser Pro
<210> 31
<211> 25
<212> DNA
<213> 埃希氏菌（Escherichia）病毒MS2
<400> 31
gcgcacatga ggatcaccca tgtgc 25
<210> 32
<211> 116
<212> PRT
<213> 埃希氏菌病毒MS2
<400> 32
Met Ala Ser Asn Phe Thr Gln Phe Val Leu Val Asp Asn Gly Gly Thr
1 5 10 15
Gly Asp Val Thr Val Ala Pro Ser Asn Phe Ala Asn Gly Ile Ala Glu
20 25 30
Ile Ser Ser Asn Ser Arg Ser Gln Ala Tyr Lys Val Thr Cys Ser Val
35 40 45
Arg Gln Ser Ser Ala Gln Asn Arg Lys Tyr Thr Ile Lys Val Glu Val
50 55 60
Pro Lys Gly Ala Trp Arg Ser Tyr Leu Asn Met Glu Leu Thr Ile Pro
65 70 75 80
Ile Phe Ala Thr Asn Ser Asp Cys Glu Leu Ile Val Lys Ala Met Gln
85 90 95
Gly Leu Leu Lys Asp Gly Asn Pro Ile Pro Ser Ala Ile Ala Ala Asn
100 105 110
Ser Gly Ile Tyr
115
<210> 33
<211> 26
<212> DNA
<213> 噬菌体PP7
<400> 33
ataaggagtt tatatggaaa ccctta 26
<210> 34
<211> 127
<212> PRT
<213> 噬菌体PP7
<400> 34
Met Ser Lys Thr Ile Val Leu Ser Val Gly Glu Ala Thr Arg Thr Leu
1 5 10 15
Thr Glu Ile Gln Ser Thr Ala Asp Arg Gln Ile Phe Glu Glu Lys Val
20 25 30
Gly Pro Leu Val Gly Arg Leu Arg Leu Thr Ala Ser Leu Arg Gln Asn
35 40 45
Gly Ala Lys Thr Ala Tyr Arg Val Asn Leu Lys Leu Asp Gln Ala Asp
50 55 60
Trp Asp Cys Ser Thr Ser Val Cys Gly Glu Leu Pro Lys Val Arg Tyr
65 70 75 80
Thr Gln Val Trp Ser His Asp Val Thr Ile Val Ala Asn Ser Thr Glu
85 90 95
Ala Ser Arg Lys Ser Leu Tyr Asp Leu Thr Lys Ser Leu Val Ala Thr
100 105 110
Ser Gln Val Glu Asp Leu Val Val Asn Leu Val Pro Leu Gly Arg
115 120 125
<210> 35
<211> 19
<212> DNA
<213> Unknown
<220>
<223> 志贺氏菌（Shigella）病毒
<400> 35
ctgaatgcct gcgagcatc 19
<210> 36
<211> 62
<212> PRT
<213> Unknown
<220>
<223> 志贺氏菌病毒
<400> 36
Met Lys Ser Ile Arg Cys Lys Asn Cys Asn Lys Leu Leu Phe Lys Ala
1 5 10 15
Asp Ser Phe Asp His Ile Glu Ile Arg Cys Pro Arg Cys Lys Arg His
20 25 30
Ile Ile Met Leu Asn Ala Cys Glu His Pro Thr Glu Lys His Cys Gly
35 40 45
Lys Arg Glu Lys Ile Thr His Ser Asp Glu Thr Val Arg Tyr
50 55 60
<210> 37
<211> 79
<212> PRT
<213> 智人
<400> 37
Arg Val Leu Tyr Val Gly Gly Leu Ala Glu Glu Val Asp Asp Lys Val
1 5 10 15
Leu His Ala Ala Phe Ile Pro Phe Gly Asp Ile Thr Asp Ile Gln Ile
20 25 30
Pro Leu Asp Tyr Glu Thr Glu Lys His Arg Gly Phe Ala Phe Val Glu
35 40 45
Phe Glu Leu Ala Glu Asp Ala Ala Ala Ala Ile Asp Asn Met Asn Glu
50 55 60
Ser Glu Leu Phe Gly Arg Thr Ile Arg Val Asn Leu Ala Lys Pro
65 70 75
<210> 38
<211> 78
<212> PRT
<213> 智人
<400> 38
Arg Ser Ile Tyr Val Gly Asn Val Asp Tyr Gly Ala Thr Ala Glu Glu
1 5 10 15
Leu Glu Ala His Phe His Gly Cys Gly Ser Val Asn Arg Val Thr Ile
20 25 30
Leu Cys Asp Lys Phe Ser Gly His Pro Lys Gly Phe Ala Tyr Ile Glu
35 40 45
Phe Ser Asp Lys Glu Ser Val Arg Thr Ser Leu Ala Leu Asp Glu Ser
50 55 60
Leu Phe Arg Gly Arg Gln Ile Lys Val Ile Pro Lys Arg Thr
65 70 75
<210> 39
<211> 73
<212> PRT
<213> 智人
<400> 39
Asn Trp Leu Val Leu Lys Asn Leu Thr Pro Gln Ile Asp Gly Ser Thr
1 5 10 15
Leu Arg Thr Leu Cys Met Gln His Gly Pro Leu Ile Thr Phe His Leu
20 25 30
Asn Leu Pro His Gly Asn Ala Leu Val Arg Tyr Ser Ser Lys Glu Glu
35 40 45
Val Val Lys Ala Gln Lys Ser Leu His Met Cys Val Leu Gly Asn Thr
50 55 60
Thr Ile Leu Ala Glu Phe Ala Ser Glu
65 70
<210> 40
<211> 79
<212> PRT
<213> 智人
<400> 40
Gly Lys Leu Phe Val Gly Gly Leu Ser Phe Asp Thr Asn Glu Gln Ser
1 5 10 15
Leu Glu Gln Val Phe Ser Lys Tyr Gly Gln Ile Ser Glu Val Val Val
20 25 30
Val Lys Asp Arg Glu Thr Gln Arg Ser Arg Gly Phe Gly Phe Val Thr
35 40 45
Phe Glu Asn Ile Asp Asp Ala Lys Asp Ala Met Met Ala Met Asn Gly
50 55 60
Lys Ser Val Asp Gly Arg Gln Ile Arg Val Asp Gln Ala Gly Lys
65 70 75
<210> 41
<211> 74
<212> PRT
<213> 智人
<400> 41
Leu Ile Pro Val Lys Gln Tyr Pro Lys Phe Asn Phe Val Gly Lys Leu
1 5 10 15
Leu Gly Pro Arg Gly Asn Ser Leu Lys Arg Leu Gln Glu Glu Thr Gly
20 25 30
Ala Lys Met Ser Ile Leu Gly Lys Gly Ser Met Arg Asp Lys Ala Lys
35 40 45
Glu Glu Glu Leu Arg Lys Ser Gly Glu Ala Lys Tyr Ala His Leu Ser
50 55 60
Asp Glu Leu His Val Leu Ile Ser Glu Gln
65 70
<210> 42
<211> 67
<212> PRT
<213> 智人
<400> 42
Leu Ile Pro Val Lys Gln Phe Pro Lys Phe Asn Phe Val Gly Lys Leu
1 5 10 15
Leu Gly Pro Arg Gly Asn Ser Leu Lys Arg Leu Gln Glu Glu Thr Leu
20 25 30
Thr Lys Met Ser Ile Leu Gly Lys Gly Ser Met Arg Asp Lys Ala Lys
35 40 45
Glu Glu Glu Leu Arg Lys Ser Gly Glu Ala Lys Tyr Phe His Leu Asn
50 55 60
Asp Asp Leu
65
<210> 43
<211> 62
<212> PRT
<213> 智人
<400> 43
Pro Glu Val Phe Gln Val Gln Thr Arg Leu Leu Lys Ala Ile Phe Gly
1 5 10 15
Pro Asp Gly Ser Arg Ile Pro Tyr Ile Glu Gln Val Ser Lys Ala Met
20 25 30
Leu Glu Leu Lys Ala Leu Glu Ser Ser Asp Leu Thr Glu Val Val Val
35 40 45
Tyr Gly Ser Tyr Leu Tyr Lys Leu Arg Thr Lys Trp Met Leu
50 55 60
<210> 44
<211> 71
<212> PRT
<213> 小家鼠（Mus musculus）
<400> 44
Gln Val Leu Gly Thr Val Lys Trp Phe Asn Val Arg Asn Gly Tyr Gly
1 5 10 15
Phe Ile Asn Arg Asn Asp Thr Lys Glu Asp Val Phe Val His Gln Thr
20 25 30
Ala Ile Lys Arg Asn Asn Pro Arg Lys Phe Leu Arg Ser Val Gly Asp
35 40 45
Gly Glu Thr Val Glu Phe Asp Val Val Glu Gly Glu Lys Gly Ala Arg
50 55 60
Ala Ala Asn Val Thr Gly Pro
65 70
<210> 45
<211> 71
<212> PRT
<213> 小家鼠
<400> 45
Lys Val Leu Gly Thr Val Lys Trp Phe Asn Val Arg Asn Gly Tyr Gly
1 5 10 15
Phe Ile Asn Arg Asn Asp Thr Lys Glu Asp Val Phe Val His Gln Thr
20 25 30
Ala Ile Lys Lys Asn Asn Pro Arg Lys Tyr Leu Arg Ser Val Gly Asp
35 40 45
Gly Glu Thr Val Glu Phe Asp Val Val Glu Gly Glu Lys Gly Ala Glu
50 55 60
Ala Ala Asn Val Thr Gly Pro
65 70
<210> 46
<211> 138
<212> PRT
<213> 智人
<400> 46
Ala Arg Phe Phe Leu Ile Lys Ser Asn Asn His Glu Asn Val Ser Leu
1 5 10 15
Ala Lys Ala Lys Gly Val Trp Ser Thr Leu Pro Val Asn Glu Lys Lys
20 25 30
Leu Asn Leu Ala Phe Arg Ser Ala Arg Ser Val Ile Leu Ile Phe Ser
35 40 45
Val Arg Glu Ser Gly Lys Phe Gln Gly Phe Ala Arg Leu Ser Ser Glu
50 55 60
Ser His His Gly Gly Ser Pro Ile His Trp Val Leu Pro Ala Gly Met
65 70 75 80
Ser Ala Lys Met Leu Gly Gly Val Phe Lys Ile Asp Trp Ile Cys Arg
85 90 95
Arg Glu Leu Pro Phe Thr Lys Ser Ala His Leu Thr Asn Pro Trp Asn
100 105 110
Glu His Lys Pro Val Lys Ile Gly Arg Asp Gly Gln Glu Ile Glu Leu
115 120 125
Glu Cys Gly Thr Gln Leu Cys Leu Leu Phe
130 135
<210> 47
<211> 343
<212> PRT
<213> 智人
<400> 47
Gly Arg Ser Arg Leu Leu Glu Asp Phe Arg Asn Asn Arg Phe Pro Asn
1 5 10 15
Leu Gln Leu Arg Asp Leu Ile Gly His Ile Val Glu Phe Ser Gln Asp
20 25 30
Gln His Gly Ser Arg Phe Ile Gln Gln Lys Leu Glu Arg Ala Thr Pro
35 40 45
Ala Glu Arg Gln Met Val Phe Asn Glu Ile Leu Gln Ala Ala Tyr Gln
50 55 60
Leu Met Thr Asp Val Phe Gly Asn Tyr Val Ile Gln Lys Phe Phe Glu
65 70 75 80
Phe Gly Ser Leu Asp Gln Lys Leu Ala Leu Ala Thr Arg Ile Arg Gly
85 90 95
His Val Leu Pro Leu Ala Leu Gln Met Tyr Gly Cys Arg Val Ile Gln
100 105 110
Lys Ala Leu Glu Ser Ile Ser Ser Asp Gln Gln Val Ile Ser Glu Met
115 120 125
Val Lys Glu Leu Asp Gly His Val Leu Lys Cys Val Lys Asp Gln Asn
130 135 140
Gly Asn His Val Val Gln Lys Cys Ile Glu Cys Val Gln Pro Gln Ser
145 150 155 160
Leu Gln Phe Ile Ile Asp Ala Phe Lys Gly Gln Val Phe Val Leu Ser
165 170 175
Thr His Pro Tyr Gly Cys Arg Val Ile Gln Arg Ile Leu Glu His Cys
180 185 190
Thr Ala Glu Gln Thr Leu Pro Ile Leu Glu Glu Leu His Gln His Thr
195 200 205
Glu Gln Leu Val Gln Asp Gln Tyr Gly Asn Tyr Val Ile Gln His Val
210 215 220
Leu Glu His Gly Arg Pro Glu Asp Lys Ser Lys Ile Val Ser Glu Ile
225 230 235 240
Arg Gly Lys Val Leu Ala Leu Ser Gln His Lys Phe Ala Ser Asn Val
245 250 255
Val Glu Lys Cys Val Thr His Ala Ser Arg Ala Glu Arg Ala Leu Leu
260 265 270
Ile Asp Glu Val Cys Cys Gln Asn Asp Gly Pro His Ser Ala Leu Tyr
275 280 285
Thr Met Met Lys Asp Gln Tyr Ala Asn Tyr Val Val Gln Lys Met Ile
290 295 300
Asp Met Ala Glu Pro Ala Gln Arg Lys Ile Ile Met His Lys Ile Arg
305 310 315 320
Pro His Ile Thr Thr Leu Arg Lys Tyr Thr Tyr Gly Lys His Ile Leu
325 330 335
Ala Lys Leu Glu Lys Tyr Tyr
340
<210> 48
<211> 23
<212> PRT
<213> 智人
<400> 48
Phe Phe Cys Asp Thr Cys Asp Arg Gly Phe Lys Asn Gln Glu Lys Tyr
1 5 10 15
Asp Lys His Met Ser Glu His
20
<210> 49
<211> 23
<212> PRT
<213> 智人
<400> 49
Cys Gln Met Cys Gln Lys Gln Cys Arg Asp Glu Asn Gly Phe Lys Cys
1 5 10 15
His Cys Met Ser Glu Ser His
20
<210> 50
<211> 79
<212> PRT
<213> 智人
<400> 50
Thr Ser Leu Lys Val Asp Asn Leu Thr Tyr Arg Thr Ser Pro Asp Thr
1 5 10 15
Leu Arg Arg Val Phe Glu Lys Tyr Gly Arg Val Gly Asp Val Tyr Ile
20 25 30
Pro Arg Asp Arg Tyr Thr Lys Glu Ser Arg Gly Phe Ala Phe Val Arg
35 40 45
Phe His Asp Lys Arg Asp Ala Glu Asp Ala Met Asp Ala Met Asp Gly
50 55 60
Ala Val Leu Asp Gly Arg Glu Leu Arg Val Gln Met Ala Arg Tyr
65 70 75
<210> 51
<211> 84
<212> PRT
<213> 黑腹果蝇（Drosophila melanogaster）
<400> 51
Gln Leu Phe Leu Gly Asn Ile Pro His His Ala Ser Glu Asp Asp Leu
1 5 10 15
Arg Glu Ile Phe Ser Arg Phe Gly Asn Val Leu Glu Leu Arg Ile Leu
20 25 30
Ser Lys Ala Gly Asn Lys Val Pro Pro Gly Met Arg Ser Pro Leu Asn
35 40 45
Tyr Gly Phe Ile Thr Tyr Asp Asp Pro Glu Ala Val Gln Lys Cys Leu
50 55 60
Ala Asn Cys Pro Leu Tyr Phe Pro Glu Asn Ser Pro Asp Gly Gln Lys
65 70 75 80
Leu Asn Val Glu
<210> 52
<211> 79
<212> PRT
<213> 智人
<400> 52
Gly Lys Leu Phe Val Gly Gly Leu Asn Phe Asn Thr Asp Glu Gln Ala
1 5 10 15
Leu Glu Asp His Phe Ser Ser Phe Gly Pro Ile Ser Glu Val Val Val
20 25 30
Val Lys Asp Arg Glu Thr Gln Arg Ser Arg Gly Phe Gly Phe Ile Thr
35 40 45
Phe Thr Asn Pro Glu His Ala Ser Val Ala Met Arg Ala Met Asn Gly
50 55 60
Glu Ser Leu Asp Gly Arg Gln Ile Arg Val Asp His Ala Gly Lys
65 70 75
<210> 53
<211> 683
<212> PRT
<213> 莫洛尼鼠类白血病病毒
<400> 53
Thr Leu Asn Ile Glu Asp Glu His Arg Leu His Glu Thr Ser Lys Glu
1 5 10 15
Pro Asp Val Ser Leu Gly Ser Thr Trp Leu Ser Asp Phe Pro Gln Ala
20 25 30
Trp Ala Glu Thr Gly Gly Met Gly Leu Ala Val Arg Gln Ala Pro Leu
35 40 45
Ile Ile Pro Leu Lys Ala Thr Ser Thr Pro Val Ser Ile Lys Gln Tyr
50 55 60
Pro Met Ser Gln Glu Ala Arg Leu Gly Ile Lys Pro His Ile Gln Arg
65 70 75 80
Leu Leu Asp Gln Gly Ile Leu Val Pro Cys Gln Ser Pro Trp Asn Thr
85 90 95
Pro Leu Leu Pro Val Lys Lys Pro Gly Thr Asn Asp Tyr Arg Pro Val
100 105 110
Gln Asp Leu Arg Glu Val Asn Lys Arg Val Glu Asp Ile His Pro Thr
115 120 125
Val Pro Asn Pro Tyr Asn Leu Leu Ser Gly Leu Pro Pro Ser His Gln
130 135 140
Trp Tyr Thr Val Leu Asp Leu Lys Asp Ala Phe Phe Cys Leu Arg Leu
145 150 155 160
His Pro Thr Ser Gln Pro Leu Phe Ala Phe Glu Trp Arg Asp Pro Glu
165 170 175
Met Gly Ile Ser Gly Gln Leu Thr Trp Thr Arg Leu Pro Gln Gly Phe
180 185 190
Lys Asn Ser Pro Thr Leu Phe Asp Glu Ala Leu His Arg Asp Leu Ala
195 200 205
Asp Phe Arg Ile Gln His Pro Asp Leu Ile Leu Leu Gln Tyr Val Asp
210 215 220
Asp Leu Leu Leu Ala Ala Thr Ser Glu Leu Asp Cys Gln Gln Gly Thr
225 230 235 240
Arg Ala Leu Leu Gln Thr Leu Gly Asn Leu Gly Tyr Arg Ala Ser Ala
245 250 255
Lys Lys Ala Gln Ile Cys Gln Lys Gln Val Lys Tyr Leu Gly Tyr Leu
260 265 270
Leu Lys Glu Gly Gln Arg Trp Leu Thr Glu Ala Arg Lys Glu Thr Val
275 280 285
Met Gly Gln Pro Thr Pro Lys Thr Pro Arg Gln Leu Arg Glu Phe Leu
290 295 300
Gly Thr Ala Gly Phe Cys Arg Leu Trp Ile Pro Gly Phe Ala Glu Met
305 310 315 320
Ala Ala Pro Leu Tyr Pro Leu Thr Lys Thr Gly Thr Leu Phe Asn Trp
325 330 335
Gly Pro Asp Gln Gln Lys Ala Tyr Gln Glu Ile Lys Gln Ala Leu Leu
340 345 350
Thr Ala Pro Ala Leu Gly Leu Pro Asp Leu Thr Lys Pro Phe Glu Leu
355 360 365
Phe Val Asp Glu Lys Gln Gly Tyr Ala Lys Gly Val Leu Thr Gln Lys
370 375 380
Leu Gly Pro Trp Arg Arg Pro Val Ala Tyr Leu Ser Lys Lys Leu Asp
385 390 395 400
Pro Val Ala Ala Gly Trp Pro Pro Cys Leu Arg Met Val Ala Ala Ile
405 410 415
Ala Val Leu Thr Lys Asp Ala Gly Lys Leu Thr Met Gly Gln Pro Leu
420 425 430
Val Ile Leu Ala Pro His Ala Val Glu Ala Leu Val Lys Gln Pro Pro
435 440 445
Asp Arg Trp Leu Ser Asn Ala Arg Met Thr His Tyr Gln Ala Leu Leu
450 455 460
Leu Asp Thr Asp Arg Val Gln Phe Gly Pro Val Val Ala Leu Asn Pro
465 470 475 480
Ala Thr Leu Leu Pro Leu Pro Glu Glu Gly Leu Gln His Asn Cys Leu
485 490 495
Asp Ile Leu Ala Glu Ala His Gly Thr Arg Pro Asp Leu Thr Asp Gln
500 505 510
Pro Leu Pro Asp Ala Asp His Thr Trp Tyr Thr Asp Gly Ser Ser Leu
515 520 525
Leu Gln Glu Gly Gln Arg Lys Ala Gly Ala Ala Val Thr Thr Glu Thr
530 535 540
Glu Val Ile Trp Ala Lys Ala Leu Pro Ala Gly Thr Ser Ala Gln Arg
545 550 555 560
Ala Glu Leu Ile Ala Leu Thr Gln Ala Leu Lys Met Ala Glu Gly Lys
565 570 575
Lys Leu Asn Val Tyr Thr Asp Ser Arg Tyr Ala Phe Ala Thr Ala His
580 585 590
Ile His Gly Glu Ile Tyr Arg Arg Arg Gly Leu Leu Thr Ser Glu Gly
595 600 605
Lys Glu Ile Lys Asn Lys Asp Glu Ile Leu Ala Leu Leu Lys Ala Leu
610 615 620
Phe Leu Pro Lys Arg Leu Ser Ile Ile His Cys Pro Gly His Gln Lys
625 630 635 640
Gly His Ser Ala Glu Ala Arg Gly Asn Arg Met Ala Asp Gln Ala Ala
645 650 655
Arg Lys Ala Ala Ile Thr Glu Thr Pro Asp Thr Ser Thr Leu Leu Ile
660 665 670
Glu Asn Ser Ser Pro Asn Ser Arg Leu Ile Asn
675 680
<210> 54
<211> 19
<212> DNA
<213> 人工的
<220>
<223> 合成的RNA间隔子
<220>
<221> misc_feature
<222> (1)..(19)
<223> 其中n为A, C, T或G
<400> 54
nnnnnnnnnn nnnnnnnnn 19
<210> 55
<211> 22
<212> DNA
<213> 人工的
<220>
<223> 合成的靶标链
<220>
<221> misc_feature
<222> (4)..(22)
<223> 其中n为A, C, T或G
<400> 55
aaannnnnnn nnnnnnnnnn nn 22
<210> 56
<211> 22
<212> DNA
<213> 人工的
<220>
<223> 合成的非靶标链
<220>
<221> misc_feature
<222> (4)..(22)
<223> 其中n为A, C, T或G
<400> 56
tttnnnnnnn nnnnnnnnnn nn 22
<210> 57
<211> 3790
<212> DNA
<213> 人工的
<220>
<223> 合成的多核苷酸
<400> 57
ccatgggcag caaactggaa aaatttacga attgttatag cctgtccaag accctgcgtt 60
tcaaagccat ccccgttggc aaaacccagg agaatattga taataaacgt ctgctggttg 120
aggatgaaaa aagagcagaa gactataagg gagtcaaaaa actgctggat cggtactacc 180
tgagctttat aaatgacgtg ctgcatagca ttaaactgaa aaatctgaat aactatatta 240
gtctgttccg caagaaaacc cgaacagaga aagaaaataa agagctggaa aacctggaga 300
tcaatctgcg taaagagatc gcaaaagctt ttaaaggaaa tgaaggttat aaaagcctgt 360
tcaaaaaaga cattattgaa accatcctgc cggaatttct ggatgataaa gacgagatag 420
cgctcgtgaa cagcttcaac gggttcacga ccgccttcac gggctttttc gataacaggg 480
aaaatatgtt ttcagaggaa gccaaaagca cctcgatagc gttccgttgc attaatgaaa 540
atttgacaag atatatcagc aacatggata ttttcgagaa agttgatgcg atctttgaca 600
aacatgaagt gcaggagatt aaggaaaaaa ttctgaacag cgattatgat gttgaggatt 660
ttttcgaggg ggaatttttt aactttgtac tgacacagga aggtatagat gtgtataatg 720
ctattatcgg cgggttcgtt accgaatccg gcgagaaaat taagggtctg aatgagtaca 780
tcaatctgta taaccaaaag accaaacaga aactgccaaa attcaaaccg ctgtacaagc 840
aagtcctgag cgatcgggaa agcttgagct tttacggtga aggttatacc agcgacgagg 900
aggtactgga ggtctttcgc aataccctga acaagaacag cgaaattttc agctccatta 960
aaaagctgga gaaactgttt aagaattttg acgagtacag cagcgcaggt atttttgtga 1020
agaacggacc tgccataagc accattagca aggatatttt tggagagtgg aatgttatcc 1080
gtgataaatg gaacgcggaa tatgatgaca tacacctgaa aaagaaggct gtggtaactg 1140
agaaatatga agacgatcgc cgcaaaagct ttaaaaaaat cggcagcttt agcctggagc 1200
agctgcagga atatgcggac gccgacctga gcgtggtcga gaaactgaag gaaattatta 1260
tccaaaaagt ggatgagatt tacaaggtat atggtagcag cgaaaaactg tttgatgcgg 1320
acttcgttct ggaaaaaagc ctgaaaaaaa atgatgctgt tgttgcgatc atgaaagacc 1380
tgctcgatag cgttaagagc tttgaaaatt acattaaagc attctttggc gagggcaaag 1440
aaacaaacag agacgaaagc ttttatggcg acttcgtcct ggcttatgac atcctgttga 1500
aggtagatca tatatatgat gcaattcgta attacgtaac ccaaaagccg tacagcaaag 1560
ataagttcaa actgtatttc cagaacccgc agtttatggg tggctgggac aaagacaagg 1620
agacagacta tcgcgccact attctgcgtt acggcagcaa gtactatctc gccatcatgg 1680
acaaaaaata tgcaaagtgt ctgcagaaaa tcgataaaga cgacgtgaac ggaaattacg 1740
aaaagattaa ttataagctg ctgccagggc ccaacaagat gttaccgaaa gtattttttt 1800
ccaaaaaatg gatggcatac tataacccga gcgaggatat acagaagatt tacaaaaatg 1860
ggaccttcaa aaagggggat atgttcaatc tgaatgactg ccacaaactg atcgattttt 1920
ttaaagatag catcagccgt tatcctaaat ggtcaaacgc gtatgatttt aatttctccg 1980
aaacggagaa atataaagac attgctggtt tctatcgcga agtcgaagaa cagggttata 2040
aagttagctt tgaatcggcc agcaagaaag aggttgataa actggtggag gagggtaagc 2100
tgtatatgtt tcagatttat aacaaagact ttagcgacaa aagccacggt actcctaatc 2160
tgcatacgat gtactttaaa ctgctgtttg atgagaataa ccacggccaa atccgtctct 2220
ccggtggagc agaacttttt atgcggcgtg cgagcctaaa aaaggaagaa ctggtggtgc 2280
atcccgccaa cagcccgatt gctaacaaaa atccagataa tcctaagaag accaccacac 2340
tgtcgtacga tgtctataag gataaacgtt tctcggaaga ccagtatgaa ttgcatatac 2400
cgatagcaat taataaatgc ccaaaaaaca ttttcaaaat caacactgaa gttcgtgtgc 2460
tgctgaaaca tgatgataat ccgtatgtga tcggaattga ccgtggggag agaaatctgc 2520
tgtatattgt agtcgttgat ggcaagggca acatcgttga gcagtatagc ctgaatgaaa 2580
taattaataa ttttaacggt atacgtatta aaaccgacta tcatagcctg ctggataaaa 2640
aggagaaaga gcgttttgag gcacgccaaa attggacgag catcgaaaac atcaaggaac 2700
tgaaggcagg atatatcagc caagtagtcc ataaaatctg tgaactggtg gagaagtacg 2760
acgctgtcat tgccctggaa gacctcaata gcggctttaa aaacagccgg gtgaaggtgg 2820
agaaacaggt ataccaaaag tttgaaaaga tgctcattga taagctgaac tatatggttg 2880
ataaaaagag caacccgtgc gccactggcg gtgcactgaa agggtaccaa attaccaata 2940
aatttgaaag ctttaaaagc atgagcacgc agaatgggtt tattttttat ataccagcat 3000
ggctgacgag caagattgac cccagcactg gttttgtcaa tctgctgaaa accaaataca 3060
caagcattgc ggatagcaaa aaatttattt cgagcttcga ccgtattatg tatgttccgg 3120
aggaagatct gtttgaattt gccctggatt ataaaaactt cagccgcacc gatgcagatt 3180
atatcaaaaa atggaagctg tacagttatg gtaatcgtat acgtatcttc cgtaatccga 3240
agaaaaacaa tgtgttcgat tgggaagagg tctgtctgac cagcgcgtat aaagaactgt 3300
tcaacaagta cggaataaat tatcagcaag gtgacattcg cgcactgctg tgtgaacagt 3360
cagataaagc attttatagc agctttatgg cgctgatgag cctgatgctc cagatgcgca 3420
acagcataac cggtcgcaca gatgttgact ttctgatcag ccctgtgaag aatagcgacg 3480
gcatcttcta cgattccagg aactatgaag cacaggaaaa cgctattctg cctaaaaatg 3540
ccgatgccaa cggcgcctat aatattgcac ggaaggttct gtgggcgatt ggacagttca 3600
agaaagcgga agatgagaag ctggataagg taaaaattgc tattagcaat aaggaatggc 3660
tggagtacgc acagacatcg gttaaacacg gtagtaaaag gccggcggcc acgaaaaagg 3720
ccggccaggc aaaaaagaaa aagggagcgg ccgcactcga gcaccaccac caccaccact 3780
gagcggccgc 3790
<210> 58
<211> 8986
<212> DNA
<213> 人工的
<220>
<223> 合成的质粒
<400> 58
tggcgaatgg gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg 60
cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc 120
ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg 180
gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc 240
acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt 300
ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc 360
ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta 420
acaaaaattt aacgcgaatt ttaacaaaat attaacgttt acaatttcag gtggcacttt 480
tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta 540
tccgctcatg aattaattct tagaaaaact catcgagcat caaatgaaac tgcaatttat 600
tcatatcagg attatcaata ccatattttt gaaaaagccg tttctgtaat gaaggagaaa 660
actcaccgag gcagttccat aggatggcaa gatcctggta tcggtctgcg attccgactc 720
gtccaacatc aatacaacct attaatttcc cctcgtcaaa aataaggtta tcaagtgaga 780
aatcaccatg agtgacgact gaatccggtg agaatggcaa aagtttatgc atttctttcc 840
agacttgttc aacaggccag ccattacgct cgtcatcaaa atcactcgca tcaaccaaac 900
cgttattcat tcgtgattgc gcctgagcga gacgaaatac gcgatcgctg ttaaaaggac 960
aattacaaac aggaatcgaa tgcaaccggc gcaggaacac tgccagcgca tcaacaatat 1020
tttcacctga atcaggatat tcttctaata cctggaatgc tgttttcccg gggatcgcag 1080
tggtgagtaa ccatgcatca tcaggagtac ggataaaatg cttgatggtc ggaagaggca 1140
taaattccgt cagccagttt agtctgacca tctcatctgt aacatcattg gcaacgctac 1200
ctttgccatg tttcagaaac aactctggcg catcgggctt cccatacaat cgatagattg 1260
tcgcacctga ttgcccgaca ttatcgcgag cccatttata cccatataaa tcagcatcca 1320
tgttggaatt taatcgcggc ctagagcaag acgtttcccg ttgaatatgg ctcataacac 1380
cccttgtatt actgtttatg taagcagaca gttttattgt tcatgaccaa aatcccttaa 1440
cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga 1500
gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg 1560
gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc 1620
agagcgcaga taccaaatac tgtccttcta gtgtagccgt agttaggcca ccacttcaag 1680
aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc 1740
agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg 1800
cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac 1860
accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga 1920
aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt 1980
ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag 2040
cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg 2100
gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta 2160
tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc 2220
agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cctgatgcgg 2280
tattttctcc ttacgcatct gtgcggtatt tcacaccgca tatatggtgc actctcagta 2340
caatctgctc tgatgccgca tagttaagcc agtatacact ccgctatcgc tacgtgactg 2400
ggtcatggct gcgccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct 2460
gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag 2520
gttttcaccg tcatcaccga aacgcgcgag gcagctgcgg taaagctcat cagcgtggtc 2580
gtgaagcgat tcacagatgt ctgcctgttc atccgcgtcc agctcgttga gtttctccag 2640
aagcgttaat gtctggcttc tgataaagcg ggccatgtta agggcggttt tttcctgttt 2700
ggtcactgat gcctccgtgt aagggggatt tctgttcatg ggggtaatga taccgatgaa 2760
acgagagagg atgctcacga tacgggttac tgatgatgaa catgcccggt tactggaacg 2820
ttgtgagggt aaacaactgg cggtatggat gcggcgggac cagagaaaaa tcactcaggg 2880
tcaatgccag cgcttcgtta atacagatgt aggtgttcca cagggtagcc agcagcatcc 2940
tgcgatgcag atccggaaca taatggtgca gggcgctgac ttccgcgttt ccagacttta 3000
cgaaacacgg aaaccgaaga ccattcatgt tgttgctcag gtcgcagacg ttttgcagca 3060
gcagtcgctt cacgttcgct cgcgtatcgg tgattcattc tgctaaccag taaggcaacc 3120
ccgccagcct agccgggtcc tcaacgacag gagcacgatc atgcgcaccc gtggggccgc 3180
catgccggcg ataatggcct gcttctcgcc gaaacgtttg gtggcgggac cagtgacgaa 3240
ggcttgagcg agggcgtgca agattccgaa taccgcaagc gacaggccga tcatcgtcgc 3300
gctccagcga aagcggtcct cgccgaaaat gacccagagc gctgccggca cctgtcctac 3360
gagttgcatg ataaagaaga cagtcataag tgcggcgacg atagtcatgc cccgcgccca 3420
ccggaaggag ctgactgggt tgaaggctct caagggcatc ggtcgagatc ccggtgccta 3480
atgagtgagc taacttacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa 3540
cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat 3600
tgggcgccag ggtggttttt cttttcacca gtgagacggg caacagctga ttgcccttca 3660
ccgcctggcc ctgagagagt tgcagcaagc ggtccacgct ggtttgcccc agcaggcgaa 3720
aatcctgttt gatggtggtt aacggcggga tataacatga gctgtcttcg gtatcgtcgt 3780
atcccactac cgagatatcc gcaccaacgc gcagcccgga ctcggtaatg gcgcgcattg 3840
cgcccagcgc catctgatcg ttggcaacca gcatcgcagt gggaacgatg ccctcattca 3900
gcatttgcat ggtttgttga aaaccggaca tggcactcca gtcgccttcc cgttccgcta 3960
tcggctgaat ttgattgcga gtgagatatt tatgccagcc agccagacgc agacgcgccg 4020
agacagaact taatgggccc gctaacagcg cgatttgctg gtgacccaat gcgaccagat 4080
gctccacgcc cagtcgcgta ccgtcttcat gggagaaaat aatactgttg atgggtgtct 4140
ggtcagagac atcaagaaat aacgccggaa cattagtgca ggcagcttcc acagcaatgg 4200
catcctggtc atccagcgga tagttaatga tcagcccact gacgcgttgc gcgagaagat 4260
tgtgcaccgc cgctttacag gcttcgacgc cgcttcgttc taccatcgac accaccacgc 4320
tggcacccag ttgatcggcg cgagatttaa tcgccgcgac aatttgcgac ggcgcgtgca 4380
gggccagact ggaggtggca acgccaatca gcaacgactg tttgcccgcc agttgttgtg 4440
ccacgcggtt gggaatgtaa ttcagctccg ccatcgccgc ttccactttt tcccgcgttt 4500
tcgcagaaac gtggctggcc tggttcacca cgcgggaaac ggtctgataa gagacaccgg 4560
catactctgc gacatcgtat aacgttactg gtttcacatt caccaccctg aattgactct 4620
cttccgggcg ctatcatgcc ataccgcgaa aggttttgcg ccattcgatg gtgtccggga 4680
tctcgacgct ctcccttatg cgactcctgc attaggaagc agcccagtag taggttgagg 4740
ccgttgagca ccgccgccgc aaggaatggt gcatgcaagg agatggcgcc caacagtccc 4800
ccggccacgg ggcctgccac catacccacg ccgaaacaag cgctcatgag cccgaagtgg 4860
cgagcccgat cttccccatc ggtgatgtcg gcgatatagg cgccagcaac cgcacctgtg 4920
gcgccggtga tgccggccac gatgcgtccg gcgtagagga tcgagatctc gatcccgcga 4980
aattaatacg actcactata ggggaattgt gagcggataa caattcccct ctagaaataa 5040
ttttgtttaa ctttaagaag gagatatacc atgggcagca aactggaaaa atttacgaat 5100
tgttatagcc tgtccaagac cctgcgtttc aaagccatcc ccgttggcaa aacccaggag 5160
aatattgata ataaacgtct gctggttgag gatgaaaaaa gagcagaaga ctataaggga 5220
gtcaaaaaac tgctggatcg gtactacctg agctttataa atgacgtgct gcatagcatt 5280
aaactgaaaa atctgaataa ctatattagt ctgttccgca agaaaacccg aacagagaaa 5340
gaaaataaag agctggaaaa cctggagatc aatctgcgta aagagatcgc aaaagctttt 5400
aaaggaaatg aaggttataa aagcctgttc aaaaaagaca ttattgaaac catcctgccg 5460
gaatttctgg atgataaaga cgagatagcg ctcgtgaaca gcttcaacgg gttcacgacc 5520
gccttcacgg gctttttcga taacagggaa aatatgtttt cagaggaagc caaaagcacc 5580
tcgatagcgt tccgttgcat taatgaaaat ttgacaagat atatcagcaa catggatatt 5640
ttcgagaaag ttgatgcgat ctttgacaaa catgaagtgc aggagattaa ggaaaaaatt 5700
ctgaacagcg attatgatgt tgaggatttt ttcgaggggg aattttttaa ctttgtactg 5760
acacaggaag gtatagatgt gtataatgct attatcggcg ggttcgttac cgaatccggc 5820
gagaaaatta agggtctgaa tgagtacatc aatctgtata accaaaagac caaacagaaa 5880
ctgccaaaat tcaaaccgct gtacaagcaa gtcctgagcg atcgggaaag cttgagcttt 5940
tacggtgaag gttataccag cgacgaggag gtactggagg tctttcgcaa taccctgaac 6000
aagaacagcg aaattttcag ctccattaaa aagctggaga aactgtttaa gaattttgac 6060
gagtacagca gcgcaggtat ttttgtgaag aacggacctg ccataagcac cattagcaag 6120
gatatttttg gagagtggaa tgttatccgt gataaatgga acgcggaata tgatgacata 6180
cacctgaaaa agaaggctgt ggtaactgag aaatatgaag acgatcgccg caaaagcttt 6240
aaaaaaatcg gcagctttag cctggagcag ctgcaggaat atgcggacgc cgacctgagc 6300
gtggtcgaga aactgaagga aattattatc caaaaagtgg atgagattta caaggtatat 6360
ggtagcagcg aaaaactgtt tgatgcggac ttcgttctgg aaaaaagcct gaaaaaaaat 6420
gatgctgttg ttgcgatcat gaaagacctg ctcgatagcg ttaagagctt tgaaaattac 6480
attaaagcat tctttggcga gggcaaagaa acaaacagag acgaaagctt ttatggcgac 6540
ttcgtcctgg cttatgacat cctgttgaag gtagatcata tatatgatgc aattcgtaat 6600
tacgtaaccc aaaagccgta cagcaaagat aagttcaaac tgtatttcca gaacccgcag 6660
tttatgggtg gctgggacaa agacaaggag acagactatc gcgccactat tctgcgttac 6720
ggcagcaagt actatctcgc catcatggac aaaaaatatg caaagtgtct gcagaaaatc 6780
gataaagacg acgtgaacgg aaattacgaa aagattaatt ataagctgct gccagggccc 6840
aacaagatgt taccgaaagt atttttttcc aaaaaatgga tggcatacta taacccgagc 6900
gaggatatac agaagattta caaaaatggg accttcaaaa agggggatat gttcaatctg 6960
aatgactgcc acaaactgat cgattttttt aaagatagca tcagccgtta tcctaaatgg 7020
tcaaacgcgt atgattttaa tttctccgaa acggagaaat ataaagacat tgctggtttc 7080
tatcgcgaag tcgaagaaca gggttataaa gttagctttg aatcggccag caagaaagag 7140
gttgataaac tggtggagga gggtaagctg tatatgtttc agatttataa caaagacttt 7200
agcgacaaaa gccacggtac tcctaatctg catacgatgt actttaaact gctgtttgat 7260
gagaataacc acggccaaat ccgtctctcc ggtggagcag aactttttat gcggcgtgcg 7320
agcctaaaaa aggaagaact ggtggtgcat cccgccaaca gcccgattgc taacaaaaat 7380
ccagataatc ctaagaagac caccacactg tcgtacgatg tctataagga taaacgtttc 7440
tcggaagacc agtatgaatt gcatataccg atagcaatta ataaatgccc aaaaaacatt 7500
ttcaaaatca acactgaagt tcgtgtgctg ctgaaacatg atgataatcc gtatgtgatc 7560
ggaattgacc gtggggagag aaatctgctg tatattgtag tcgttgatgg caagggcaac 7620
atcgttgagc agtatagcct gaatgaaata attaataatt ttaacggtat acgtattaaa 7680
accgactatc atagcctgct ggataaaaag gagaaagagc gttttgaggc acgccaaaat 7740
tggacgagca tcgaaaacat caaggaactg aaggcaggat atatcagcca agtagtccat 7800
aaaatctgtg aactggtgga gaagtacgac gctgtcattg ccctggaaga cctcaatagc 7860
ggctttaaaa acagccgggt gaaggtggag aaacaggtat accaaaagtt tgaaaagatg 7920
ctcattgata agctgaacta tatggttgat aaaaagagca acccgtgcgc cactggcggt 7980
gcactgaaag ggtaccaaat taccaataaa tttgaaagct ttaaaagcat gagcacgcag 8040
aatgggttta ttttttatat accagcatgg ctgacgagca agattgaccc cagcactggt 8100
tttgtcaatc tgctgaaaac caaatacaca agcattgcgg atagcaaaaa atttatttcg 8160
agcttcgacc gtattatgta tgttccggag gaagatctgt ttgaatttgc cctggattat 8220
aaaaacttca gccgcaccga tgcagattat atcaaaaaat ggaagctgta cagttatggt 8280
aatcgtatac gtatcttccg taatccgaag aaaaacaatg tgttcgattg ggaagaggtc 8340
tgtctgacca gcgcgtataa agaactgttc aacaagtacg gaataaatta tcagcaaggt 8400
gacattcgcg cactgctgtg tgaacagtca gataaagcat tttatagcag ctttatggcg 8460
ctgatgagcc tgatgctcca gatgcgcaac agcataaccg gtcgcacaga tgttgacttt 8520
ctgatcagcc ctgtgaagaa tagcgacggc atcttctacg attccaggaa ctatgaagca 8580
caggaaaacg ctattctgcc taaaaatgcc gatgccaacg gcgcctataa tattgcacgg 8640
aaggttctgt gggcgattgg acagttcaag aaagcggaag atgagaagct ggataaggta 8700
aaaattgcta ttagcaataa ggaatggctg gagtacgcac agacatcggt taaacacggt 8760
agtaaaaggc cggcggccac gaaaaaggcc ggccaggcaa aaaagaaaaa gggagcggcc 8820
gcactcgagc accaccacca ccaccactga gatccggctg ctaacaaagc ccgaaaggaa 8880
gctgagttgg ctgctgccac cgctgagcaa taactagcat aaccccttgg ggcctctaaa 8940
cgggtcttga ggggtttttt gctgaaagga ggaactatat ccggat 8986
<210> 59
<211> 43
<212> RNA
<213> 人工的
<220>
<223> 合成的CRISPR RNA
<400> 59
aauuucuacu aaguguagau ggaaucccuu cugcagcacc ugg 43
<210> 60
<211> 28
<212> DNA
<213> 人工的
<220>
<223> 合成的原间隔子
<400> 60
ctttcggaat cccttctgca gcacctgg 28
<210> 61
<211> 133
<212> DNA
<213> 智人
<400> 61
gtaagtatca aggttacaag acaggtttaa ggagaccaat agaaactggg cttgtcgaga 60
cagagaagac tcttgcgttt ctgataggca cctattggtc ttactgacat ccactttgcc 120
tttctctcca cag 133
<210> 62
<211> 99
<212> DNA
<213> 猿猴病毒40
<400> 62
gtaagtttag tctttttgtc ttttatttca ggtcccggat ccggtggtgg tgcaaatcaa 60
agaactgctc ctcagtggat gttgccttta cttctaggc 99
<210> 63
<211> 1963
<212> PRT
<213> 人工的
<220>
<223> 合成的REDRAW编辑器
<400> 63
Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys
1 5 10 15
Arg Lys Val Thr Leu Asn Ile Glu Asp Glu Tyr Arg Leu His Glu Thr
20 25 30
Ser Lys Glu Pro Asp Val Ser Leu Gly Ser Thr Trp Leu Ser Asp Phe
35 40 45
Pro Gln Ala Trp Ala Glu Thr Gly Gly Met Gly Leu Ala Val Arg Gln
50 55 60
Ala Pro Leu Ile Ile Pro Leu Lys Ala Thr Ser Thr Pro Val Ser Ile
65 70 75 80
Lys Gln Tyr Pro Met Ser Gln Glu Ala Arg Leu Gly Ile Lys Pro His
85 90 95
Ile Gln Arg Leu Leu Asp Gln Gly Ile Leu Val Pro Cys Gln Ser Pro
100 105 110
Trp Asn Thr Pro Leu Leu Pro Val Lys Lys Pro Gly Thr Asn Asp Tyr
115 120 125
Arg Pro Val Gln Asp Leu Arg Glu Val Asn Lys Arg Val Glu Asp Ile
130 135 140
His Pro Thr Val Pro Asn Pro Tyr Asn Leu Leu Ser Gly Leu Pro Pro
145 150 155 160
Ser His Gln Trp Tyr Thr Val Leu Asp Leu Lys Asp Ala Phe Phe Cys
165 170 175
Leu Arg Leu His Pro Thr Ser Gln Pro Leu Phe Ala Phe Glu Trp Arg
180 185 190
Asp Pro Glu Met Gly Ile Ser Gly Gln Leu Thr Trp Thr Arg Leu Pro
195 200 205
Gln Gly Phe Lys Asn Ser Pro Thr Leu Phe Asn Glu Ala Leu His Arg
210 215 220
Asp Leu Ala Asp Phe Arg Ile Gln His Pro Asp Leu Ile Leu Leu Gln
225 230 235 240
Tyr Val Asp Asp Leu Leu Leu Ala Ala Thr Ser Glu Leu Asp Cys Gln
245 250 255
Gln Gly Thr Arg Ala Leu Leu Gln Thr Leu Gly Asn Leu Gly Tyr Arg
260 265 270
Ala Ser Ala Lys Lys Ala Gln Ile Cys Gln Lys Gln Val Lys Tyr Leu
275 280 285
Gly Tyr Leu Leu Lys Glu Gly Gln Arg Trp Leu Thr Glu Ala Arg Lys
290 295 300
Glu Thr Val Met Gly Gln Pro Thr Pro Lys Thr Pro Arg Gln Leu Arg
305 310 315 320
Glu Phe Leu Gly Lys Ala Gly Phe Cys Arg Leu Phe Ile Pro Gly Phe
325 330 335
Ala Glu Met Ala Ala Pro Leu Tyr Pro Leu Thr Lys Pro Gly Thr Leu
340 345 350
Phe Asn Trp Gly Pro Asp Gln Gln Lys Ala Tyr Gln Glu Ile Lys Gln
355 360 365
Ala Leu Leu Thr Ala Pro Ala Leu Gly Leu Pro Asp Leu Thr Lys Pro
370 375 380
Phe Glu Leu Phe Val Asp Glu Lys Gln Gly Tyr Ala Lys Gly Val Leu
385 390 395 400
Thr Gln Lys Leu Gly Pro Trp Arg Arg Pro Val Ala Tyr Leu Ser Lys
405 410 415
Lys Leu Asp Pro Val Ala Ala Gly Trp Pro Pro Cys Leu Arg Met Val
420 425 430
Ala Ala Ile Ala Val Leu Thr Lys Asp Ala Gly Lys Leu Thr Met Gly
435 440 445
Gln Pro Leu Val Ile Leu Ala Pro His Ala Val Glu Ala Leu Val Lys
450 455 460
Gln Pro Pro Asp Arg Trp Leu Ser Asn Ala Arg Met Thr His Tyr Gln
465 470 475 480
Ala Leu Leu Leu Asp Thr Asp Arg Val Gln Phe Gly Pro Val Val Ala
485 490 495
Leu Asn Pro Ala Thr Leu Leu Pro Leu Pro Glu Glu Gly Leu Gln His
500 505 510
Asn Cys Leu Asp Ile Leu Ala Glu Ala His Gly Thr Arg Pro Asp Leu
515 520 525
Thr Asp Gln Pro Leu Pro Asp Ala Asp His Thr Trp Tyr Thr Asp Gly
530 535 540
Ser Ser Leu Leu Gln Glu Gly Gln Arg Lys Ala Gly Ala Ala Val Thr
545 550 555 560
Thr Glu Thr Glu Val Ile Trp Ala Lys Ala Leu Pro Ala Gly Thr Ser
565 570 575
Ala Gln Arg Ala Glu Leu Ile Ala Leu Thr Gln Ala Leu Lys Met Ala
580 585 590
Glu Gly Lys Lys Leu Asn Val Tyr Thr Asp Ser Arg Tyr Ala Phe Ala
595 600 605
Thr Ala His Ile His Gly Glu Ile Tyr Arg Arg Arg Gly Trp Leu Thr
610 615 620
Ser Glu Gly Lys Glu Ile Lys Asn Lys Asp Glu Ile Leu Ala Leu Leu
625 630 635 640
Lys Ala Leu Phe Leu Pro Lys Arg Leu Ser Ile Ile His Cys Pro Gly
645 650 655
His Gln Lys Gly His Ser Ala Glu Ala Arg Gly Asn Arg Met Ala Asp
660 665 670
Gln Ala Ala Arg Lys Ala Ala Ile Thr Glu Thr Pro Asp Thr Ser Thr
675 680 685
Leu Leu Ile Glu Asn Ser Ser Pro Ser Gly Ser Glu Thr Pro Gly Thr
690 695 700
Ser Glu Ser Ala Thr Pro Glu Ser Ser Lys Leu Glu Lys Phe Thr Asn
705 710 715 720
Cys Tyr Ser Leu Ser Lys Thr Leu Arg Phe Lys Ala Ile Pro Val Gly
725 730 735
Lys Thr Gln Glu Asn Ile Asp Asn Lys Arg Leu Leu Val Glu Asp Glu
740 745 750
Lys Arg Ala Glu Asp Tyr Lys Gly Val Lys Lys Leu Leu Asp Arg Tyr
755 760 765
Tyr Leu Ser Phe Ile Asn Asp Val Leu His Ser Ile Lys Leu Lys Asn
770 775 780
Leu Asn Asn Tyr Ile Ser Leu Phe Arg Lys Lys Thr Arg Thr Glu Lys
785 790 795 800
Glu Asn Lys Glu Leu Glu Asn Leu Glu Ile Asn Leu Arg Lys Glu Ile
805 810 815
Ala Lys Ala Phe Lys Gly Asn Glu Gly Tyr Lys Ser Leu Phe Lys Lys
820 825 830
Asp Ile Ile Glu Thr Ile Leu Pro Glu Phe Leu Asp Asp Lys Asp Glu
835 840 845
Ile Ala Leu Val Asn Ser Phe Asn Gly Phe Thr Thr Ala Phe Thr Gly
850 855 860
Phe Phe Asp Asn Arg Glu Asn Met Phe Ser Glu Glu Ala Lys Ser Thr
865 870 875 880
Ser Ile Ala Phe Arg Cys Ile Asn Glu Asn Leu Thr Arg Tyr Ile Ser
885 890 895
Asn Met Asp Ile Phe Glu Lys Val Asp Ala Ile Phe Asp Lys His Glu
900 905 910
Val Gln Glu Ile Lys Glu Lys Ile Leu Asn Ser Asp Tyr Asp Val Glu
915 920 925
Asp Phe Phe Glu Gly Glu Phe Phe Asn Phe Val Leu Thr Gln Glu Gly
930 935 940
Ile Asp Val Tyr Asn Ala Ile Ile Gly Gly Phe Val Thr Glu Ser Gly
945 950 955 960
Glu Lys Ile Lys Gly Leu Asn Glu Tyr Ile Asn Leu Tyr Asn Gln Lys
965 970 975
Thr Lys Gln Lys Leu Pro Lys Phe Lys Pro Leu Tyr Lys Gln Val Leu
980 985 990
Ser Asp Arg Glu Ser Leu Ser Phe Tyr Gly Glu Gly Tyr Thr Ser Asp
995 1000 1005
Glu Glu Val Leu Glu Val Phe Arg Asn Thr Leu Asn Lys Asn Ser
1010 1015 1020
Glu Ile Phe Ser Ser Ile Lys Lys Leu Glu Lys Leu Phe Lys Asn
1025 1030 1035
Phe Asp Glu Tyr Ser Ser Ala Gly Ile Phe Val Lys Asn Gly Pro
1040 1045 1050
Ala Ile Ser Thr Ile Ser Lys Asp Ile Phe Gly Glu Trp Asn Val
1055 1060 1065
Ile Arg Asp Lys Trp Asn Ala Glu Tyr Asp Asp Ile His Leu Lys
1070 1075 1080
Lys Lys Ala Val Val Thr Glu Lys Tyr Glu Asp Asp Arg Arg Lys
1085 1090 1095
Ser Phe Lys Lys Ile Gly Ser Phe Ser Leu Glu Gln Leu Gln Glu
1100 1105 1110
Tyr Ala Asp Ala Asp Leu Ser Val Val Glu Lys Leu Lys Glu Ile
1115 1120 1125
Ile Ile Gln Lys Val Asp Glu Ile Tyr Lys Val Tyr Gly Ser Ser
1130 1135 1140
Glu Lys Leu Phe Asp Ala Asp Phe Val Leu Glu Lys Ser Leu Lys
1145 1150 1155
Lys Asn Asp Ala Val Val Ala Ile Met Lys Asp Leu Leu Asp Ser
1160 1165 1170
Val Lys Ser Phe Glu Asn Tyr Ile Lys Ala Phe Phe Gly Glu Gly
1175 1180 1185
Lys Glu Thr Asn Arg Asp Glu Ser Phe Tyr Gly Asp Phe Val Leu
1190 1195 1200
Ala Tyr Asp Ile Leu Leu Lys Val Asp His Ile Tyr Asp Ala Ile
1205 1210 1215
Arg Asn Tyr Val Thr Gln Lys Pro Tyr Ser Lys Asp Lys Phe Lys
1220 1225 1230
Leu Tyr Phe Gln Asn Pro Gln Phe Met Arg Gly Trp Asp Lys Asp
1235 1240 1245
Lys Glu Thr Asp Tyr Arg Ala Thr Ile Leu Arg Tyr Gly Ser Lys
1250 1255 1260
Tyr Tyr Leu Ala Ile Met Asp Lys Lys Tyr Ala Lys Cys Leu Gln
1265 1270 1275
Lys Ile Asp Lys Asp Asp Val Asn Gly Asn Tyr Glu Lys Ile Asn
1280 1285 1290
Tyr Lys Leu Leu Pro Gly Pro Asn Lys Met Leu Pro Arg Val Phe
1295 1300 1305
Phe Ser Lys Lys Trp Met Ala Tyr Tyr Asn Pro Ser Glu Asp Ile
1310 1315 1320
Gln Lys Ile Tyr Lys Asn Gly Thr Phe Lys Lys Gly Asp Met Phe
1325 1330 1335
Asn Leu Asn Asp Cys His Lys Leu Ile Asp Phe Phe Lys Asp Ser
1340 1345 1350
Ile Ser Arg Tyr Pro Lys Trp Ser Asn Ala Tyr Asp Phe Asn Phe
1355 1360 1365
Ser Glu Thr Glu Lys Tyr Lys Asp Ile Ala Gly Phe Tyr Arg Glu
1370 1375 1380
Val Glu Glu Gln Gly Tyr Lys Val Ser Phe Glu Ser Ala Ser Lys
1385 1390 1395
Lys Glu Val Asp Lys Leu Val Glu Glu Gly Lys Leu Tyr Met Phe
1400 1405 1410
Gln Ile Tyr Asn Lys Asp Phe Ser Asp Lys Ser His Gly Thr Pro
1415 1420 1425
Asn Leu His Thr Met Tyr Phe Lys Leu Leu Phe Asp Glu Asn Asn
1430 1435 1440
His Gly Gln Ile Arg Leu Ser Gly Gly Ala Glu Leu Phe Met Arg
1445 1450 1455
Arg Ala Ser Leu Lys Lys Glu Glu Leu Val Val His Pro Ala Asn
1460 1465 1470
Ser Pro Ile Ala Asn Lys Asn Pro Asp Asn Pro Lys Lys Thr Thr
1475 1480 1485
Thr Leu Ser Tyr Asp Val Tyr Lys Asp Lys Arg Phe Ser Glu Asp
1490 1495 1500
Gln Tyr Glu Leu His Ile Pro Ile Ala Ile Asn Lys Cys Pro Lys
1505 1510 1515
Asn Ile Phe Lys Ile Asn Thr Glu Val Arg Val Leu Leu Lys His
1520 1525 1530
Asp Asp Asn Pro Tyr Val Ile Gly Ile Asp Arg Gly Glu Arg Asn
1535 1540 1545
Leu Leu Tyr Ile Val Val Val Asp Gly Lys Gly Asn Ile Val Glu
1550 1555 1560
Gln Tyr Ser Leu Asn Glu Ile Ile Asn Asn Phe Asn Gly Ile Arg
1565 1570 1575
Ile Lys Thr Asp Tyr His Ser Leu Leu Asp Lys Lys Glu Lys Glu
1580 1585 1590
Arg Phe Glu Ala Arg Gln Asn Trp Thr Ser Ile Glu Asn Ile Lys
1595 1600 1605
Glu Leu Lys Ala Gly Tyr Ile Ser Gln Val Val His Lys Ile Cys
1610 1615 1620
Glu Leu Val Glu Lys Tyr Asp Ala Val Ile Ala Leu Glu Asp Leu
1625 1630 1635
Asn Ser Gly Phe Lys Asn Ser Arg Val Lys Val Glu Lys Gln Val
1640 1645 1650
Tyr Gln Lys Phe Glu Lys Met Leu Ile Asp Lys Leu Asn Tyr Met
1655 1660 1665
Val Asp Lys Lys Ser Asn Pro Cys Ala Thr Gly Gly Ala Leu Lys
1670 1675 1680
Gly Tyr Gln Ile Thr Asn Lys Phe Glu Ser Phe Lys Ser Met Ser
1685 1690 1695
Thr Gln Asn Gly Phe Ile Phe Tyr Ile Pro Ala Trp Leu Thr Ser
1700 1705 1710
Lys Ile Asp Pro Ser Thr Gly Phe Val Asn Leu Leu Lys Thr Lys
1715 1720 1725
Tyr Thr Ser Ile Ala Asp Ser Lys Lys Phe Ile Ser Ser Phe Asp
1730 1735 1740
Arg Ile Met Tyr Val Pro Glu Glu Asp Leu Phe Glu Phe Ala Leu
1745 1750 1755
Asp Tyr Lys Asn Phe Ser Arg Thr Asp Ala Asp Tyr Ile Lys Lys
1760 1765 1770
Trp Lys Leu Tyr Ser Tyr Gly Asn Arg Ile Arg Ile Phe Arg Asn
1775 1780 1785
Pro Lys Lys Asn Asn Val Phe Asp Trp Glu Glu Val Cys Leu Thr
1790 1795 1800
Ser Ala Tyr Lys Glu Leu Phe Asn Lys Tyr Gly Ile Asn Tyr Gln
1805 1810 1815
Gln Gly Asp Ile Arg Ala Leu Leu Cys Glu Gln Ser Asp Lys Ala
1820 1825 1830
Phe Tyr Ser Ser Phe Met Ala Leu Met Ser Leu Met Leu Gln Met
1835 1840 1845
Ala Asn Ser Ile Thr Gly Arg Thr Asp Val Asp Phe Leu Ile Ser
1850 1855 1860
Pro Val Lys Asn Ser Asp Gly Ile Phe Tyr Asp Ser Arg Asn Tyr
1865 1870 1875
Glu Ala Gln Glu Asn Ala Ile Leu Pro Lys Asn Ala Asp Ala Asn
1880 1885 1890
Gly Ala Tyr Asn Ile Ala Arg Lys Val Leu Trp Ala Ile Gly Gln
1895 1900 1905
Phe Lys Lys Ala Glu Asp Glu Lys Leu Asp Lys Val Lys Ile Ala
1910 1915 1920
Ile Ser Asn Lys Glu Trp Leu Glu Tyr Ala Gln Thr Ser Val Lys
1925 1930 1935
His Ser Gly Gly Ser Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu
1940 1945 1950
Pro Lys Lys Lys Arg Lys Val Gly Ser Gly
1955 1960
<210> 64
<211> 1979
<212> PRT
<213> 人工的
<220>
<223> 合成的REDRAW编辑器
<400> 64
Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys
1 5 10 15
Arg Lys Val Thr Leu Asn Ile Glu Asp Glu Tyr Arg Leu His Glu Thr
20 25 30
Ser Lys Glu Pro Asp Val Ser Leu Gly Ser Thr Trp Leu Ser Asp Phe
35 40 45
Pro Gln Ala Trp Ala Glu Thr Gly Gly Met Gly Leu Ala Val Arg Gln
50 55 60
Ala Pro Leu Ile Ile Pro Leu Lys Ala Thr Ser Thr Pro Val Ser Ile
65 70 75 80
Lys Gln Tyr Pro Met Ser Gln Glu Ala Arg Leu Gly Ile Lys Pro His
85 90 95
Ile Gln Arg Leu Leu Asp Gln Gly Ile Leu Val Pro Cys Gln Ser Pro
100 105 110
Trp Asn Thr Pro Leu Leu Pro Val Lys Lys Pro Gly Thr Asn Asp Tyr
115 120 125
Arg Pro Val Gln Asp Leu Arg Glu Val Asn Lys Arg Val Glu Asp Ile
130 135 140
His Pro Thr Val Pro Asn Pro Tyr Asn Leu Leu Ser Gly Leu Pro Pro
145 150 155 160
Ser His Gln Trp Tyr Thr Val Leu Asp Leu Lys Asp Ala Phe Phe Cys
165 170 175
Leu Arg Leu His Pro Thr Ser Gln Pro Leu Phe Ala Phe Glu Trp Arg
180 185 190
Asp Pro Glu Met Gly Ile Ser Gly Gln Leu Thr Trp Thr Arg Leu Pro
195 200 205
Gln Gly Phe Lys Asn Ser Pro Thr Leu Phe Asn Glu Ala Leu His Arg
210 215 220
Asp Leu Ala Asp Phe Arg Ile Gln His Pro Asp Leu Ile Leu Leu Gln
225 230 235 240
Tyr Val Asp Asp Leu Leu Leu Ala Ala Thr Ser Glu Leu Asp Cys Gln
245 250 255
Gln Gly Thr Arg Ala Leu Leu Gln Thr Leu Gly Asn Leu Gly Tyr Arg
260 265 270
Ala Ser Ala Lys Lys Ala Gln Ile Cys Gln Lys Gln Val Lys Tyr Leu
275 280 285
Gly Tyr Leu Leu Lys Glu Gly Gln Arg Trp Leu Thr Glu Ala Arg Lys
290 295 300
Glu Thr Val Met Gly Gln Pro Thr Pro Lys Thr Pro Arg Gln Leu Arg
305 310 315 320
Glu Phe Leu Gly Lys Ala Gly Phe Cys Arg Leu Phe Ile Pro Gly Phe
325 330 335
Ala Glu Met Ala Ala Pro Leu Tyr Pro Leu Thr Lys Pro Gly Thr Leu
340 345 350
Phe Asn Trp Gly Pro Asp Gln Gln Lys Ala Tyr Gln Glu Ile Lys Gln
355 360 365
Ala Leu Leu Thr Ala Pro Ala Leu Gly Leu Pro Asp Leu Thr Lys Pro
370 375 380
Phe Glu Leu Phe Val Asp Glu Lys Gln Gly Tyr Ala Lys Gly Val Leu
385 390 395 400
Thr Gln Lys Leu Gly Pro Trp Arg Arg Pro Val Ala Tyr Leu Ser Lys
405 410 415
Lys Leu Asp Pro Val Ala Ala Gly Trp Pro Pro Cys Leu Arg Met Val
420 425 430
Ala Ala Ile Ala Val Leu Thr Lys Asp Ala Gly Lys Leu Thr Met Gly
435 440 445
Gln Pro Leu Val Ile Leu Ala Pro His Ala Val Glu Ala Leu Val Lys
450 455 460
Gln Pro Pro Asp Arg Trp Leu Ser Asn Ala Arg Met Thr His Tyr Gln
465 470 475 480
Ala Leu Leu Leu Asp Thr Asp Arg Val Gln Phe Gly Pro Val Val Ala
485 490 495
Leu Asn Pro Ala Thr Leu Leu Pro Leu Pro Glu Glu Gly Leu Gln His
500 505 510
Asn Cys Leu Asp Ile Leu Ala Glu Ala His Gly Thr Arg Pro Asp Leu
515 520 525
Thr Asp Gln Pro Leu Pro Asp Ala Asp His Thr Trp Tyr Thr Asp Gly
530 535 540
Ser Ser Leu Leu Gln Glu Gly Gln Arg Lys Ala Gly Ala Ala Val Thr
545 550 555 560
Thr Glu Thr Glu Val Ile Trp Ala Lys Ala Leu Pro Ala Gly Thr Ser
565 570 575
Ala Gln Arg Ala Glu Leu Ile Ala Leu Thr Gln Ala Leu Lys Met Ala
580 585 590
Glu Gly Lys Lys Leu Asn Val Tyr Thr Asp Ser Arg Tyr Ala Phe Ala
595 600 605
Thr Ala His Ile His Gly Glu Ile Tyr Arg Arg Arg Gly Trp Leu Thr
610 615 620
Ser Glu Gly Lys Glu Ile Lys Asn Lys Asp Glu Ile Leu Ala Leu Leu
625 630 635 640
Lys Ala Leu Phe Leu Pro Lys Arg Leu Ser Ile Ile His Cys Pro Gly
645 650 655
His Gln Lys Gly His Ser Ala Glu Ala Arg Gly Asn Arg Met Ala Asp
660 665 670
Gln Ala Ala Arg Lys Ala Ala Ile Thr Glu Thr Pro Asp Thr Ser Thr
675 680 685
Leu Leu Ile Glu Asn Ser Ser Pro Ser Gly Gly Ser Ser Gly Gly Ser
690 695 700
Ser Gly Ser Glu Thr Pro Gly Thr Ser Glu Ser Ala Thr Pro Glu Ser
705 710 715 720
Ser Gly Gly Ser Ser Gly Gly Ser Ser Lys Leu Glu Lys Phe Thr Asn
725 730 735
Cys Tyr Ser Leu Ser Lys Thr Leu Arg Phe Lys Ala Ile Pro Val Gly
740 745 750
Lys Thr Gln Glu Asn Ile Asp Asn Lys Arg Leu Leu Val Glu Asp Glu
755 760 765
Lys Arg Ala Glu Asp Tyr Lys Gly Val Lys Lys Leu Leu Asp Arg Tyr
770 775 780
Tyr Leu Ser Phe Ile Asn Asp Val Leu His Ser Ile Lys Leu Lys Asn
785 790 795 800
Leu Asn Asn Tyr Ile Ser Leu Phe Arg Lys Lys Thr Arg Thr Glu Lys
805 810 815
Glu Asn Lys Glu Leu Glu Asn Leu Glu Ile Asn Leu Arg Lys Glu Ile
820 825 830
Ala Lys Ala Phe Lys Gly Asn Glu Gly Tyr Lys Ser Leu Phe Lys Lys
835 840 845
Asp Ile Ile Glu Thr Ile Leu Pro Glu Phe Leu Asp Asp Lys Asp Glu
850 855 860
Ile Ala Leu Val Asn Ser Phe Asn Gly Phe Thr Thr Ala Phe Thr Gly
865 870 875 880
Phe Phe Asp Asn Arg Glu Asn Met Phe Ser Glu Glu Ala Lys Ser Thr
885 890 895
Ser Ile Ala Phe Arg Cys Ile Asn Glu Asn Leu Thr Arg Tyr Ile Ser
900 905 910
Asn Met Asp Ile Phe Glu Lys Val Asp Ala Ile Phe Asp Lys His Glu
915 920 925
Val Gln Glu Ile Lys Glu Lys Ile Leu Asn Ser Asp Tyr Asp Val Glu
930 935 940
Asp Phe Phe Glu Gly Glu Phe Phe Asn Phe Val Leu Thr Gln Glu Gly
945 950 955 960
Ile Asp Val Tyr Asn Ala Ile Ile Gly Gly Phe Val Thr Glu Ser Gly
965 970 975
Glu Lys Ile Lys Gly Leu Asn Glu Tyr Ile Asn Leu Tyr Asn Gln Lys
980 985 990
Thr Lys Gln Lys Leu Pro Lys Phe Lys Pro Leu Tyr Lys Gln Val Leu
995 1000 1005
Ser Asp Arg Glu Ser Leu Ser Phe Tyr Gly Glu Gly Tyr Thr Ser
1010 1015 1020
Asp Glu Glu Val Leu Glu Val Phe Arg Asn Thr Leu Asn Lys Asn
1025 1030 1035
Ser Glu Ile Phe Ser Ser Ile Lys Lys Leu Glu Lys Leu Phe Lys
1040 1045 1050
Asn Phe Asp Glu Tyr Ser Ser Ala Gly Ile Phe Val Lys Asn Gly
1055 1060 1065
Pro Ala Ile Ser Thr Ile Ser Lys Asp Ile Phe Gly Glu Trp Asn
1070 1075 1080
Val Ile Arg Asp Lys Trp Asn Ala Glu Tyr Asp Asp Ile His Leu
1085 1090 1095
Lys Lys Lys Ala Val Val Thr Glu Lys Tyr Glu Asp Asp Arg Arg
1100 1105 1110
Lys Ser Phe Lys Lys Ile Gly Ser Phe Ser Leu Glu Gln Leu Gln
1115 1120 1125
Glu Tyr Ala Asp Ala Asp Leu Ser Val Val Glu Lys Leu Lys Glu
1130 1135 1140
Ile Ile Ile Gln Lys Val Asp Glu Ile Tyr Lys Val Tyr Gly Ser
1145 1150 1155
Ser Glu Lys Leu Phe Asp Ala Asp Phe Val Leu Glu Lys Ser Leu
1160 1165 1170
Lys Lys Asn Asp Ala Val Val Ala Ile Met Lys Asp Leu Leu Asp
1175 1180 1185
Ser Val Lys Ser Phe Glu Asn Tyr Ile Lys Ala Phe Phe Gly Glu
1190 1195 1200
Gly Lys Glu Thr Asn Arg Asp Glu Ser Phe Tyr Gly Asp Phe Val
1205 1210 1215
Leu Ala Tyr Asp Ile Leu Leu Lys Val Asp His Ile Tyr Asp Ala
1220 1225 1230
Ile Arg Asn Tyr Val Thr Gln Lys Pro Tyr Ser Lys Asp Lys Phe
1235 1240 1245
Lys Leu Tyr Phe Gln Asn Pro Gln Phe Met Arg Gly Trp Asp Lys
1250 1255 1260
Asp Lys Glu Thr Asp Tyr Arg Ala Thr Ile Leu Arg Tyr Gly Ser
1265 1270 1275
Lys Tyr Tyr Leu Ala Ile Met Asp Lys Lys Tyr Ala Lys Cys Leu
1280 1285 1290
Gln Lys Ile Asp Lys Asp Asp Val Asn Gly Asn Tyr Glu Lys Ile
1295 1300 1305
Asn Tyr Lys Leu Leu Pro Gly Pro Asn Lys Met Leu Pro Arg Val
1310 1315 1320
Phe Phe Ser Lys Lys Trp Met Ala Tyr Tyr Asn Pro Ser Glu Asp
1325 1330 1335
Ile Gln Lys Ile Tyr Lys Asn Gly Thr Phe Lys Lys Gly Asp Met
1340 1345 1350
Phe Asn Leu Asn Asp Cys His Lys Leu Ile Asp Phe Phe Lys Asp
1355 1360 1365
Ser Ile Ser Arg Tyr Pro Lys Trp Ser Asn Ala Tyr Asp Phe Asn
1370 1375 1380
Phe Ser Glu Thr Glu Lys Tyr Lys Asp Ile Ala Gly Phe Tyr Arg
1385 1390 1395
Glu Val Glu Glu Gln Gly Tyr Lys Val Ser Phe Glu Ser Ala Ser
1400 1405 1410
Lys Lys Glu Val Asp Lys Leu Val Glu Glu Gly Lys Leu Tyr Met
1415 1420 1425
Phe Gln Ile Tyr Asn Lys Asp Phe Ser Asp Lys Ser His Gly Thr
1430 1435 1440
Pro Asn Leu His Thr Met Tyr Phe Lys Leu Leu Phe Asp Glu Asn
1445 1450 1455
Asn His Gly Gln Ile Arg Leu Ser Gly Gly Ala Glu Leu Phe Met
1460 1465 1470
Arg Arg Ala Ser Leu Lys Lys Glu Glu Leu Val Val His Pro Ala
1475 1480 1485
Asn Ser Pro Ile Ala Asn Lys Asn Pro Asp Asn Pro Lys Lys Thr
1490 1495 1500
Thr Thr Leu Ser Tyr Asp Val Tyr Lys Asp Lys Arg Phe Ser Glu
1505 1510 1515
Asp Gln Tyr Glu Leu His Ile Pro Ile Ala Ile Asn Lys Cys Pro
1520 1525 1530
Lys Asn Ile Phe Lys Ile Asn Thr Glu Val Arg Val Leu Leu Lys
1535 1540 1545
His Asp Asp Asn Pro Tyr Val Ile Gly Ile Asp Arg Gly Glu Arg
1550 1555 1560
Asn Leu Leu Tyr Ile Val Val Val Asp Gly Lys Gly Asn Ile Val
1565 1570 1575
Glu Gln Tyr Ser Leu Asn Glu Ile Ile Asn Asn Phe Asn Gly Ile
1580 1585 1590
Arg Ile Lys Thr Asp Tyr His Ser Leu Leu Asp Lys Lys Glu Lys
1595 1600 1605
Glu Arg Phe Glu Ala Arg Gln Asn Trp Thr Ser Ile Glu Asn Ile
1610 1615 1620
Lys Glu Leu Lys Ala Gly Tyr Ile Ser Gln Val Val His Lys Ile
1625 1630 1635
Cys Glu Leu Val Glu Lys Tyr Asp Ala Val Ile Ala Leu Glu Asp
1640 1645 1650
Leu Asn Ser Gly Phe Lys Asn Ser Arg Val Lys Val Glu Lys Gln
1655 1660 1665
Val Tyr Gln Lys Phe Glu Lys Met Leu Ile Asp Lys Leu Asn Tyr
1670 1675 1680
Met Val Asp Lys Lys Ser Asn Pro Cys Ala Thr Gly Gly Ala Leu
1685 1690 1695
Lys Gly Tyr Gln Ile Thr Asn Lys Phe Glu Ser Phe Lys Ser Met
1700 1705 1710
Ser Thr Gln Asn Gly Phe Ile Phe Tyr Ile Pro Ala Trp Leu Thr
1715 1720 1725
Ser Lys Ile Asp Pro Ser Thr Gly Phe Val Asn Leu Leu Lys Thr
1730 1735 1740
Lys Tyr Thr Ser Ile Ala Asp Ser Lys Lys Phe Ile Ser Ser Phe
1745 1750 1755
Asp Arg Ile Met Tyr Val Pro Glu Glu Asp Leu Phe Glu Phe Ala
1760 1765 1770
Leu Asp Tyr Lys Asn Phe Ser Arg Thr Asp Ala Asp Tyr Ile Lys
1775 1780 1785
Lys Trp Lys Leu Tyr Ser Tyr Gly Asn Arg Ile Arg Ile Phe Arg
1790 1795 1800
Asn Pro Lys Lys Asn Asn Val Phe Asp Trp Glu Glu Val Cys Leu
1805 1810 1815
Thr Ser Ala Tyr Lys Glu Leu Phe Asn Lys Tyr Gly Ile Asn Tyr
1820 1825 1830
Gln Gln Gly Asp Ile Arg Ala Leu Leu Cys Glu Gln Ser Asp Lys
1835 1840 1845
Ala Phe Tyr Ser Ser Phe Met Ala Leu Met Ser Leu Met Leu Gln
1850 1855 1860
Met Ala Asn Ser Ile Thr Gly Arg Thr Asp Val Asp Phe Leu Ile
1865 1870 1875
Ser Pro Val Lys Asn Ser Asp Gly Ile Phe Tyr Asp Ser Arg Asn
1880 1885 1890
Tyr Glu Ala Gln Glu Asn Ala Ile Leu Pro Lys Asn Ala Asp Ala
1895 1900 1905
Asn Gly Ala Tyr Asn Ile Ala Arg Lys Val Leu Trp Ala Ile Gly
1910 1915 1920
Gln Phe Lys Lys Ala Glu Asp Glu Lys Leu Asp Lys Val Lys Ile
1925 1930 1935
Ala Ile Ser Asn Lys Glu Trp Leu Glu Tyr Ala Gln Thr Ser Val
1940 1945 1950
Lys His Ser Gly Gly Ser Lys Arg Thr Ala Asp Gly Ser Glu Phe
1955 1960 1965
Glu Pro Lys Lys Lys Arg Lys Val Gly Ser Gly
1970 1975
<210> 65
<211> 1963
<212> PRT
<213> 人工的
<220>
<223> 合成的REDRAW编辑器
<400> 65
Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys
1 5 10 15
Arg Lys Val Ser Lys Leu Glu Lys Phe Thr Asn Cys Tyr Ser Leu Ser
20 25 30
Lys Thr Leu Arg Phe Lys Ala Ile Pro Val Gly Lys Thr Gln Glu Asn
35 40 45
Ile Asp Asn Lys Arg Leu Leu Val Glu Asp Glu Lys Arg Ala Glu Asp
50 55 60
Tyr Lys Gly Val Lys Lys Leu Leu Asp Arg Tyr Tyr Leu Ser Phe Ile
65 70 75 80
Asn Asp Val Leu His Ser Ile Lys Leu Lys Asn Leu Asn Asn Tyr Ile
85 90 95
Ser Leu Phe Arg Lys Lys Thr Arg Thr Glu Lys Glu Asn Lys Glu Leu
100 105 110
Glu Asn Leu Glu Ile Asn Leu Arg Lys Glu Ile Ala Lys Ala Phe Lys
115 120 125
Gly Asn Glu Gly Tyr Lys Ser Leu Phe Lys Lys Asp Ile Ile Glu Thr
130 135 140
Ile Leu Pro Glu Phe Leu Asp Asp Lys Asp Glu Ile Ala Leu Val Asn
145 150 155 160
Ser Phe Asn Gly Phe Thr Thr Ala Phe Thr Gly Phe Phe Asp Asn Arg
165 170 175
Glu Asn Met Phe Ser Glu Glu Ala Lys Ser Thr Ser Ile Ala Phe Arg
180 185 190
Cys Ile Asn Glu Asn Leu Thr Arg Tyr Ile Ser Asn Met Asp Ile Phe
195 200 205
Glu Lys Val Asp Ala Ile Phe Asp Lys His Glu Val Gln Glu Ile Lys
210 215 220
Glu Lys Ile Leu Asn Ser Asp Tyr Asp Val Glu Asp Phe Phe Glu Gly
225 230 235 240
Glu Phe Phe Asn Phe Val Leu Thr Gln Glu Gly Ile Asp Val Tyr Asn
245 250 255
Ala Ile Ile Gly Gly Phe Val Thr Glu Ser Gly Glu Lys Ile Lys Gly
260 265 270
Leu Asn Glu Tyr Ile Asn Leu Tyr Asn Gln Lys Thr Lys Gln Lys Leu
275 280 285
Pro Lys Phe Lys Pro Leu Tyr Lys Gln Val Leu Ser Asp Arg Glu Ser
290 295 300
Leu Ser Phe Tyr Gly Glu Gly Tyr Thr Ser Asp Glu Glu Val Leu Glu
305 310 315 320
Val Phe Arg Asn Thr Leu Asn Lys Asn Ser Glu Ile Phe Ser Ser Ile
325 330 335
Lys Lys Leu Glu Lys Leu Phe Lys Asn Phe Asp Glu Tyr Ser Ser Ala
340 345 350
Gly Ile Phe Val Lys Asn Gly Pro Ala Ile Ser Thr Ile Ser Lys Asp
355 360 365
Ile Phe Gly Glu Trp Asn Val Ile Arg Asp Lys Trp Asn Ala Glu Tyr
370 375 380
Asp Asp Ile His Leu Lys Lys Lys Ala Val Val Thr Glu Lys Tyr Glu
385 390 395 400
Asp Asp Arg Arg Lys Ser Phe Lys Lys Ile Gly Ser Phe Ser Leu Glu
405 410 415
Gln Leu Gln Glu Tyr Ala Asp Ala Asp Leu Ser Val Val Glu Lys Leu
420 425 430
Lys Glu Ile Ile Ile Gln Lys Val Asp Glu Ile Tyr Lys Val Tyr Gly
435 440 445
Ser Ser Glu Lys Leu Phe Asp Ala Asp Phe Val Leu Glu Lys Ser Leu
450 455 460
Lys Lys Asn Asp Ala Val Val Ala Ile Met Lys Asp Leu Leu Asp Ser
465 470 475 480
Val Lys Ser Phe Glu Asn Tyr Ile Lys Ala Phe Phe Gly Glu Gly Lys
485 490 495
Glu Thr Asn Arg Asp Glu Ser Phe Tyr Gly Asp Phe Val Leu Ala Tyr
500 505 510
Asp Ile Leu Leu Lys Val Asp His Ile Tyr Asp Ala Ile Arg Asn Tyr
515 520 525
Val Thr Gln Lys Pro Tyr Ser Lys Asp Lys Phe Lys Leu Tyr Phe Gln
530 535 540
Asn Pro Gln Phe Met Arg Gly Trp Asp Lys Asp Lys Glu Thr Asp Tyr
545 550 555 560
Arg Ala Thr Ile Leu Arg Tyr Gly Ser Lys Tyr Tyr Leu Ala Ile Met
565 570 575
Asp Lys Lys Tyr Ala Lys Cys Leu Gln Lys Ile Asp Lys Asp Asp Val
580 585 590
Asn Gly Asn Tyr Glu Lys Ile Asn Tyr Lys Leu Leu Pro Gly Pro Asn
595 600 605
Lys Met Leu Pro Arg Val Phe Phe Ser Lys Lys Trp Met Ala Tyr Tyr
610 615 620
Asn Pro Ser Glu Asp Ile Gln Lys Ile Tyr Lys Asn Gly Thr Phe Lys
625 630 635 640
Lys Gly Asp Met Phe Asn Leu Asn Asp Cys His Lys Leu Ile Asp Phe
645 650 655
Phe Lys Asp Ser Ile Ser Arg Tyr Pro Lys Trp Ser Asn Ala Tyr Asp
660 665 670
Phe Asn Phe Ser Glu Thr Glu Lys Tyr Lys Asp Ile Ala Gly Phe Tyr
675 680 685
Arg Glu Val Glu Glu Gln Gly Tyr Lys Val Ser Phe Glu Ser Ala Ser
690 695 700
Lys Lys Glu Val Asp Lys Leu Val Glu Glu Gly Lys Leu Tyr Met Phe
705 710 715 720
Gln Ile Tyr Asn Lys Asp Phe Ser Asp Lys Ser His Gly Thr Pro Asn
725 730 735
Leu His Thr Met Tyr Phe Lys Leu Leu Phe Asp Glu Asn Asn His Gly
740 745 750
Gln Ile Arg Leu Ser Gly Gly Ala Glu Leu Phe Met Arg Arg Ala Ser
755 760 765
Leu Lys Lys Glu Glu Leu Val Val His Pro Ala Asn Ser Pro Ile Ala
770 775 780
Asn Lys Asn Pro Asp Asn Pro Lys Lys Thr Thr Thr Leu Ser Tyr Asp
785 790 795 800
Val Tyr Lys Asp Lys Arg Phe Ser Glu Asp Gln Tyr Glu Leu His Ile
805 810 815
Pro Ile Ala Ile Asn Lys Cys Pro Lys Asn Ile Phe Lys Ile Asn Thr
820 825 830
Glu Val Arg Val Leu Leu Lys His Asp Asp Asn Pro Tyr Val Ile Gly
835 840 845
Ile Asp Arg Gly Glu Arg Asn Leu Leu Tyr Ile Val Val Val Asp Gly
850 855 860
Lys Gly Asn Ile Val Glu Gln Tyr Ser Leu Asn Glu Ile Ile Asn Asn
865 870 875 880
Phe Asn Gly Ile Arg Ile Lys Thr Asp Tyr His Ser Leu Leu Asp Lys
885 890 895
Lys Glu Lys Glu Arg Phe Glu Ala Arg Gln Asn Trp Thr Ser Ile Glu
900 905 910
Asn Ile Lys Glu Leu Lys Ala Gly Tyr Ile Ser Gln Val Val His Lys
915 920 925
Ile Cys Glu Leu Val Glu Lys Tyr Asp Ala Val Ile Ala Leu Glu Asp
930 935 940
Leu Asn Ser Gly Phe Lys Asn Ser Arg Val Lys Val Glu Lys Gln Val
945 950 955 960
Tyr Gln Lys Phe Glu Lys Met Leu Ile Asp Lys Leu Asn Tyr Met Val
965 970 975
Asp Lys Lys Ser Asn Pro Cys Ala Thr Gly Gly Ala Leu Lys Gly Tyr
980 985 990
Gln Ile Thr Asn Lys Phe Glu Ser Phe Lys Ser Met Ser Thr Gln Asn
995 1000 1005
Gly Phe Ile Phe Tyr Ile Pro Ala Trp Leu Thr Ser Lys Ile Asp
1010 1015 1020
Pro Ser Thr Gly Phe Val Asn Leu Leu Lys Thr Lys Tyr Thr Ser
1025 1030 1035
Ile Ala Asp Ser Lys Lys Phe Ile Ser Ser Phe Asp Arg Ile Met
1040 1045 1050
Tyr Val Pro Glu Glu Asp Leu Phe Glu Phe Ala Leu Asp Tyr Lys
1055 1060 1065
Asn Phe Ser Arg Thr Asp Ala Asp Tyr Ile Lys Lys Trp Lys Leu
1070 1075 1080
Tyr Ser Tyr Gly Asn Arg Ile Arg Ile Phe Arg Asn Pro Lys Lys
1085 1090 1095
Asn Asn Val Phe Asp Trp Glu Glu Val Cys Leu Thr Ser Ala Tyr
1100 1105 1110
Lys Glu Leu Phe Asn Lys Tyr Gly Ile Asn Tyr Gln Gln Gly Asp
1115 1120 1125
Ile Arg Ala Leu Leu Cys Glu Gln Ser Asp Lys Ala Phe Tyr Ser
1130 1135 1140
Ser Phe Met Ala Leu Met Ser Leu Met Leu Gln Met Ala Asn Ser
1145 1150 1155
Ile Thr Gly Arg Thr Asp Val Asp Phe Leu Ile Ser Pro Val Lys
1160 1165 1170
Asn Ser Asp Gly Ile Phe Tyr Asp Ser Arg Asn Tyr Glu Ala Gln
1175 1180 1185
Glu Asn Ala Ile Leu Pro Lys Asn Ala Asp Ala Asn Gly Ala Tyr
1190 1195 1200
Asn Ile Ala Arg Lys Val Leu Trp Ala Ile Gly Gln Phe Lys Lys
1205 1210 1215
Ala Glu Asp Glu Lys Leu Asp Lys Val Lys Ile Ala Ile Ser Asn
1220 1225 1230
Lys Glu Trp Leu Glu Tyr Ala Gln Thr Ser Val Lys His Ser Gly
1235 1240 1245
Ser Glu Thr Pro Gly Thr Ser Glu Ser Ala Thr Pro Glu Ser Thr
1250 1255 1260
Leu Asn Ile Glu Asp Glu Tyr Arg Leu His Glu Thr Ser Lys Glu
1265 1270 1275
Pro Asp Val Ser Leu Gly Ser Thr Trp Leu Ser Asp Phe Pro Gln
1280 1285 1290
Ala Trp Ala Glu Thr Gly Gly Met Gly Leu Ala Val Arg Gln Ala
1295 1300 1305
Pro Leu Ile Ile Pro Leu Lys Ala Thr Ser Thr Pro Val Ser Ile
1310 1315 1320
Lys Gln Tyr Pro Met Ser Gln Glu Ala Arg Leu Gly Ile Lys Pro
1325 1330 1335
His Ile Gln Arg Leu Leu Asp Gln Gly Ile Leu Val Pro Cys Gln
1340 1345 1350
Ser Pro Trp Asn Thr Pro Leu Leu Pro Val Lys Lys Pro Gly Thr
1355 1360 1365
Asn Asp Tyr Arg Pro Val Gln Asp Leu Arg Glu Val Asn Lys Arg
1370 1375 1380
Val Glu Asp Ile His Pro Thr Val Pro Asn Pro Tyr Asn Leu Leu
1385 1390 1395
Ser Gly Leu Pro Pro Ser His Gln Trp Tyr Thr Val Leu Asp Leu
1400 1405 1410
Lys Asp Ala Phe Phe Cys Leu Arg Leu His Pro Thr Ser Gln Pro
1415 1420 1425
Leu Phe Ala Phe Glu Trp Arg Asp Pro Glu Met Gly Ile Ser Gly
1430 1435 1440
Gln Leu Thr Trp Thr Arg Leu Pro Gln Gly Phe Lys Asn Ser Pro
1445 1450 1455
Thr Leu Phe Asn Glu Ala Leu His Arg Asp Leu Ala Asp Phe Arg
1460 1465 1470
Ile Gln His Pro Asp Leu Ile Leu Leu Gln Tyr Val Asp Asp Leu
1475 1480 1485
Leu Leu Ala Ala Thr Ser Glu Leu Asp Cys Gln Gln Gly Thr Arg
1490 1495 1500
Ala Leu Leu Gln Thr Leu Gly Asn Leu Gly Tyr Arg Ala Ser Ala
1505 1510 1515
Lys Lys Ala Gln Ile Cys Gln Lys Gln Val Lys Tyr Leu Gly Tyr
1520 1525 1530
Leu Leu Lys Glu Gly Gln Arg Trp Leu Thr Glu Ala Arg Lys Glu
1535 1540 1545
Thr Val Met Gly Gln Pro Thr Pro Lys Thr Pro Arg Gln Leu Arg
1550 1555 1560
Glu Phe Leu Gly Lys Ala Gly Phe Cys Arg Leu Phe Ile Pro Gly
1565 1570 1575
Phe Ala Glu Met Ala Ala Pro Leu Tyr Pro Leu Thr Lys Pro Gly
1580 1585 1590
Thr Leu Phe Asn Trp Gly Pro Asp Gln Gln Lys Ala Tyr Gln Glu
1595 1600 1605
Ile Lys Gln Ala Leu Leu Thr Ala Pro Ala Leu Gly Leu Pro Asp
1610 1615 1620
Leu Thr Lys Pro Phe Glu Leu Phe Val Asp Glu Lys Gln Gly Tyr
1625 1630 1635
Ala Lys Gly Val Leu Thr Gln Lys Leu Gly Pro Trp Arg Arg Pro
1640 1645 1650
Val Ala Tyr Leu Ser Lys Lys Leu Asp Pro Val Ala Ala Gly Trp
1655 1660 1665
Pro Pro Cys Leu Arg Met Val Ala Ala Ile Ala Val Leu Thr Lys
1670 1675 1680
Asp Ala Gly Lys Leu Thr Met Gly Gln Pro Leu Val Ile Leu Ala
1685 1690 1695
Pro His Ala Val Glu Ala Leu Val Lys Gln Pro Pro Asp Arg Trp
1700 1705 1710
Leu Ser Asn Ala Arg Met Thr His Tyr Gln Ala Leu Leu Leu Asp
1715 1720 1725
Thr Asp Arg Val Gln Phe Gly Pro Val Val Ala Leu Asn Pro Ala
1730 1735 1740
Thr Leu Leu Pro Leu Pro Glu Glu Gly Leu Gln His Asn Cys Leu
1745 1750 1755
Asp Ile Leu Ala Glu Ala His Gly Thr Arg Pro Asp Leu Thr Asp
1760 1765 1770
Gln Pro Leu Pro Asp Ala Asp His Thr Trp Tyr Thr Asp Gly Ser
1775 1780 1785
Ser Leu Leu Gln Glu Gly Gln Arg Lys Ala Gly Ala Ala Val Thr
1790 1795 1800
Thr Glu Thr Glu Val Ile Trp Ala Lys Ala Leu Pro Ala Gly Thr
1805 1810 1815
Ser Ala Gln Arg Ala Glu Leu Ile Ala Leu Thr Gln Ala Leu Lys
1820 1825 1830
Met Ala Glu Gly Lys Lys Leu Asn Val Tyr Thr Asp Ser Arg Tyr
1835 1840 1845
Ala Phe Ala Thr Ala His Ile His Gly Glu Ile Tyr Arg Arg Arg
1850 1855 1860
Gly Trp Leu Thr Ser Glu Gly Lys Glu Ile Lys Asn Lys Asp Glu
1865 1870 1875
Ile Leu Ala Leu Leu Lys Ala Leu Phe Leu Pro Lys Arg Leu Ser
1880 1885 1890
Ile Ile His Cys Pro Gly His Gln Lys Gly His Ser Ala Glu Ala
1895 1900 1905
Arg Gly Asn Arg Met Ala Asp Gln Ala Ala Arg Lys Ala Ala Ile
1910 1915 1920
Thr Glu Thr Pro Asp Thr Ser Thr Leu Leu Ile Glu Asn Ser Ser
1925 1930 1935
Pro Ser Gly Gly Ser Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu
1940 1945 1950
Pro Lys Lys Lys Arg Lys Val Gly Ser Gly
1955 1960
<210> 66
<211> 1979
<212> PRT
<213> 人工的
<220>
<223> 合成的REDRAW编辑器
<400> 66
Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys
1 5 10 15
Arg Lys Val Ser Lys Leu Glu Lys Phe Thr Asn Cys Tyr Ser Leu Ser
20 25 30
Lys Thr Leu Arg Phe Lys Ala Ile Pro Val Gly Lys Thr Gln Glu Asn
35 40 45
Ile Asp Asn Lys Arg Leu Leu Val Glu Asp Glu Lys Arg Ala Glu Asp
50 55 60
Tyr Lys Gly Val Lys Lys Leu Leu Asp Arg Tyr Tyr Leu Ser Phe Ile
65 70 75 80
Asn Asp Val Leu His Ser Ile Lys Leu Lys Asn Leu Asn Asn Tyr Ile
85 90 95
Ser Leu Phe Arg Lys Lys Thr Arg Thr Glu Lys Glu Asn Lys Glu Leu
100 105 110
Glu Asn Leu Glu Ile Asn Leu Arg Lys Glu Ile Ala Lys Ala Phe Lys
115 120 125
Gly Asn Glu Gly Tyr Lys Ser Leu Phe Lys Lys Asp Ile Ile Glu Thr
130 135 140
Ile Leu Pro Glu Phe Leu Asp Asp Lys Asp Glu Ile Ala Leu Val Asn
145 150 155 160
Ser Phe Asn Gly Phe Thr Thr Ala Phe Thr Gly Phe Phe Asp Asn Arg
165 170 175
Glu Asn Met Phe Ser Glu Glu Ala Lys Ser Thr Ser Ile Ala Phe Arg
180 185 190
Cys Ile Asn Glu Asn Leu Thr Arg Tyr Ile Ser Asn Met Asp Ile Phe
195 200 205
Glu Lys Val Asp Ala Ile Phe Asp Lys His Glu Val Gln Glu Ile Lys
210 215 220
Glu Lys Ile Leu Asn Ser Asp Tyr Asp Val Glu Asp Phe Phe Glu Gly
225 230 235 240
Glu Phe Phe Asn Phe Val Leu Thr Gln Glu Gly Ile Asp Val Tyr Asn
245 250 255
Ala Ile Ile Gly Gly Phe Val Thr Glu Ser Gly Glu Lys Ile Lys Gly
260 265 270
Leu Asn Glu Tyr Ile Asn Leu Tyr Asn Gln Lys Thr Lys Gln Lys Leu
275 280 285
Pro Lys Phe Lys Pro Leu Tyr Lys Gln Val Leu Ser Asp Arg Glu Ser
290 295 300
Leu Ser Phe Tyr Gly Glu Gly Tyr Thr Ser Asp Glu Glu Val Leu Glu
305 310 315 320
Val Phe Arg Asn Thr Leu Asn Lys Asn Ser Glu Ile Phe Ser Ser Ile
325 330 335
Lys Lys Leu Glu Lys Leu Phe Lys Asn Phe Asp Glu Tyr Ser Ser Ala
340 345 350
Gly Ile Phe Val Lys Asn Gly Pro Ala Ile Ser Thr Ile Ser Lys Asp
355 360 365
Ile Phe Gly Glu Trp Asn Val Ile Arg Asp Lys Trp Asn Ala Glu Tyr
370 375 380
Asp Asp Ile His Leu Lys Lys Lys Ala Val Val Thr Glu Lys Tyr Glu
385 390 395 400
Asp Asp Arg Arg Lys Ser Phe Lys Lys Ile Gly Ser Phe Ser Leu Glu
405 410 415
Gln Leu Gln Glu Tyr Ala Asp Ala Asp Leu Ser Val Val Glu Lys Leu
420 425 430
Lys Glu Ile Ile Ile Gln Lys Val Asp Glu Ile Tyr Lys Val Tyr Gly
435 440 445
Ser Ser Glu Lys Leu Phe Asp Ala Asp Phe Val Leu Glu Lys Ser Leu
450 455 460
Lys Lys Asn Asp Ala Val Val Ala Ile Met Lys Asp Leu Leu Asp Ser
465 470 475 480
Val Lys Ser Phe Glu Asn Tyr Ile Lys Ala Phe Phe Gly Glu Gly Lys
485 490 495
Glu Thr Asn Arg Asp Glu Ser Phe Tyr Gly Asp Phe Val Leu Ala Tyr
500 505 510
Asp Ile Leu Leu Lys Val Asp His Ile Tyr Asp Ala Ile Arg Asn Tyr
515 520 525
Val Thr Gln Lys Pro Tyr Ser Lys Asp Lys Phe Lys Leu Tyr Phe Gln
530 535 540
Asn Pro Gln Phe Met Arg Gly Trp Asp Lys Asp Lys Glu Thr Asp Tyr
545 550 555 560
Arg Ala Thr Ile Leu Arg Tyr Gly Ser Lys Tyr Tyr Leu Ala Ile Met
565 570 575
Asp Lys Lys Tyr Ala Lys Cys Leu Gln Lys Ile Asp Lys Asp Asp Val
580 585 590
Asn Gly Asn Tyr Glu Lys Ile Asn Tyr Lys Leu Leu Pro Gly Pro Asn
595 600 605
Lys Met Leu Pro Arg Val Phe Phe Ser Lys Lys Trp Met Ala Tyr Tyr
610 615 620
Asn Pro Ser Glu Asp Ile Gln Lys Ile Tyr Lys Asn Gly Thr Phe Lys
625 630 635 640
Lys Gly Asp Met Phe Asn Leu Asn Asp Cys His Lys Leu Ile Asp Phe
645 650 655
Phe Lys Asp Ser Ile Ser Arg Tyr Pro Lys Trp Ser Asn Ala Tyr Asp
660 665 670
Phe Asn Phe Ser Glu Thr Glu Lys Tyr Lys Asp Ile Ala Gly Phe Tyr
675 680 685
Arg Glu Val Glu Glu Gln Gly Tyr Lys Val Ser Phe Glu Ser Ala Ser
690 695 700
Lys Lys Glu Val Asp Lys Leu Val Glu Glu Gly Lys Leu Tyr Met Phe
705 710 715 720
Gln Ile Tyr Asn Lys Asp Phe Ser Asp Lys Ser His Gly Thr Pro Asn
725 730 735
Leu His Thr Met Tyr Phe Lys Leu Leu Phe Asp Glu Asn Asn His Gly
740 745 750
Gln Ile Arg Leu Ser Gly Gly Ala Glu Leu Phe Met Arg Arg Ala Ser
755 760 765
Leu Lys Lys Glu Glu Leu Val Val His Pro Ala Asn Ser Pro Ile Ala
770 775 780
Asn Lys Asn Pro Asp Asn Pro Lys Lys Thr Thr Thr Leu Ser Tyr Asp
785 790 795 800
Val Tyr Lys Asp Lys Arg Phe Ser Glu Asp Gln Tyr Glu Leu His Ile
805 810 815
Pro Ile Ala Ile Asn Lys Cys Pro Lys Asn Ile Phe Lys Ile Asn Thr
820 825 830
Glu Val Arg Val Leu Leu Lys His Asp Asp Asn Pro Tyr Val Ile Gly
835 840 845
Ile Asp Arg Gly Glu Arg Asn Leu Leu Tyr Ile Val Val Val Asp Gly
850 855 860
Lys Gly Asn Ile Val Glu Gln Tyr Ser Leu Asn Glu Ile Ile Asn Asn
865 870 875 880
Phe Asn Gly Ile Arg Ile Lys Thr Asp Tyr His Ser Leu Leu Asp Lys
885 890 895
Lys Glu Lys Glu Arg Phe Glu Ala Arg Gln Asn Trp Thr Ser Ile Glu
900 905 910
Asn Ile Lys Glu Leu Lys Ala Gly Tyr Ile Ser Gln Val Val His Lys
915 920 925
Ile Cys Glu Leu Val Glu Lys Tyr Asp Ala Val Ile Ala Leu Glu Asp
930 935 940
Leu Asn Ser Gly Phe Lys Asn Ser Arg Val Lys Val Glu Lys Gln Val
945 950 955 960
Tyr Gln Lys Phe Glu Lys Met Leu Ile Asp Lys Leu Asn Tyr Met Val
965 970 975
Asp Lys Lys Ser Asn Pro Cys Ala Thr Gly Gly Ala Leu Lys Gly Tyr
980 985 990
Gln Ile Thr Asn Lys Phe Glu Ser Phe Lys Ser Met Ser Thr Gln Asn
995 1000 1005
Gly Phe Ile Phe Tyr Ile Pro Ala Trp Leu Thr Ser Lys Ile Asp
1010 1015 1020
Pro Ser Thr Gly Phe Val Asn Leu Leu Lys Thr Lys Tyr Thr Ser
1025 1030 1035
Ile Ala Asp Ser Lys Lys Phe Ile Ser Ser Phe Asp Arg Ile Met
1040 1045 1050
Tyr Val Pro Glu Glu Asp Leu Phe Glu Phe Ala Leu Asp Tyr Lys
1055 1060 1065
Asn Phe Ser Arg Thr Asp Ala Asp Tyr Ile Lys Lys Trp Lys Leu
1070 1075 1080
Tyr Ser Tyr Gly Asn Arg Ile Arg Ile Phe Arg Asn Pro Lys Lys
1085 1090 1095
Asn Asn Val Phe Asp Trp Glu Glu Val Cys Leu Thr Ser Ala Tyr
1100 1105 1110
Lys Glu Leu Phe Asn Lys Tyr Gly Ile Asn Tyr Gln Gln Gly Asp
1115 1120 1125
Ile Arg Ala Leu Leu Cys Glu Gln Ser Asp Lys Ala Phe Tyr Ser
1130 1135 1140
Ser Phe Met Ala Leu Met Ser Leu Met Leu Gln Met Ala Asn Ser
1145 1150 1155
Ile Thr Gly Arg Thr Asp Val Asp Phe Leu Ile Ser Pro Val Lys
1160 1165 1170
Asn Ser Asp Gly Ile Phe Tyr Asp Ser Arg Asn Tyr Glu Ala Gln
1175 1180 1185
Glu Asn Ala Ile Leu Pro Lys Asn Ala Asp Ala Asn Gly Ala Tyr
1190 1195 1200
Asn Ile Ala Arg Lys Val Leu Trp Ala Ile Gly Gln Phe Lys Lys
1205 1210 1215
Ala Glu Asp Glu Lys Leu Asp Lys Val Lys Ile Ala Ile Ser Asn
1220 1225 1230
Lys Glu Trp Leu Glu Tyr Ala Gln Thr Ser Val Lys His Ser Gly
1235 1240 1245
Gly Ser Ser Gly Gly Ser Ser Gly Ser Glu Thr Pro Gly Thr Ser
1250 1255 1260
Glu Ser Ala Thr Pro Glu Ser Ser Gly Gly Ser Ser Gly Gly Ser
1265 1270 1275
Thr Leu Asn Ile Glu Asp Glu Tyr Arg Leu His Glu Thr Ser Lys
1280 1285 1290
Glu Pro Asp Val Ser Leu Gly Ser Thr Trp Leu Ser Asp Phe Pro
1295 1300 1305
Gln Ala Trp Ala Glu Thr Gly Gly Met Gly Leu Ala Val Arg Gln
1310 1315 1320
Ala Pro Leu Ile Ile Pro Leu Lys Ala Thr Ser Thr Pro Val Ser
1325 1330 1335
Ile Lys Gln Tyr Pro Met Ser Gln Glu Ala Arg Leu Gly Ile Lys
1340 1345 1350
Pro His Ile Gln Arg Leu Leu Asp Gln Gly Ile Leu Val Pro Cys
1355 1360 1365
Gln Ser Pro Trp Asn Thr Pro Leu Leu Pro Val Lys Lys Pro Gly
1370 1375 1380
Thr Asn Asp Tyr Arg Pro Val Gln Asp Leu Arg Glu Val Asn Lys
1385 1390 1395
Arg Val Glu Asp Ile His Pro Thr Val Pro Asn Pro Tyr Asn Leu
1400 1405 1410
Leu Ser Gly Leu Pro Pro Ser His Gln Trp Tyr Thr Val Leu Asp
1415 1420 1425
Leu Lys Asp Ala Phe Phe Cys Leu Arg Leu His Pro Thr Ser Gln
1430 1435 1440
Pro Leu Phe Ala Phe Glu Trp Arg Asp Pro Glu Met Gly Ile Ser
1445 1450 1455
Gly Gln Leu Thr Trp Thr Arg Leu Pro Gln Gly Phe Lys Asn Ser
1460 1465 1470
Pro Thr Leu Phe Asn Glu Ala Leu His Arg Asp Leu Ala Asp Phe
1475 1480 1485
Arg Ile Gln His Pro Asp Leu Ile Leu Leu Gln Tyr Val Asp Asp
1490 1495 1500
Leu Leu Leu Ala Ala Thr Ser Glu Leu Asp Cys Gln Gln Gly Thr
1505 1510 1515
Arg Ala Leu Leu Gln Thr Leu Gly Asn Leu Gly Tyr Arg Ala Ser
1520 1525 1530
Ala Lys Lys Ala Gln Ile Cys Gln Lys Gln Val Lys Tyr Leu Gly
1535 1540 1545
Tyr Leu Leu Lys Glu Gly Gln Arg Trp Leu Thr Glu Ala Arg Lys
1550 1555 1560
Glu Thr Val Met Gly Gln Pro Thr Pro Lys Thr Pro Arg Gln Leu
1565 1570 1575
Arg Glu Phe Leu Gly Lys Ala Gly Phe Cys Arg Leu Phe Ile Pro
1580 1585 1590
Gly Phe Ala Glu Met Ala Ala Pro Leu Tyr Pro Leu Thr Lys Pro
1595 1600 1605
Gly Thr Leu Phe Asn Trp Gly Pro Asp Gln Gln Lys Ala Tyr Gln
1610 1615 1620
Glu Ile Lys Gln Ala Leu Leu Thr Ala Pro Ala Leu Gly Leu Pro
1625 1630 1635
Asp Leu Thr Lys Pro Phe Glu Leu Phe Val Asp Glu Lys Gln Gly
1640 1645 1650
Tyr Ala Lys Gly Val Leu Thr Gln Lys Leu Gly Pro Trp Arg Arg
1655 1660 1665
Pro Val Ala Tyr Leu Ser Lys Lys Leu Asp Pro Val Ala Ala Gly
1670 1675 1680
Trp Pro Pro Cys Leu Arg Met Val Ala Ala Ile Ala Val Leu Thr
1685 1690 1695
Lys Asp Ala Gly Lys Leu Thr Met Gly Gln Pro Leu Val Ile Leu
1700 1705 1710
Ala Pro His Ala Val Glu Ala Leu Val Lys Gln Pro Pro Asp Arg
1715 1720 1725
Trp Leu Ser Asn Ala Arg Met Thr His Tyr Gln Ala Leu Leu Leu
1730 1735 1740
Asp Thr Asp Arg Val Gln Phe Gly Pro Val Val Ala Leu Asn Pro
1745 1750 1755
Ala Thr Leu Leu Pro Leu Pro Glu Glu Gly Leu Gln His Asn Cys
1760 1765 1770
Leu Asp Ile Leu Ala Glu Ala His Gly Thr Arg Pro Asp Leu Thr
1775 1780 1785
Asp Gln Pro Leu Pro Asp Ala Asp His Thr Trp Tyr Thr Asp Gly
1790 1795 1800
Ser Ser Leu Leu Gln Glu Gly Gln Arg Lys Ala Gly Ala Ala Val
1805 1810 1815
Thr Thr Glu Thr Glu Val Ile Trp Ala Lys Ala Leu Pro Ala Gly
1820 1825 1830
Thr Ser Ala Gln Arg Ala Glu Leu Ile Ala Leu Thr Gln Ala Leu
1835 1840 1845
Lys Met Ala Glu Gly Lys Lys Leu Asn Val Tyr Thr Asp Ser Arg
1850 1855 1860
Tyr Ala Phe Ala Thr Ala His Ile His Gly Glu Ile Tyr Arg Arg
1865 1870 1875
Arg Gly Trp Leu Thr Ser Glu Gly Lys Glu Ile Lys Asn Lys Asp
1880 1885 1890
Glu Ile Leu Ala Leu Leu Lys Ala Leu Phe Leu Pro Lys Arg Leu
1895 1900 1905
Ser Ile Ile His Cys Pro Gly His Gln Lys Gly His Ser Ala Glu
1910 1915 1920
Ala Arg Gly Asn Arg Met Ala Asp Gln Ala Ala Arg Lys Ala Ala
1925 1930 1935
Ile Thr Glu Thr Pro Asp Thr Ser Thr Leu Leu Ile Glu Asn Ser
1940 1945 1950
Ser Pro Ser Gly Gly Ser Lys Arg Thr Ala Asp Gly Ser Glu Phe
1955 1960 1965
Glu Pro Lys Lys Lys Arg Lys Val Gly Ser Gly
1970 1975
<210> 67
<211> 1963
<212> PRT
<213> 人工的
<220>
<223> 合成的REDRAW编辑器
<400> 67
Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys
1 5 10 15
Arg Lys Val Thr Leu Asn Ile Glu Asp Glu Tyr Arg Leu His Glu Thr
20 25 30
Ser Lys Glu Pro Asp Val Ser Leu Gly Ser Thr Trp Leu Ser Asp Phe
35 40 45
Pro Gln Ala Trp Ala Glu Thr Gly Gly Met Gly Leu Ala Val Arg Gln
50 55 60
Ala Pro Leu Ile Ile Pro Leu Lys Ala Thr Ser Thr Pro Val Ser Ile
65 70 75 80
Lys Gln Tyr Pro Met Ser Gln Glu Ala Arg Leu Gly Ile Lys Pro His
85 90 95
Ile Gln Arg Leu Leu Asp Gln Gly Ile Leu Val Pro Cys Gln Ser Pro
100 105 110
Trp Asn Thr Pro Leu Leu Pro Val Lys Lys Pro Gly Thr Asn Asp Tyr
115 120 125
Arg Pro Val Gln Asp Leu Arg Glu Val Asn Lys Arg Val Glu Asp Ile
130 135 140
His Pro Thr Val Pro Asn Pro Tyr Asn Leu Leu Ser Gly Leu Pro Pro
145 150 155 160
Ser His Gln Trp Tyr Thr Val Leu Asp Leu Lys Asp Ala Phe Phe Cys
165 170 175
Leu Arg Leu His Pro Thr Ser Gln Pro Leu Phe Ala Phe Glu Trp Arg
180 185 190
Asp Pro Glu Met Gly Ile Ser Gly Gln Leu Thr Trp Thr Arg Leu Pro
195 200 205
Gln Gly Phe Lys Asn Ser Pro Thr Leu Phe Asn Glu Ala Leu His Arg
210 215 220
Asp Leu Ala Asp Phe Arg Ile Gln His Pro Asp Leu Ile Leu Leu Gln
225 230 235 240
Tyr Val Asp Asp Leu Leu Leu Ala Ala Thr Ser Glu Leu Asp Cys Gln
245 250 255
Gln Gly Thr Arg Ala Leu Leu Gln Thr Leu Gly Asn Leu Gly Tyr Arg
260 265 270
Ala Ser Ala Lys Lys Ala Gln Ile Cys Gln Lys Gln Val Lys Tyr Leu
275 280 285
Gly Tyr Leu Leu Lys Glu Gly Gln Arg Trp Leu Thr Glu Ala Arg Lys
290 295 300
Glu Thr Val Met Gly Gln Pro Thr Pro Lys Thr Pro Arg Gln Leu Arg
305 310 315 320
Glu Phe Leu Gly Lys Ala Gly Phe Cys Arg Leu Phe Ile Pro Gly Phe
325 330 335
Ala Glu Met Ala Ala Pro Leu Tyr Pro Leu Thr Lys Pro Gly Thr Leu
340 345 350
Phe Asn Trp Gly Pro Asp Gln Gln Lys Ala Tyr Gln Glu Ile Lys Gln
355 360 365
Ala Leu Leu Thr Ala Pro Ala Leu Gly Leu Pro Asp Leu Thr Lys Pro
370 375 380
Phe Glu Leu Phe Val Asp Glu Lys Gln Gly Tyr Ala Lys Gly Val Leu
385 390 395 400
Thr Gln Lys Leu Gly Pro Trp Arg Arg Pro Val Ala Tyr Leu Ser Lys
405 410 415
Lys Leu Asp Pro Val Ala Ala Gly Trp Pro Pro Cys Leu Arg Met Val
420 425 430
Ala Ala Ile Ala Val Leu Thr Lys Asp Ala Gly Lys Leu Thr Met Gly
435 440 445
Gln Pro Leu Val Ile Leu Ala Pro His Ala Val Glu Ala Leu Val Lys
450 455 460
Gln Pro Pro Asp Arg Trp Leu Ser Asn Ala Arg Met Thr His Tyr Gln
465 470 475 480
Ala Leu Leu Leu Asp Thr Asp Arg Val Gln Phe Gly Pro Val Val Ala
485 490 495
Leu Asn Pro Ala Thr Leu Leu Pro Leu Pro Glu Glu Gly Leu Gln His
500 505 510
Asn Cys Leu Asp Ile Leu Ala Glu Ala His Gly Thr Arg Pro Asp Leu
515 520 525
Thr Asp Gln Pro Leu Pro Asp Ala Asp His Thr Trp Tyr Thr Asp Gly
530 535 540
Ser Ser Leu Leu Gln Glu Gly Gln Arg Lys Ala Gly Ala Ala Val Thr
545 550 555 560
Thr Glu Thr Glu Val Ile Trp Ala Lys Ala Leu Pro Ala Gly Thr Ser
565 570 575
Ala Gln Arg Ala Glu Leu Ile Ala Leu Thr Gln Ala Leu Lys Met Ala
580 585 590
Glu Gly Lys Lys Leu Asn Val Tyr Thr Asp Ser Arg Tyr Ala Phe Ala
595 600 605
Thr Ala His Ile His Gly Glu Ile Tyr Arg Arg Arg Gly Trp Leu Thr
610 615 620
Ser Glu Gly Lys Glu Ile Lys Asn Lys Asp Glu Ile Leu Ala Leu Leu
625 630 635 640
Lys Ala Leu Phe Leu Pro Lys Arg Leu Ser Ile Ile His Cys Pro Gly
645 650 655
His Gln Lys Gly His Ser Ala Glu Ala Arg Gly Asn Arg Met Ala Asp
660 665 670
Gln Ala Ala Arg Lys Ala Ala Ile Thr Glu Thr Pro Asp Thr Ser Thr
675 680 685
Leu Leu Ile Glu Asn Ser Ser Pro Ser Gly Ser Glu Thr Pro Gly Thr
690 695 700
Ser Glu Ser Ala Thr Pro Glu Ser Ser Lys Leu Glu Lys Phe Thr Asn
705 710 715 720
Cys Tyr Ser Leu Ser Lys Thr Leu Arg Phe Lys Ala Ile Pro Val Gly
725 730 735
Lys Thr Gln Glu Asn Ile Asp Asn Lys Arg Leu Leu Val Glu Asp Glu
740 745 750
Lys Arg Ala Glu Asp Tyr Lys Gly Val Lys Lys Leu Leu Asp Arg Tyr
755 760 765
Tyr Leu Ser Phe Ile Asn Asp Val Leu His Ser Ile Lys Leu Lys Asn
770 775 780
Leu Asn Asn Tyr Ile Ser Leu Phe Arg Lys Lys Thr Arg Thr Glu Lys
785 790 795 800
Glu Asn Lys Glu Leu Glu Asn Leu Glu Ile Asn Leu Arg Lys Glu Ile
805 810 815
Ala Lys Ala Phe Lys Gly Asn Glu Gly Tyr Lys Ser Leu Phe Lys Lys
820 825 830
Asp Ile Ile Glu Thr Ile Leu Pro Glu Phe Leu Asp Asp Lys Asp Glu
835 840 845
Ile Ala Leu Val Asn Ser Phe Asn Gly Phe Thr Thr Ala Phe Thr Gly
850 855 860
Phe Phe Asp Asn Arg Glu Asn Met Phe Ser Glu Glu Ala Lys Ser Thr
865 870 875 880
Ser Ile Ala Phe Arg Cys Ile Asn Glu Asn Leu Thr Arg Tyr Ile Ser
885 890 895
Asn Met Asp Ile Phe Glu Lys Val Asp Ala Ile Phe Asp Lys His Glu
900 905 910
Val Gln Glu Ile Lys Glu Lys Ile Leu Asn Ser Asp Tyr Asp Val Glu
915 920 925
Asp Phe Phe Glu Gly Glu Phe Phe Asn Phe Val Leu Thr Gln Glu Gly
930 935 940
Ile Asp Val Tyr Asn Ala Ile Ile Gly Gly Phe Val Thr Glu Ser Gly
945 950 955 960
Glu Lys Ile Lys Gly Leu Asn Glu Tyr Ile Asn Leu Tyr Asn Gln Lys
965 970 975
Thr Lys Gln Lys Leu Pro Lys Phe Lys Pro Leu Tyr Lys Gln Val Leu
980 985 990
Ser Asp Arg Glu Ser Leu Ser Phe Tyr Gly Glu Gly Tyr Thr Ser Asp
995 1000 1005
Glu Glu Val Leu Glu Val Phe Arg Asn Thr Leu Asn Lys Asn Ser
1010 1015 1020
Glu Ile Phe Ser Ser Ile Lys Lys Leu Glu Lys Leu Phe Lys Asn
1025 1030 1035
Phe Asp Glu Tyr Ser Ser Ala Gly Ile Phe Val Lys Asn Gly Pro
1040 1045 1050
Ala Ile Ser Thr Ile Ser Lys Asp Ile Phe Gly Glu Trp Asn Val
1055 1060 1065
Ile Arg Asp Lys Trp Asn Ala Glu Tyr Asp Asp Ile His Leu Lys
1070 1075 1080
Lys Lys Ala Val Val Thr Glu Lys Tyr Glu Asp Asp Arg Arg Lys
1085 1090 1095
Ser Phe Lys Lys Ile Gly Ser Phe Ser Leu Glu Gln Leu Gln Glu
1100 1105 1110
Tyr Ala Asp Ala Asp Leu Ser Val Val Glu Lys Leu Lys Glu Ile
1115 1120 1125
Ile Ile Gln Lys Val Asp Glu Ile Tyr Lys Val Tyr Gly Ser Ser
1130 1135 1140
Glu Lys Leu Phe Asp Ala Asp Phe Val Leu Glu Lys Ser Leu Lys
1145 1150 1155
Lys Asn Asp Ala Val Val Ala Ile Met Lys Asp Leu Leu Asp Ser
1160 1165 1170
Val Lys Ser Phe Glu Asn Tyr Ile Lys Ala Phe Phe Gly Glu Gly
1175 1180 1185
Lys Glu Thr Asn Arg Asp Glu Ser Phe Tyr Gly Asp Phe Val Leu
1190 1195 1200
Ala Tyr Asp Ile Leu Leu Lys Val Asp His Ile Tyr Asp Ala Ile
1205 1210 1215
Arg Asn Tyr Val Thr Gln Lys Pro Tyr Ser Lys Asp Lys Phe Lys
1220 1225 1230
Leu Tyr Phe Gln Asn Pro Gln Phe Met Arg Gly Trp Asp Lys Asp
1235 1240 1245
Val Glu Thr Asp Tyr Arg Ala Thr Ile Leu Arg Tyr Gly Ser Lys
1250 1255 1260
Tyr Tyr Leu Ala Ile Met Asp Lys Lys Tyr Ala Lys Cys Leu Gln
1265 1270 1275
Lys Ile Asp Lys Asp Asp Val Asn Gly Asn Tyr Glu Lys Ile Asn
1280 1285 1290
Tyr Lys Leu Leu Pro Gly Pro Asn Lys Met Leu Pro Arg Val Phe
1295 1300 1305
Phe Ser Lys Lys Trp Met Ala Tyr Tyr Asn Pro Ser Glu Asp Ile
1310 1315 1320
Gln Lys Ile Tyr Lys Asn Gly Thr Phe Lys Lys Gly Asp Met Phe
1325 1330 1335
Asn Leu Asn Asp Cys His Lys Leu Ile Asp Phe Phe Lys Asp Ser
1340 1345 1350
Ile Ser Arg Tyr Pro Lys Trp Ser Asn Ala Tyr Asp Phe Asn Phe
1355 1360 1365
Ser Glu Thr Glu Lys Tyr Lys Asp Ile Ala Gly Phe Tyr Arg Glu
1370 1375 1380
Val Glu Glu Gln Gly Tyr Lys Val Ser Phe Glu Ser Ala Ser Lys
1385 1390 1395
Lys Glu Val Asp Lys Leu Val Glu Glu Gly Lys Leu Tyr Met Phe
1400 1405 1410
Gln Ile Tyr Asn Lys Asp Phe Ser Asp Lys Ser His Gly Thr Pro
1415 1420 1425
Asn Leu His Thr Met Tyr Phe Lys Leu Leu Phe Asp Glu Asn Asn
1430 1435 1440
His Gly Gln Ile Arg Leu Ser Gly Gly Ala Glu Leu Phe Met Arg
1445 1450 1455
Arg Ala Ser Leu Lys Lys Glu Glu Leu Val Val His Pro Ala Asn
1460 1465 1470
Ser Pro Ile Ala Asn Lys Asn Pro Asp Asn Pro Lys Lys Thr Thr
1475 1480 1485
Thr Leu Ser Tyr Asp Val Tyr Lys Asp Lys Arg Phe Ser Glu Asp
1490 1495 1500
Gln Tyr Glu Leu His Ile Pro Ile Ala Ile Asn Lys Cys Pro Lys
1505 1510 1515
Asn Ile Phe Lys Ile Asn Thr Glu Val Arg Val Leu Leu Lys His
1520 1525 1530
Asp Asp Asn Pro Tyr Val Ile Gly Ile Asp Arg Gly Glu Arg Asn
1535 1540 1545
Leu Leu Tyr Ile Val Val Val Asp Gly Lys Gly Asn Ile Val Glu
1550 1555 1560
Gln Tyr Ser Leu Asn Glu Ile Ile Asn Asn Phe Asn Gly Ile Arg
1565 1570 1575
Ile Lys Thr Asp Tyr His Ser Leu Leu Asp Lys Lys Glu Lys Glu
1580 1585 1590
Arg Phe Glu Ala Arg Gln Asn Trp Thr Ser Ile Glu Asn Ile Lys
1595 1600 1605
Glu Leu Lys Ala Gly Tyr Ile Ser Gln Val Val His Lys Ile Cys
1610 1615 1620
Glu Leu Val Glu Lys Tyr Asp Ala Val Ile Ala Leu Glu Asp Leu
1625 1630 1635
Asn Ser Gly Phe Lys Asn Ser Arg Val Lys Val Glu Lys Gln Val
1640 1645 1650
Tyr Gln Lys Phe Glu Lys Met Leu Ile Asp Lys Leu Asn Tyr Met
1655 1660 1665
Val Asp Lys Lys Ser Asn Pro Cys Ala Thr Gly Gly Ala Leu Lys
1670 1675 1680
Gly Tyr Gln Ile Thr Asn Lys Phe Glu Ser Phe Lys Ser Met Ser
1685 1690 1695
Thr Gln Asn Gly Phe Ile Phe Tyr Ile Pro Ala Trp Leu Thr Ser
1700 1705 1710
Lys Ile Asp Pro Ser Thr Gly Phe Val Asn Leu Leu Lys Thr Lys
1715 1720 1725
Tyr Thr Ser Ile Ala Asp Ser Lys Lys Phe Ile Ser Ser Phe Asp
1730 1735 1740
Arg Ile Met Tyr Val Pro Glu Glu Asp Leu Phe Glu Phe Ala Leu
1745 1750 1755
Asp Tyr Lys Asn Phe Ser Arg Thr Asp Ala Asp Tyr Ile Lys Lys
1760 1765 1770
Trp Lys Leu Tyr Ser Tyr Gly Asn Arg Ile Arg Ile Phe Arg Asn
1775 1780 1785
Pro Lys Lys Asn Asn Val Phe Asp Trp Glu Glu Val Cys Leu Thr
1790 1795 1800
Ser Ala Tyr Lys Glu Leu Phe Asn Lys Tyr Gly Ile Asn Tyr Gln
1805 1810 1815
Gln Gly Asp Ile Arg Ala Leu Leu Cys Glu Gln Ser Asp Lys Ala
1820 1825 1830
Phe Tyr Ser Ser Phe Met Ala Leu Met Ser Leu Met Leu Gln Met
1835 1840 1845
Ala Asn Ser Ile Thr Gly Arg Thr Asp Val Asp Phe Leu Ile Ser
1850 1855 1860
Pro Val Lys Asn Ser Asp Gly Ile Phe Tyr Asp Ser Arg Asn Tyr
1865 1870 1875
Glu Ala Gln Glu Asn Ala Ile Leu Pro Lys Asn Ala Asp Ala Asn
1880 1885 1890
Gly Ala Tyr Asn Ile Ala Arg Lys Val Leu Trp Ala Ile Gly Gln
1895 1900 1905
Phe Lys Lys Ala Glu Asp Glu Lys Leu Asp Lys Val Lys Ile Ala
1910 1915 1920
Ile Ser Asn Lys Glu Trp Leu Glu Tyr Ala Gln Thr Ser Val Lys
1925 1930 1935
His Ser Gly Gly Ser Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu
1940 1945 1950
Pro Lys Lys Lys Arg Lys Val Gly Ser Gly
1955 1960
<210> 68
<211> 1979
<212> PRT
<213> 人工的
<220>
<223> 合成的REDRAW编辑器
<400> 68
Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys
1 5 10 15
Arg Lys Val Thr Leu Asn Ile Glu Asp Glu Tyr Arg Leu His Glu Thr
20 25 30
Ser Lys Glu Pro Asp Val Ser Leu Gly Ser Thr Trp Leu Ser Asp Phe
35 40 45
Pro Gln Ala Trp Ala Glu Thr Gly Gly Met Gly Leu Ala Val Arg Gln
50 55 60
Ala Pro Leu Ile Ile Pro Leu Lys Ala Thr Ser Thr Pro Val Ser Ile
65 70 75 80
Lys Gln Tyr Pro Met Ser Gln Glu Ala Arg Leu Gly Ile Lys Pro His
85 90 95
Ile Gln Arg Leu Leu Asp Gln Gly Ile Leu Val Pro Cys Gln Ser Pro
100 105 110
Trp Asn Thr Pro Leu Leu Pro Val Lys Lys Pro Gly Thr Asn Asp Tyr
115 120 125
Arg Pro Val Gln Asp Leu Arg Glu Val Asn Lys Arg Val Glu Asp Ile
130 135 140
His Pro Thr Val Pro Asn Pro Tyr Asn Leu Leu Ser Gly Leu Pro Pro
145 150 155 160
Ser His Gln Trp Tyr Thr Val Leu Asp Leu Lys Asp Ala Phe Phe Cys
165 170 175
Leu Arg Leu His Pro Thr Ser Gln Pro Leu Phe Ala Phe Glu Trp Arg
180 185 190
Asp Pro Glu Met Gly Ile Ser Gly Gln Leu Thr Trp Thr Arg Leu Pro
195 200 205
Gln Gly Phe Lys Asn Ser Pro Thr Leu Phe Asn Glu Ala Leu His Arg
210 215 220
Asp Leu Ala Asp Phe Arg Ile Gln His Pro Asp Leu Ile Leu Leu Gln
225 230 235 240
Tyr Val Asp Asp Leu Leu Leu Ala Ala Thr Ser Glu Leu Asp Cys Gln
245 250 255
Gln Gly Thr Arg Ala Leu Leu Gln Thr Leu Gly Asn Leu Gly Tyr Arg
260 265 270
Ala Ser Ala Lys Lys Ala Gln Ile Cys Gln Lys Gln Val Lys Tyr Leu
275 280 285
Gly Tyr Leu Leu Lys Glu Gly Gln Arg Trp Leu Thr Glu Ala Arg Lys
290 295 300
Glu Thr Val Met Gly Gln Pro Thr Pro Lys Thr Pro Arg Gln Leu Arg
305 310 315 320
Glu Phe Leu Gly Lys Ala Gly Phe Cys Arg Leu Phe Ile Pro Gly Phe
325 330 335
Ala Glu Met Ala Ala Pro Leu Tyr Pro Leu Thr Lys Pro Gly Thr Leu
340 345 350
Phe Asn Trp Gly Pro Asp Gln Gln Lys Ala Tyr Gln Glu Ile Lys Gln
355 360 365
Ala Leu Leu Thr Ala Pro Ala Leu Gly Leu Pro Asp Leu Thr Lys Pro
370 375 380
Phe Glu Leu Phe Val Asp Glu Lys Gln Gly Tyr Ala Lys Gly Val Leu
385 390 395 400
Thr Gln Lys Leu Gly Pro Trp Arg Arg Pro Val Ala Tyr Leu Ser Lys
405 410 415
Lys Leu Asp Pro Val Ala Ala Gly Trp Pro Pro Cys Leu Arg Met Val
420 425 430
Ala Ala Ile Ala Val Leu Thr Lys Asp Ala Gly Lys Leu Thr Met Gly
435 440 445
Gln Pro Leu Val Ile Leu Ala Pro His Ala Val Glu Ala Leu Val Lys
450 455 460
Gln Pro Pro Asp Arg Trp Leu Ser Asn Ala Arg Met Thr His Tyr Gln
465 470 475 480
Ala Leu Leu Leu Asp Thr Asp Arg Val Gln Phe Gly Pro Val Val Ala
485 490 495
Leu Asn Pro Ala Thr Leu Leu Pro Leu Pro Glu Glu Gly Leu Gln His
500 505 510
Asn Cys Leu Asp Ile Leu Ala Glu Ala His Gly Thr Arg Pro Asp Leu
515 520 525
Thr Asp Gln Pro Leu Pro Asp Ala Asp His Thr Trp Tyr Thr Asp Gly
530 535 540
Ser Ser Leu Leu Gln Glu Gly Gln Arg Lys Ala Gly Ala Ala Val Thr
545 550 555 560
Thr Glu Thr Glu Val Ile Trp Ala Lys Ala Leu Pro Ala Gly Thr Ser
565 570 575
Ala Gln Arg Ala Glu Leu Ile Ala Leu Thr Gln Ala Leu Lys Met Ala
580 585 590
Glu Gly Lys Lys Leu Asn Val Tyr Thr Asp Ser Arg Tyr Ala Phe Ala
595 600 605
Thr Ala His Ile His Gly Glu Ile Tyr Arg Arg Arg Gly Trp Leu Thr
610 615 620
Ser Glu Gly Lys Glu Ile Lys Asn Lys Asp Glu Ile Leu Ala Leu Leu
625 630 635 640
Lys Ala Leu Phe Leu Pro Lys Arg Leu Ser Ile Ile His Cys Pro Gly
645 650 655
His Gln Lys Gly His Ser Ala Glu Ala Arg Gly Asn Arg Met Ala Asp
660 665 670
Gln Ala Ala Arg Lys Ala Ala Ile Thr Glu Thr Pro Asp Thr Ser Thr
675 680 685
Leu Leu Ile Glu Asn Ser Ser Pro Ser Gly Gly Ser Ser Gly Gly Ser
690 695 700
Ser Gly Ser Glu Thr Pro Gly Thr Ser Glu Ser Ala Thr Pro Glu Ser
705 710 715 720
Ser Gly Gly Ser Ser Gly Gly Ser Ser Lys Leu Glu Lys Phe Thr Asn
725 730 735
Cys Tyr Ser Leu Ser Lys Thr Leu Arg Phe Lys Ala Ile Pro Val Gly
740 745 750
Lys Thr Gln Glu Asn Ile Asp Asn Lys Arg Leu Leu Val Glu Asp Glu
755 760 765
Lys Arg Ala Glu Asp Tyr Lys Gly Val Lys Lys Leu Leu Asp Arg Tyr
770 775 780
Tyr Leu Ser Phe Ile Asn Asp Val Leu His Ser Ile Lys Leu Lys Asn
785 790 795 800
Leu Asn Asn Tyr Ile Ser Leu Phe Arg Lys Lys Thr Arg Thr Glu Lys
805 810 815
Glu Asn Lys Glu Leu Glu Asn Leu Glu Ile Asn Leu Arg Lys Glu Ile
820 825 830
Ala Lys Ala Phe Lys Gly Asn Glu Gly Tyr Lys Ser Leu Phe Lys Lys
835 840 845
Asp Ile Ile Glu Thr Ile Leu Pro Glu Phe Leu Asp Asp Lys Asp Glu
850 855 860
Ile Ala Leu Val Asn Ser Phe Asn Gly Phe Thr Thr Ala Phe Thr Gly
865 870 875 880
Phe Phe Asp Asn Arg Glu Asn Met Phe Ser Glu Glu Ala Lys Ser Thr
885 890 895
Ser Ile Ala Phe Arg Cys Ile Asn Glu Asn Leu Thr Arg Tyr Ile Ser
900 905 910
Asn Met Asp Ile Phe Glu Lys Val Asp Ala Ile Phe Asp Lys His Glu
915 920 925
Val Gln Glu Ile Lys Glu Lys Ile Leu Asn Ser Asp Tyr Asp Val Glu
930 935 940
Asp Phe Phe Glu Gly Glu Phe Phe Asn Phe Val Leu Thr Gln Glu Gly
945 950 955 960
Ile Asp Val Tyr Asn Ala Ile Ile Gly Gly Phe Val Thr Glu Ser Gly
965 970 975
Glu Lys Ile Lys Gly Leu Asn Glu Tyr Ile Asn Leu Tyr Asn Gln Lys
980 985 990
Thr Lys Gln Lys Leu Pro Lys Phe Lys Pro Leu Tyr Lys Gln Val Leu
995 1000 1005
Ser Asp Arg Glu Ser Leu Ser Phe Tyr Gly Glu Gly Tyr Thr Ser
1010 1015 1020
Asp Glu Glu Val Leu Glu Val Phe Arg Asn Thr Leu Asn Lys Asn
1025 1030 1035
Ser Glu Ile Phe Ser Ser Ile Lys Lys Leu Glu Lys Leu Phe Lys
1040 1045 1050
Asn Phe Asp Glu Tyr Ser Ser Ala Gly Ile Phe Val Lys Asn Gly
1055 1060 1065
Pro Ala Ile Ser Thr Ile Ser Lys Asp Ile Phe Gly Glu Trp Asn
1070 1075 1080
Val Ile Arg Asp Lys Trp Asn Ala Glu Tyr Asp Asp Ile His Leu
1085 1090 1095
Lys Lys Lys Ala Val Val Thr Glu Lys Tyr Glu Asp Asp Arg Arg
1100 1105 1110
Lys Ser Phe Lys Lys Ile Gly Ser Phe Ser Leu Glu Gln Leu Gln
1115 1120 1125
Glu Tyr Ala Asp Ala Asp Leu Ser Val Val Glu Lys Leu Lys Glu
1130 1135 1140
Ile Ile Ile Gln Lys Val Asp Glu Ile Tyr Lys Val Tyr Gly Ser
1145 1150 1155
Ser Glu Lys Leu Phe Asp Ala Asp Phe Val Leu Glu Lys Ser Leu
1160 1165 1170
Lys Lys Asn Asp Ala Val Val Ala Ile Met Lys Asp Leu Leu Asp
1175 1180 1185
Ser Val Lys Ser Phe Glu Asn Tyr Ile Lys Ala Phe Phe Gly Glu
1190 1195 1200
Gly Lys Glu Thr Asn Arg Asp Glu Ser Phe Tyr Gly Asp Phe Val
1205 1210 1215
Leu Ala Tyr Asp Ile Leu Leu Lys Val Asp His Ile Tyr Asp Ala
1220 1225 1230
Ile Arg Asn Tyr Val Thr Gln Lys Pro Tyr Ser Lys Asp Lys Phe
1235 1240 1245
Lys Leu Tyr Phe Gln Asn Pro Gln Phe Met Arg Gly Trp Asp Lys
1250 1255 1260
Asp Val Glu Thr Asp Tyr Arg Ala Thr Ile Leu Arg Tyr Gly Ser
1265 1270 1275
Lys Tyr Tyr Leu Ala Ile Met Asp Lys Lys Tyr Ala Lys Cys Leu
1280 1285 1290
Gln Lys Ile Asp Lys Asp Asp Val Asn Gly Asn Tyr Glu Lys Ile
1295 1300 1305
Asn Tyr Lys Leu Leu Pro Gly Pro Asn Lys Met Leu Pro Arg Val
1310 1315 1320
Phe Phe Ser Lys Lys Trp Met Ala Tyr Tyr Asn Pro Ser Glu Asp
1325 1330 1335
Ile Gln Lys Ile Tyr Lys Asn Gly Thr Phe Lys Lys Gly Asp Met
1340 1345 1350
Phe Asn Leu Asn Asp Cys His Lys Leu Ile Asp Phe Phe Lys Asp
1355 1360 1365
Ser Ile Ser Arg Tyr Pro Lys Trp Ser Asn Ala Tyr Asp Phe Asn
1370 1375 1380
Phe Ser Glu Thr Glu Lys Tyr Lys Asp Ile Ala Gly Phe Tyr Arg
1385 1390 1395
Glu Val Glu Glu Gln Gly Tyr Lys Val Ser Phe Glu Ser Ala Ser
1400 1405 1410
Lys Lys Glu Val Asp Lys Leu Val Glu Glu Gly Lys Leu Tyr Met
1415 1420 1425
Phe Gln Ile Tyr Asn Lys Asp Phe Ser Asp Lys Ser His Gly Thr
1430 1435 1440
Pro Asn Leu His Thr Met Tyr Phe Lys Leu Leu Phe Asp Glu Asn
1445 1450 1455
Asn His Gly Gln Ile Arg Leu Ser Gly Gly Ala Glu Leu Phe Met
1460 1465 1470
Arg Arg Ala Ser Leu Lys Lys Glu Glu Leu Val Val His Pro Ala
1475 1480 1485
Asn Ser Pro Ile Ala Asn Lys Asn Pro Asp Asn Pro Lys Lys Thr
1490 1495 1500
Thr Thr Leu Ser Tyr Asp Val Tyr Lys Asp Lys Arg Phe Ser Glu
1505 1510 1515
Asp Gln Tyr Glu Leu His Ile Pro Ile Ala Ile Asn Lys Cys Pro
1520 1525 1530
Lys Asn Ile Phe Lys Ile Asn Thr Glu Val Arg Val Leu Leu Lys
1535 1540 1545
His Asp Asp Asn Pro Tyr Val Ile Gly Ile Asp Arg Gly Glu Arg
1550 1555 1560
Asn Leu Leu Tyr Ile Val Val Val Asp Gly Lys Gly Asn Ile Val
1565 1570 1575
Glu Gln Tyr Ser Leu Asn Glu Ile Ile Asn Asn Phe Asn Gly Ile
1580 1585 1590
Arg Ile Lys Thr Asp Tyr His Ser Leu Leu Asp Lys Lys Glu Lys
1595 1600 1605
Glu Arg Phe Glu Ala Arg Gln Asn Trp Thr Ser Ile Glu Asn Ile
1610 1615 1620
Lys Glu Leu Lys Ala Gly Tyr Ile Ser Gln Val Val His Lys Ile
1625 1630 1635
Cys Glu Leu Val Glu Lys Tyr Asp Ala Val Ile Ala Leu Glu Asp
1640 1645 1650
Leu Asn Ser Gly Phe Lys Asn Ser Arg Val Lys Val Glu Lys Gln
1655 1660 1665
Val Tyr Gln Lys Phe Glu Lys Met Leu Ile Asp Lys Leu Asn Tyr
1670 1675 1680
Met Val Asp Lys Lys Ser Asn Pro Cys Ala Thr Gly Gly Ala Leu
1685 1690 1695
Lys Gly Tyr Gln Ile Thr Asn Lys Phe Glu Ser Phe Lys Ser Met
1700 1705 1710
Ser Thr Gln Asn Gly Phe Ile Phe Tyr Ile Pro Ala Trp Leu Thr
1715 1720 1725
Ser Lys Ile Asp Pro Ser Thr Gly Phe Val Asn Leu Leu Lys Thr
1730 1735 1740
Lys Tyr Thr Ser Ile Ala Asp Ser Lys Lys Phe Ile Ser Ser Phe
1745 1750 1755
Asp Arg Ile Met Tyr Val Pro Glu Glu Asp Leu Phe Glu Phe Ala
1760 1765 1770
Leu Asp Tyr Lys Asn Phe Ser Arg Thr Asp Ala Asp Tyr Ile Lys
1775 1780 1785
Lys Trp Lys Leu Tyr Ser Tyr Gly Asn Arg Ile Arg Ile Phe Arg
1790 1795 1800
Asn Pro Lys Lys Asn Asn Val Phe Asp Trp Glu Glu Val Cys Leu
1805 1810 1815
Thr Ser Ala Tyr Lys Glu Leu Phe Asn Lys Tyr Gly Ile Asn Tyr
1820 1825 1830
Gln Gln Gly Asp Ile Arg Ala Leu Leu Cys Glu Gln Ser Asp Lys
1835 1840 1845
Ala Phe Tyr Ser Ser Phe Met Ala Leu Met Ser Leu Met Leu Gln
1850 1855 1860
Met Ala Asn Ser Ile Thr Gly Arg Thr Asp Val Asp Phe Leu Ile
1865 1870 1875
Ser Pro Val Lys Asn Ser Asp Gly Ile Phe Tyr Asp Ser Arg Asn
1880 1885 1890
Tyr Glu Ala Gln Glu Asn Ala Ile Leu Pro Lys Asn Ala Asp Ala
1895 1900 1905
Asn Gly Ala Tyr Asn Ile Ala Arg Lys Val Leu Trp Ala Ile Gly
1910 1915 1920
Gln Phe Lys Lys Ala Glu Asp Glu Lys Leu Asp Lys Val Lys Ile
1925 1930 1935
Ala Ile Ser Asn Lys Glu Trp Leu Glu Tyr Ala Gln Thr Ser Val
1940 1945 1950
Lys His Ser Gly Gly Ser Lys Arg Thr Ala Asp Gly Ser Glu Phe
1955 1960 1965
Glu Pro Lys Lys Lys Arg Lys Val Gly Ser Gly
1970 1975
<210> 69
<211> 1963
<212> PRT
<213> 人工的
<220>
<223> 合成的REDRAW编辑器
<400> 69
Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys
1 5 10 15
Arg Lys Val Ser Lys Leu Glu Lys Phe Thr Asn Cys Tyr Ser Leu Ser
20 25 30
Lys Thr Leu Arg Phe Lys Ala Ile Pro Val Gly Lys Thr Gln Glu Asn
35 40 45
Ile Asp Asn Lys Arg Leu Leu Val Glu Asp Glu Lys Arg Ala Glu Asp
50 55 60
Tyr Lys Gly Val Lys Lys Leu Leu Asp Arg Tyr Tyr Leu Ser Phe Ile
65 70 75 80
Asn Asp Val Leu His Ser Ile Lys Leu Lys Asn Leu Asn Asn Tyr Ile
85 90 95
Ser Leu Phe Arg Lys Lys Thr Arg Thr Glu Lys Glu Asn Lys Glu Leu
100 105 110
Glu Asn Leu Glu Ile Asn Leu Arg Lys Glu Ile Ala Lys Ala Phe Lys
115 120 125
Gly Asn Glu Gly Tyr Lys Ser Leu Phe Lys Lys Asp Ile Ile Glu Thr
130 135 140
Ile Leu Pro Glu Phe Leu Asp Asp Lys Asp Glu Ile Ala Leu Val Asn
145 150 155 160
Ser Phe Asn Gly Phe Thr Thr Ala Phe Thr Gly Phe Phe Asp Asn Arg
165 170 175
Glu Asn Met Phe Ser Glu Glu Ala Lys Ser Thr Ser Ile Ala Phe Arg
180 185 190
Cys Ile Asn Glu Asn Leu Thr Arg Tyr Ile Ser Asn Met Asp Ile Phe
195 200 205
Glu Lys Val Asp Ala Ile Phe Asp Lys His Glu Val Gln Glu Ile Lys
210 215 220
Glu Lys Ile Leu Asn Ser Asp Tyr Asp Val Glu Asp Phe Phe Glu Gly
225 230 235 240
Glu Phe Phe Asn Phe Val Leu Thr Gln Glu Gly Ile Asp Val Tyr Asn
245 250 255
Ala Ile Ile Gly Gly Phe Val Thr Glu Ser Gly Glu Lys Ile Lys Gly
260 265 270
Leu Asn Glu Tyr Ile Asn Leu Tyr Asn Gln Lys Thr Lys Gln Lys Leu
275 280 285
Pro Lys Phe Lys Pro Leu Tyr Lys Gln Val Leu Ser Asp Arg Glu Ser
290 295 300
Leu Ser Phe Tyr Gly Glu Gly Tyr Thr Ser Asp Glu Glu Val Leu Glu
305 310 315 320
Val Phe Arg Asn Thr Leu Asn Lys Asn Ser Glu Ile Phe Ser Ser Ile
325 330 335
Lys Lys Leu Glu Lys Leu Phe Lys Asn Phe Asp Glu Tyr Ser Ser Ala
340 345 350
Gly Ile Phe Val Lys Asn Gly Pro Ala Ile Ser Thr Ile Ser Lys Asp
355 360 365
Ile Phe Gly Glu Trp Asn Val Ile Arg Asp Lys Trp Asn Ala Glu Tyr
370 375 380
Asp Asp Ile His Leu Lys Lys Lys Ala Val Val Thr Glu Lys Tyr Glu
385 390 395 400
Asp Asp Arg Arg Lys Ser Phe Lys Lys Ile Gly Ser Phe Ser Leu Glu
405 410 415
Gln Leu Gln Glu Tyr Ala Asp Ala Asp Leu Ser Val Val Glu Lys Leu
420 425 430
Lys Glu Ile Ile Ile Gln Lys Val Asp Glu Ile Tyr Lys Val Tyr Gly
435 440 445
Ser Ser Glu Lys Leu Phe Asp Ala Asp Phe Val Leu Glu Lys Ser Leu
450 455 460
Lys Lys Asn Asp Ala Val Val Ala Ile Met Lys Asp Leu Leu Asp Ser
465 470 475 480
Val Lys Ser Phe Glu Asn Tyr Ile Lys Ala Phe Phe Gly Glu Gly Lys
485 490 495
Glu Thr Asn Arg Asp Glu Ser Phe Tyr Gly Asp Phe Val Leu Ala Tyr
500 505 510
Asp Ile Leu Leu Lys Val Asp His Ile Tyr Asp Ala Ile Arg Asn Tyr
515 520 525
Val Thr Gln Lys Pro Tyr Ser Lys Asp Lys Phe Lys Leu Tyr Phe Gln
530 535 540
Asn Pro Gln Phe Met Arg Gly Trp Asp Lys Asp Val Glu Thr Asp Tyr
545 550 555 560
Arg Ala Thr Ile Leu Arg Tyr Gly Ser Lys Tyr Tyr Leu Ala Ile Met
565 570 575
Asp Lys Lys Tyr Ala Lys Cys Leu Gln Lys Ile Asp Lys Asp Asp Val
580 585 590
Asn Gly Asn Tyr Glu Lys Ile Asn Tyr Lys Leu Leu Pro Gly Pro Asn
595 600 605
Lys Met Leu Pro Arg Val Phe Phe Ser Lys Lys Trp Met Ala Tyr Tyr
610 615 620
Asn Pro Ser Glu Asp Ile Gln Lys Ile Tyr Lys Asn Gly Thr Phe Lys
625 630 635 640
Lys Gly Asp Met Phe Asn Leu Asn Asp Cys His Lys Leu Ile Asp Phe
645 650 655
Phe Lys Asp Ser Ile Ser Arg Tyr Pro Lys Trp Ser Asn Ala Tyr Asp
660 665 670
Phe Asn Phe Ser Glu Thr Glu Lys Tyr Lys Asp Ile Ala Gly Phe Tyr
675 680 685
Arg Glu Val Glu Glu Gln Gly Tyr Lys Val Ser Phe Glu Ser Ala Ser
690 695 700
Lys Lys Glu Val Asp Lys Leu Val Glu Glu Gly Lys Leu Tyr Met Phe
705 710 715 720
Gln Ile Tyr Asn Lys Asp Phe Ser Asp Lys Ser His Gly Thr Pro Asn
725 730 735
Leu His Thr Met Tyr Phe Lys Leu Leu Phe Asp Glu Asn Asn His Gly
740 745 750
Gln Ile Arg Leu Ser Gly Gly Ala Glu Leu Phe Met Arg Arg Ala Ser
755 760 765
Leu Lys Lys Glu Glu Leu Val Val His Pro Ala Asn Ser Pro Ile Ala
770 775 780
Asn Lys Asn Pro Asp Asn Pro Lys Lys Thr Thr Thr Leu Ser Tyr Asp
785 790 795 800
Val Tyr Lys Asp Lys Arg Phe Ser Glu Asp Gln Tyr Glu Leu His Ile
805 810 815
Pro Ile Ala Ile Asn Lys Cys Pro Lys Asn Ile Phe Lys Ile Asn Thr
820 825 830
Glu Val Arg Val Leu Leu Lys His Asp Asp Asn Pro Tyr Val Ile Gly
835 840 845
Ile Asp Arg Gly Glu Arg Asn Leu Leu Tyr Ile Val Val Val Asp Gly
850 855 860
Lys Gly Asn Ile Val Glu Gln Tyr Ser Leu Asn Glu Ile Ile Asn Asn
865 870 875 880
Phe Asn Gly Ile Arg Ile Lys Thr Asp Tyr His Ser Leu Leu Asp Lys
885 890 895
Lys Glu Lys Glu Arg Phe Glu Ala Arg Gln Asn Trp Thr Ser Ile Glu
900 905 910
Asn Ile Lys Glu Leu Lys Ala Gly Tyr Ile Ser Gln Val Val His Lys
915 920 925
Ile Cys Glu Leu Val Glu Lys Tyr Asp Ala Val Ile Ala Leu Glu Asp
930 935 940
Leu Asn Ser Gly Phe Lys Asn Ser Arg Val Lys Val Glu Lys Gln Val
945 950 955 960
Tyr Gln Lys Phe Glu Lys Met Leu Ile Asp Lys Leu Asn Tyr Met Val
965 970 975
Asp Lys Lys Ser Asn Pro Cys Ala Thr Gly Gly Ala Leu Lys Gly Tyr
980 985 990
Gln Ile Thr Asn Lys Phe Glu Ser Phe Lys Ser Met Ser Thr Gln Asn
995 1000 1005
Gly Phe Ile Phe Tyr Ile Pro Ala Trp Leu Thr Ser Lys Ile Asp
1010 1015 1020
Pro Ser Thr Gly Phe Val Asn Leu Leu Lys Thr Lys Tyr Thr Ser
1025 1030 1035
Ile Ala Asp Ser Lys Lys Phe Ile Ser Ser Phe Asp Arg Ile Met
1040 1045 1050
Tyr Val Pro Glu Glu Asp Leu Phe Glu Phe Ala Leu Asp Tyr Lys
1055 1060 1065
Asn Phe Ser Arg Thr Asp Ala Asp Tyr Ile Lys Lys Trp Lys Leu
1070 1075 1080
Tyr Ser Tyr Gly Asn Arg Ile Arg Ile Phe Arg Asn Pro Lys Lys
1085 1090 1095
Asn Asn Val Phe Asp Trp Glu Glu Val Cys Leu Thr Ser Ala Tyr
1100 1105 1110
Lys Glu Leu Phe Asn Lys Tyr Gly Ile Asn Tyr Gln Gln Gly Asp
1115 1120 1125
Ile Arg Ala Leu Leu Cys Glu Gln Ser Asp Lys Ala Phe Tyr Ser
1130 1135 1140
Ser Phe Met Ala Leu Met Ser Leu Met Leu Gln Met Ala Asn Ser
1145 1150 1155
Ile Thr Gly Arg Thr Asp Val Asp Phe Leu Ile Ser Pro Val Lys
1160 1165 1170
Asn Ser Asp Gly Ile Phe Tyr Asp Ser Arg Asn Tyr Glu Ala Gln
1175 1180 1185
Glu Asn Ala Ile Leu Pro Lys Asn Ala Asp Ala Asn Gly Ala Tyr
1190 1195 1200
Asn Ile Ala Arg Lys Val Leu Trp Ala Ile Gly Gln Phe Lys Lys
1205 1210 1215
Ala Glu Asp Glu Lys Leu Asp Lys Val Lys Ile Ala Ile Ser Asn
1220 1225 1230
Lys Glu Trp Leu Glu Tyr Ala Gln Thr Ser Val Lys His Ser Gly
1235 1240 1245
Ser Glu Thr Pro Gly Thr Ser Glu Ser Ala Thr Pro Glu Ser Thr
1250 1255 1260
Leu Asn Ile Glu Asp Glu Tyr Arg Leu His Glu Thr Ser Lys Glu
1265 1270 1275
Pro Asp Val Ser Leu Gly Ser Thr Trp Leu Ser Asp Phe Pro Gln
1280 1285 1290
Ala Trp Ala Glu Thr Gly Gly Met Gly Leu Ala Val Arg Gln Ala
1295 1300 1305
Pro Leu Ile Ile Pro Leu Lys Ala Thr Ser Thr Pro Val Ser Ile
1310 1315 1320
Lys Gln Tyr Pro Met Ser Gln Glu Ala Arg Leu Gly Ile Lys Pro
1325 1330 1335
His Ile Gln Arg Leu Leu Asp Gln Gly Ile Leu Val Pro Cys Gln
1340 1345 1350
Ser Pro Trp Asn Thr Pro Leu Leu Pro Val Lys Lys Pro Gly Thr
1355 1360 1365
Asn Asp Tyr Arg Pro Val Gln Asp Leu Arg Glu Val Asn Lys Arg
1370 1375 1380
Val Glu Asp Ile His Pro Thr Val Pro Asn Pro Tyr Asn Leu Leu
1385 1390 1395
Ser Gly Leu Pro Pro Ser His Gln Trp Tyr Thr Val Leu Asp Leu
1400 1405 1410
Lys Asp Ala Phe Phe Cys Leu Arg Leu His Pro Thr Ser Gln Pro
1415 1420 1425
Leu Phe Ala Phe Glu Trp Arg Asp Pro Glu Met Gly Ile Ser Gly
1430 1435 1440
Gln Leu Thr Trp Thr Arg Leu Pro Gln Gly Phe Lys Asn Ser Pro
1445 1450 1455
Thr Leu Phe Asn Glu Ala Leu His Arg Asp Leu Ala Asp Phe Arg
1460 1465 1470
Ile Gln His Pro Asp Leu Ile Leu Leu Gln Tyr Val Asp Asp Leu
1475 1480 1485
Leu Leu Ala Ala Thr Ser Glu Leu Asp Cys Gln Gln Gly Thr Arg
1490 1495 1500
Ala Leu Leu Gln Thr Leu Gly Asn Leu Gly Tyr Arg Ala Ser Ala
1505 1510 1515
Lys Lys Ala Gln Ile Cys Gln Lys Gln Val Lys Tyr Leu Gly Tyr
1520 1525 1530
Leu Leu Lys Glu Gly Gln Arg Trp Leu Thr Glu Ala Arg Lys Glu
1535 1540 1545
Thr Val Met Gly Gln Pro Thr Pro Lys Thr Pro Arg Gln Leu Arg
1550 1555 1560
Glu Phe Leu Gly Lys Ala Gly Phe Cys Arg Leu Phe Ile Pro Gly
1565 1570 1575
Phe Ala Glu Met Ala Ala Pro Leu Tyr Pro Leu Thr Lys Pro Gly
1580 1585 1590
Thr Leu Phe Asn Trp Gly Pro Asp Gln Gln Lys Ala Tyr Gln Glu
1595 1600 1605
Ile Lys Gln Ala Leu Leu Thr Ala Pro Ala Leu Gly Leu Pro Asp
1610 1615 1620
Leu Thr Lys Pro Phe Glu Leu Phe Val Asp Glu Lys Gln Gly Tyr
1625 1630 1635
Ala Lys Gly Val Leu Thr Gln Lys Leu Gly Pro Trp Arg Arg Pro
1640 1645 1650
Val Ala Tyr Leu Ser Lys Lys Leu Asp Pro Val Ala Ala Gly Trp
1655 1660 1665
Pro Pro Cys Leu Arg Met Val Ala Ala Ile Ala Val Leu Thr Lys
1670 1675 1680
Asp Ala Gly Lys Leu Thr Met Gly Gln Pro Leu Val Ile Leu Ala
1685 1690 1695
Pro His Ala Val Glu Ala Leu Val Lys Gln Pro Pro Asp Arg Trp
1700 1705 1710
Leu Ser Asn Ala Arg Met Thr His Tyr Gln Ala Leu Leu Leu Asp
1715 1720 1725
Thr Asp Arg Val Gln Phe Gly Pro Val Val Ala Leu Asn Pro Ala
1730 1735 1740
Thr Leu Leu Pro Leu Pro Glu Glu Gly Leu Gln His Asn Cys Leu
1745 1750 1755
Asp Ile Leu Ala Glu Ala His Gly Thr Arg Pro Asp Leu Thr Asp
1760 1765 1770
Gln Pro Leu Pro Asp Ala Asp His Thr Trp Tyr Thr Asp Gly Ser
1775 1780 1785
Ser Leu Leu Gln Glu Gly Gln Arg Lys Ala Gly Ala Ala Val Thr
1790 1795 1800
Thr Glu Thr Glu Val Ile Trp Ala Lys Ala Leu Pro Ala Gly Thr
1805 1810 1815
Ser Ala Gln Arg Ala Glu Leu Ile Ala Leu Thr Gln Ala Leu Lys
1820 1825 1830
Met Ala Glu Gly Lys Lys Leu Asn Val Tyr Thr Asp Ser Arg Tyr
1835 1840 1845
Ala Phe Ala Thr Ala His Ile His Gly Glu Ile Tyr Arg Arg Arg
1850 1855 1860
Gly Trp Leu Thr Ser Glu Gly Lys Glu Ile Lys Asn Lys Asp Glu
1865 1870 1875
Ile Leu Ala Leu Leu Lys Ala Leu Phe Leu Pro Lys Arg Leu Ser
1880 1885 1890
Ile Ile His Cys Pro Gly His Gln Lys Gly His Ser Ala Glu Ala
1895 1900 1905
Arg Gly Asn Arg Met Ala Asp Gln Ala Ala Arg Lys Ala Ala Ile
1910 1915 1920
Thr Glu Thr Pro Asp Thr Ser Thr Leu Leu Ile Glu Asn Ser Ser
1925 1930 1935
Pro Ser Gly Gly Ser Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu
1940 1945 1950
Pro Lys Lys Lys Arg Lys Val Gly Ser Gly
1955 1960
<210> 70
<211> 1979
<212> PRT
<213> 人工的
<220>
<223> 合成的REDRAW编辑器
<400> 70
Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys
1 5 10 15
Arg Lys Val Ser Lys Leu Glu Lys Phe Thr Asn Cys Tyr Ser Leu Ser
20 25 30
Lys Thr Leu Arg Phe Lys Ala Ile Pro Val Gly Lys Thr Gln Glu Asn
35 40 45
Ile Asp Asn Lys Arg Leu Leu Val Glu Asp Glu Lys Arg Ala Glu Asp
50 55 60
Tyr Lys Gly Val Lys Lys Leu Leu Asp Arg Tyr Tyr Leu Ser Phe Ile
65 70 75 80
Asn Asp Val Leu His Ser Ile Lys Leu Lys Asn Leu Asn Asn Tyr Ile
85 90 95
Ser Leu Phe Arg Lys Lys Thr Arg Thr Glu Lys Glu Asn Lys Glu Leu
100 105 110
Glu Asn Leu Glu Ile Asn Leu Arg Lys Glu Ile Ala Lys Ala Phe Lys
115 120 125
Gly Asn Glu Gly Tyr Lys Ser Leu Phe Lys Lys Asp Ile Ile Glu Thr
130 135 140
Ile Leu Pro Glu Phe Leu Asp Asp Lys Asp Glu Ile Ala Leu Val Asn
145 150 155 160
Ser Phe Asn Gly Phe Thr Thr Ala Phe Thr Gly Phe Phe Asp Asn Arg
165 170 175
Glu Asn Met Phe Ser Glu Glu Ala Lys Ser Thr Ser Ile Ala Phe Arg
180 185 190
Cys Ile Asn Glu Asn Leu Thr Arg Tyr Ile Ser Asn Met Asp Ile Phe
195 200 205
Glu Lys Val Asp Ala Ile Phe Asp Lys His Glu Val Gln Glu Ile Lys
210 215 220
Glu Lys Ile Leu Asn Ser Asp Tyr Asp Val Glu Asp Phe Phe Glu Gly
225 230 235 240
Glu Phe Phe Asn Phe Val Leu Thr Gln Glu Gly Ile Asp Val Tyr Asn
245 250 255
Ala Ile Ile Gly Gly Phe Val Thr Glu Ser Gly Glu Lys Ile Lys Gly
260 265 270
Leu Asn Glu Tyr Ile Asn Leu Tyr Asn Gln Lys Thr Lys Gln Lys Leu
275 280 285
Pro Lys Phe Lys Pro Leu Tyr Lys Gln Val Leu Ser Asp Arg Glu Ser
290 295 300
Leu Ser Phe Tyr Gly Glu Gly Tyr Thr Ser Asp Glu Glu Val Leu Glu
305 310 315 320
Val Phe Arg Asn Thr Leu Asn Lys Asn Ser Glu Ile Phe Ser Ser Ile
325 330 335
Lys Lys Leu Glu Lys Leu Phe Lys Asn Phe Asp Glu Tyr Ser Ser Ala
340 345 350
Gly Ile Phe Val Lys Asn Gly Pro Ala Ile Ser Thr Ile Ser Lys Asp
355 360 365
Ile Phe Gly Glu Trp Asn Val Ile Arg Asp Lys Trp Asn Ala Glu Tyr
370 375 380
Asp Asp Ile His Leu Lys Lys Lys Ala Val Val Thr Glu Lys Tyr Glu
385 390 395 400
Asp Asp Arg Arg Lys Ser Phe Lys Lys Ile Gly Ser Phe Ser Leu Glu
405 410 415
Gln Leu Gln Glu Tyr Ala Asp Ala Asp Leu Ser Val Val Glu Lys Leu
420 425 430
Lys Glu Ile Ile Ile Gln Lys Val Asp Glu Ile Tyr Lys Val Tyr Gly
435 440 445
Ser Ser Glu Lys Leu Phe Asp Ala Asp Phe Val Leu Glu Lys Ser Leu
450 455 460
Lys Lys Asn Asp Ala Val Val Ala Ile Met Lys Asp Leu Leu Asp Ser
465 470 475 480
Val Lys Ser Phe Glu Asn Tyr Ile Lys Ala Phe Phe Gly Glu Gly Lys
485 490 495
Glu Thr Asn Arg Asp Glu Ser Phe Tyr Gly Asp Phe Val Leu Ala Tyr
500 505 510
Asp Ile Leu Leu Lys Val Asp His Ile Tyr Asp Ala Ile Arg Asn Tyr
515 520 525
Val Thr Gln Lys Pro Tyr Ser Lys Asp Lys Phe Lys Leu Tyr Phe Gln
530 535 540
Asn Pro Gln Phe Met Arg Gly Trp Asp Lys Asp Val Glu Thr Asp Tyr
545 550 555 560
Arg Ala Thr Ile Leu Arg Tyr Gly Ser Lys Tyr Tyr Leu Ala Ile Met
565 570 575
Asp Lys Lys Tyr Ala Lys Cys Leu Gln Lys Ile Asp Lys Asp Asp Val
580 585 590
Asn Gly Asn Tyr Glu Lys Ile Asn Tyr Lys Leu Leu Pro Gly Pro Asn
595 600 605
Lys Met Leu Pro Arg Val Phe Phe Ser Lys Lys Trp Met Ala Tyr Tyr
610 615 620
Asn Pro Ser Glu Asp Ile Gln Lys Ile Tyr Lys Asn Gly Thr Phe Lys
625 630 635 640
Lys Gly Asp Met Phe Asn Leu Asn Asp Cys His Lys Leu Ile Asp Phe
645 650 655
Phe Lys Asp Ser Ile Ser Arg Tyr Pro Lys Trp Ser Asn Ala Tyr Asp
660 665 670
Phe Asn Phe Ser Glu Thr Glu Lys Tyr Lys Asp Ile Ala Gly Phe Tyr
675 680 685
Arg Glu Val Glu Glu Gln Gly Tyr Lys Val Ser Phe Glu Ser Ala Ser
690 695 700
Lys Lys Glu Val Asp Lys Leu Val Glu Glu Gly Lys Leu Tyr Met Phe
705 710 715 720
Gln Ile Tyr Asn Lys Asp Phe Ser Asp Lys Ser His Gly Thr Pro Asn
725 730 735
Leu His Thr Met Tyr Phe Lys Leu Leu Phe Asp Glu Asn Asn His Gly
740 745 750
Gln Ile Arg Leu Ser Gly Gly Ala Glu Leu Phe Met Arg Arg Ala Ser
755 760 765
Leu Lys Lys Glu Glu Leu Val Val His Pro Ala Asn Ser Pro Ile Ala
770 775 780
Asn Lys Asn Pro Asp Asn Pro Lys Lys Thr Thr Thr Leu Ser Tyr Asp
785 790 795 800
Val Tyr Lys Asp Lys Arg Phe Ser Glu Asp Gln Tyr Glu Leu His Ile
805 810 815
Pro Ile Ala Ile Asn Lys Cys Pro Lys Asn Ile Phe Lys Ile Asn Thr
820 825 830
Glu Val Arg Val Leu Leu Lys His Asp Asp Asn Pro Tyr Val Ile Gly
835 840 845
Ile Asp Arg Gly Glu Arg Asn Leu Leu Tyr Ile Val Val Val Asp Gly
850 855 860
Lys Gly Asn Ile Val Glu Gln Tyr Ser Leu Asn Glu Ile Ile Asn Asn
865 870 875 880
Phe Asn Gly Ile Arg Ile Lys Thr Asp Tyr His Ser Leu Leu Asp Lys
885 890 895
Lys Glu Lys Glu Arg Phe Glu Ala Arg Gln Asn Trp Thr Ser Ile Glu
900 905 910
Asn Ile Lys Glu Leu Lys Ala Gly Tyr Ile Ser Gln Val Val His Lys
915 920 925
Ile Cys Glu Leu Val Glu Lys Tyr Asp Ala Val Ile Ala Leu Glu Asp
930 935 940
Leu Asn Ser Gly Phe Lys Asn Ser Arg Val Lys Val Glu Lys Gln Val
945 950 955 960
Tyr Gln Lys Phe Glu Lys Met Leu Ile Asp Lys Leu Asn Tyr Met Val
965 970 975
Asp Lys Lys Ser Asn Pro Cys Ala Thr Gly Gly Ala Leu Lys Gly Tyr
980 985 990
Gln Ile Thr Asn Lys Phe Glu Ser Phe Lys Ser Met Ser Thr Gln Asn
995 1000 1005
Gly Phe Ile Phe Tyr Ile Pro Ala Trp Leu Thr Ser Lys Ile Asp
1010 1015 1020
Pro Ser Thr Gly Phe Val Asn Leu Leu Lys Thr Lys Tyr Thr Ser
1025 1030 1035
Ile Ala Asp Ser Lys Lys Phe Ile Ser Ser Phe Asp Arg Ile Met
1040 1045 1050
Tyr Val Pro Glu Glu Asp Leu Phe Glu Phe Ala Leu Asp Tyr Lys
1055 1060 1065
Asn Phe Ser Arg Thr Asp Ala Asp Tyr Ile Lys Lys Trp Lys Leu
1070 1075 1080
Tyr Ser Tyr Gly Asn Arg Ile Arg Ile Phe Arg Asn Pro Lys Lys
1085 1090 1095
Asn Asn Val Phe Asp Trp Glu Glu Val Cys Leu Thr Ser Ala Tyr
1100 1105 1110
Lys Glu Leu Phe Asn Lys Tyr Gly Ile Asn Tyr Gln Gln Gly Asp
1115 1120 1125
Ile Arg Ala Leu Leu Cys Glu Gln Ser Asp Lys Ala Phe Tyr Ser
1130 1135 1140
Ser Phe Met Ala Leu Met Ser Leu Met Leu Gln Met Ala Asn Ser
1145 1150 1155
Ile Thr Gly Arg Thr Asp Val Asp Phe Leu Ile Ser Pro Val Lys
1160 1165 1170
Asn Ser Asp Gly Ile Phe Tyr Asp Ser Arg Asn Tyr Glu Ala Gln
1175 1180 1185
Glu Asn Ala Ile Leu Pro Lys Asn Ala Asp Ala Asn Gly Ala Tyr
1190 1195 1200
Asn Ile Ala Arg Lys Val Leu Trp Ala Ile Gly Gln Phe Lys Lys
1205 1210 1215
Ala Glu Asp Glu Lys Leu Asp Lys Val Lys Ile Ala Ile Ser Asn
1220 1225 1230
Lys Glu Trp Leu Glu Tyr Ala Gln Thr Ser Val Lys His Ser Gly
1235 1240 1245
Gly Ser Ser Gly Gly Ser Ser Gly Ser Glu Thr Pro Gly Thr Ser
1250 1255 1260
Glu Ser Ala Thr Pro Glu Ser Ser Gly Gly Ser Ser Gly Gly Ser
1265 1270 1275
Thr Leu Asn Ile Glu Asp Glu Tyr Arg Leu His Glu Thr Ser Lys
1280 1285 1290
Glu Pro Asp Val Ser Leu Gly Ser Thr Trp Leu Ser Asp Phe Pro
1295 1300 1305
Gln Ala Trp Ala Glu Thr Gly Gly Met Gly Leu Ala Val Arg Gln
1310 1315 1320
Ala Pro Leu Ile Ile Pro Leu Lys Ala Thr Ser Thr Pro Val Ser
1325 1330 1335
Ile Lys Gln Tyr Pro Met Ser Gln Glu Ala Arg Leu Gly Ile Lys
1340 1345 1350
Pro His Ile Gln Arg Leu Leu Asp Gln Gly Ile Leu Val Pro Cys
1355 1360 1365
Gln Ser Pro Trp Asn Thr Pro Leu Leu Pro Val Lys Lys Pro Gly
1370 1375 1380
Thr Asn Asp Tyr Arg Pro Val Gln Asp Leu Arg Glu Val Asn Lys
1385 1390 1395
Arg Val Glu Asp Ile His Pro Thr Val Pro Asn Pro Tyr Asn Leu
1400 1405 1410
Leu Ser Gly Leu Pro Pro Ser His Gln Trp Tyr Thr Val Leu Asp
1415 1420 1425
Leu Lys Asp Ala Phe Phe Cys Leu Arg Leu His Pro Thr Ser Gln
1430 1435 1440
Pro Leu Phe Ala Phe Glu Trp Arg Asp Pro Glu Met Gly Ile Ser
1445 1450 1455
Gly Gln Leu Thr Trp Thr Arg Leu Pro Gln Gly Phe Lys Asn Ser
1460 1465 1470
Pro Thr Leu Phe Asn Glu Ala Leu His Arg Asp Leu Ala Asp Phe
1475 1480 1485
Arg Ile Gln His Pro Asp Leu Ile Leu Leu Gln Tyr Val Asp Asp
1490 1495 1500
Leu Leu Leu Ala Ala Thr Ser Glu Leu Asp Cys Gln Gln Gly Thr
1505 1510 1515
Arg Ala Leu Leu Gln Thr Leu Gly Asn Leu Gly Tyr Arg Ala Ser
1520 1525 1530
Ala Lys Lys Ala Gln Ile Cys Gln Lys Gln Val Lys Tyr Leu Gly
1535 1540 1545
Tyr Leu Leu Lys Glu Gly Gln Arg Trp Leu Thr Glu Ala Arg Lys
1550 1555 1560
Glu Thr Val Met Gly Gln Pro Thr Pro Lys Thr Pro Arg Gln Leu
1565 1570 1575
Arg Glu Phe Leu Gly Lys Ala Gly Phe Cys Arg Leu Phe Ile Pro
1580 1585 1590
Gly Phe Ala Glu Met Ala Ala Pro Leu Tyr Pro Leu Thr Lys Pro
1595 1600 1605
Gly Thr Leu Phe Asn Trp Gly Pro Asp Gln Gln Lys Ala Tyr Gln
1610 1615 1620
Glu Ile Lys Gln Ala Leu Leu Thr Ala Pro Ala Leu Gly Leu Pro
1625 1630 1635
Asp Leu Thr Lys Pro Phe Glu Leu Phe Val Asp Glu Lys Gln Gly
1640 1645 1650
Tyr Ala Lys Gly Val Leu Thr Gln Lys Leu Gly Pro Trp Arg Arg
1655 1660 1665
Pro Val Ala Tyr Leu Ser Lys Lys Leu Asp Pro Val Ala Ala Gly
1670 1675 1680
Trp Pro Pro Cys Leu Arg Met Val Ala Ala Ile Ala Val Leu Thr
1685 1690 1695
Lys Asp Ala Gly Lys Leu Thr Met Gly Gln Pro Leu Val Ile Leu
1700 1705 1710
Ala Pro His Ala Val Glu Ala Leu Val Lys Gln Pro Pro Asp Arg
1715 1720 1725
Trp Leu Ser Asn Ala Arg Met Thr His Tyr Gln Ala Leu Leu Leu
1730 1735 1740
Asp Thr Asp Arg Val Gln Phe Gly Pro Val Val Ala Leu Asn Pro
1745 1750 1755
Ala Thr Leu Leu Pro Leu Pro Glu Glu Gly Leu Gln His Asn Cys
1760 1765 1770
Leu Asp Ile Leu Ala Glu Ala His Gly Thr Arg Pro Asp Leu Thr
1775 1780 1785
Asp Gln Pro Leu Pro Asp Ala Asp His Thr Trp Tyr Thr Asp Gly
1790 1795 1800
Ser Ser Leu Leu Gln Glu Gly Gln Arg Lys Ala Gly Ala Ala Val
1805 1810 1815
Thr Thr Glu Thr Glu Val Ile Trp Ala Lys Ala Leu Pro Ala Gly
1820 1825 1830
Thr Ser Ala Gln Arg Ala Glu Leu Ile Ala Leu Thr Gln Ala Leu
1835 1840 1845
Lys Met Ala Glu Gly Lys Lys Leu Asn Val Tyr Thr Asp Ser Arg
1850 1855 1860
Tyr Ala Phe Ala Thr Ala His Ile His Gly Glu Ile Tyr Arg Arg
1865 1870 1875
Arg Gly Trp Leu Thr Ser Glu Gly Lys Glu Ile Lys Asn Lys Asp
1880 1885 1890
Glu Ile Leu Ala Leu Leu Lys Ala Leu Phe Leu Pro Lys Arg Leu
1895 1900 1905
Ser Ile Ile His Cys Pro Gly His Gln Lys Gly His Ser Ala Glu
1910 1915 1920
Ala Arg Gly Asn Arg Met Ala Asp Gln Ala Ala Arg Lys Ala Ala
1925 1930 1935
Ile Thr Glu Thr Pro Asp Thr Ser Thr Leu Leu Ile Glu Asn Ser
1940 1945 1950
Ser Pro Ser Gly Gly Ser Lys Arg Thr Ala Asp Gly Ser Glu Phe
1955 1960 1965
Glu Pro Lys Lys Lys Arg Lys Val Gly Ser Gly
1970 1975
<210> 71
<211> 1963
<212> PRT
<213> 人工的
<220>
<223> 合成的REDRAW编辑器
<400> 71
Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys
1 5 10 15
Arg Lys Val Thr Leu Asn Ile Glu Asp Glu Tyr Arg Leu His Glu Thr
20 25 30
Ser Lys Glu Pro Asp Val Ser Leu Gly Ser Thr Trp Leu Ser Asp Phe
35 40 45
Pro Gln Ala Trp Ala Glu Thr Gly Gly Met Gly Leu Ala Val Arg Gln
50 55 60
Ala Pro Leu Ile Ile Pro Leu Lys Ala Thr Ser Thr Pro Val Ser Ile
65 70 75 80
Lys Gln Tyr Pro Met Ser Gln Glu Ala Arg Leu Gly Ile Lys Pro His
85 90 95
Ile Gln Arg Leu Leu Asp Gln Gly Ile Leu Val Pro Cys Gln Ser Pro
100 105 110
Trp Asn Thr Pro Leu Leu Pro Val Lys Lys Pro Gly Thr Asn Asp Tyr
115 120 125
Arg Pro Val Gln Asp Leu Arg Glu Val Asn Lys Arg Val Glu Asp Ile
130 135 140
His Pro Thr Val Pro Asn Pro Tyr Asn Leu Leu Ser Gly Leu Pro Pro
145 150 155 160
Ser His Gln Trp Tyr Thr Val Leu Asp Leu Lys Asp Ala Phe Phe Cys
165 170 175
Leu Arg Leu His Pro Thr Ser Gln Pro Leu Phe Ala Phe Glu Trp Arg
180 185 190
Asp Pro Glu Met Gly Ile Ser Gly Gln Leu Thr Trp Thr Arg Leu Pro
195 200 205
Gln Gly Phe Lys Asn Ser Pro Thr Leu Phe Asn Glu Ala Leu His Arg
210 215 220
Asp Leu Ala Asp Phe Arg Ile Gln His Pro Asp Leu Ile Leu Leu Gln
225 230 235 240
Tyr Val Asp Asp Leu Leu Leu Ala Ala Thr Ser Glu Leu Asp Cys Gln
245 250 255
Gln Gly Thr Arg Ala Leu Leu Gln Thr Leu Gly Asn Leu Gly Tyr Arg
260 265 270
Ala Ser Ala Lys Lys Ala Gln Ile Cys Gln Lys Gln Val Lys Tyr Leu
275 280 285
Gly Tyr Leu Leu Lys Glu Gly Gln Arg Trp Leu Thr Glu Ala Arg Lys
290 295 300
Glu Thr Val Met Gly Gln Pro Thr Pro Lys Thr Pro Arg Gln Leu Arg
305 310 315 320
Glu Phe Leu Gly Lys Ala Gly Phe Cys Arg Leu Phe Ile Pro Gly Phe
325 330 335
Ala Glu Met Ala Ala Pro Leu Tyr Pro Leu Thr Lys Pro Gly Thr Leu
340 345 350
Phe Asn Trp Gly Pro Asp Gln Gln Lys Ala Tyr Gln Glu Ile Lys Gln
355 360 365
Ala Leu Leu Thr Ala Pro Ala Leu Gly Leu Pro Asp Leu Thr Lys Pro
370 375 380
Phe Glu Leu Phe Val Asp Glu Lys Gln Gly Tyr Ala Lys Gly Val Leu
385 390 395 400
Thr Gln Lys Leu Gly Pro Trp Arg Arg Pro Val Ala Tyr Leu Ser Lys
405 410 415
Lys Leu Asp Pro Val Ala Ala Gly Trp Pro Pro Cys Leu Arg Met Val
420 425 430
Ala Ala Ile Ala Val Leu Thr Lys Asp Ala Gly Lys Leu Thr Met Gly
435 440 445
Gln Pro Leu Val Ile Leu Ala Pro His Ala Val Glu Ala Leu Val Lys
450 455 460
Gln Pro Pro Asp Arg Trp Leu Ser Asn Ala Arg Met Thr His Tyr Gln
465 470 475 480
Ala Leu Leu Leu Asp Thr Asp Arg Val Gln Phe Gly Pro Val Val Ala
485 490 495
Leu Asn Pro Ala Thr Leu Leu Pro Leu Pro Glu Glu Gly Leu Gln His
500 505 510
Asn Cys Leu Asp Ile Leu Ala Glu Ala His Gly Thr Arg Pro Asp Leu
515 520 525
Thr Asp Gln Pro Leu Pro Asp Ala Asp His Thr Trp Tyr Thr Asp Gly
530 535 540
Ser Ser Leu Leu Gln Glu Gly Gln Arg Lys Ala Gly Ala Ala Val Thr
545 550 555 560
Thr Glu Thr Glu Val Ile Trp Ala Lys Ala Leu Pro Ala Gly Thr Ser
565 570 575
Ala Gln Arg Ala Glu Leu Ile Ala Leu Thr Gln Ala Leu Lys Met Ala
580 585 590
Glu Gly Lys Lys Leu Asn Val Tyr Thr Asp Ser Arg Tyr Ala Phe Ala
595 600 605
Thr Ala His Ile His Gly Glu Ile Tyr Arg Arg Arg Gly Trp Leu Thr
610 615 620
Ser Glu Gly Lys Glu Ile Lys Asn Lys Asp Glu Ile Leu Ala Leu Leu
625 630 635 640
Lys Ala Leu Phe Leu Pro Lys Arg Leu Ser Ile Ile His Cys Pro Gly
645 650 655
His Gln Lys Gly His Ser Ala Glu Ala Arg Gly Asn Arg Met Ala Asp
660 665 670
Gln Ala Ala Arg Lys Ala Ala Ile Thr Glu Thr Pro Asp Thr Ser Thr
675 680 685
Leu Leu Ile Glu Asn Ser Ser Pro Ser Gly Ser Glu Thr Pro Gly Thr
690 695 700
Ser Glu Ser Ala Thr Pro Glu Ser Ser Lys Leu Glu Lys Phe Thr Asn
705 710 715 720
Cys Tyr Ser Leu Ser Lys Thr Leu Arg Phe Lys Ala Ile Pro Val Gly
725 730 735
Lys Thr Gln Glu Asn Ile Asp Asn Lys Arg Leu Leu Val Glu Asp Glu
740 745 750
Lys Arg Ala Glu Asp Tyr Lys Gly Val Lys Lys Leu Leu Asp Arg Tyr
755 760 765
Tyr Leu Ser Phe Ile Asn Asp Val Leu His Ser Ile Lys Leu Lys Asn
770 775 780
Leu Asn Asn Tyr Ile Ser Leu Phe Arg Lys Lys Thr Arg Thr Glu Lys
785 790 795 800
Glu Asn Lys Glu Leu Glu Asn Leu Glu Ile Asn Leu Arg Lys Glu Ile
805 810 815
Ala Lys Ala Phe Lys Gly Asn Glu Gly Tyr Lys Ser Leu Phe Lys Lys
820 825 830
Asp Ile Ile Glu Thr Ile Leu Pro Glu Phe Leu Asp Asp Lys Asp Glu
835 840 845
Ile Ala Leu Val Asn Ser Phe Asn Gly Phe Thr Thr Ala Phe Thr Gly
850 855 860
Phe Phe Asp Asn Arg Glu Asn Met Phe Ser Glu Glu Ala Lys Ser Thr
865 870 875 880
Ser Ile Ala Phe Arg Cys Ile Asn Glu Asn Leu Thr Arg Tyr Ile Ser
885 890 895
Asn Met Asp Ile Phe Glu Lys Val Asp Ala Ile Phe Asp Lys His Glu
900 905 910
Val Gln Glu Ile Lys Glu Lys Ile Leu Asn Ser Asp Tyr Asp Val Glu
915 920 925
Asp Phe Phe Glu Gly Glu Phe Phe Asn Phe Val Leu Thr Gln Glu Gly
930 935 940
Ile Asp Val Tyr Asn Ala Ile Ile Gly Gly Phe Val Thr Glu Ser Gly
945 950 955 960
Glu Lys Ile Lys Gly Leu Asn Glu Tyr Ile Asn Leu Tyr Asn Gln Lys
965 970 975
Thr Lys Gln Lys Leu Pro Lys Phe Lys Pro Leu Tyr Lys Gln Val Leu
980 985 990
Ser Asp Arg Glu Ser Leu Ser Phe Tyr Gly Glu Gly Tyr Thr Ser Asp
995 1000 1005
Glu Glu Val Leu Glu Val Phe Arg Asn Thr Leu Asn Lys Asn Ser
1010 1015 1020
Glu Ile Phe Ser Ser Ile Lys Lys Leu Glu Lys Leu Phe Lys Asn
1025 1030 1035
Phe Asp Glu Tyr Ser Ser Ala Gly Ile Phe Val Lys Asn Gly Pro
1040 1045 1050
Ala Ile Ser Thr Ile Ser Lys Asp Ile Phe Gly Glu Trp Asn Val
1055 1060 1065
Ile Arg Asp Lys Trp Asn Ala Glu Tyr Asp Asp Ile His Leu Lys
1070 1075 1080
Lys Lys Ala Val Val Thr Glu Lys Tyr Glu Asp Asp Arg Arg Lys
1085 1090 1095
Ser Phe Lys Lys Ile Gly Ser Phe Ser Leu Glu Gln Leu Gln Glu
1100 1105 1110
Tyr Ala Asp Ala Asp Leu Ser Val Val Glu Lys Leu Lys Glu Ile
1115 1120 1125
Ile Ile Gln Lys Val Asp Glu Ile Tyr Lys Val Tyr Gly Ser Ser
1130 1135 1140
Glu Lys Leu Phe Asp Ala Asp Phe Val Leu Glu Lys Ser Leu Lys
1145 1150 1155
Lys Asn Asp Ala Val Val Ala Ile Met Lys Asp Leu Leu Asp Ser
1160 1165 1170
Val Lys Ser Phe Glu Asn Tyr Ile Lys Ala Phe Phe Gly Glu Gly
1175 1180 1185
Lys Glu Thr Asn Arg Asp Glu Ser Phe Tyr Gly Asp Phe Val Leu
1190 1195 1200
Ala Tyr Asp Ile Leu Leu Lys Val Asp His Ile Tyr Asp Ala Ile
1205 1210 1215
Arg Asn Tyr Val Thr Gln Lys Pro Tyr Ser Lys Asp Lys Phe Lys
1220 1225 1230
Leu Tyr Phe Gln Asn Pro Gln Phe Met Gly Gly Trp Asp Lys Asp
1235 1240 1245
Lys Glu Thr Asp Tyr Arg Ala Thr Ile Leu Arg Tyr Gly Ser Lys
1250 1255 1260
Tyr Tyr Leu Ala Ile Met Asp Lys Lys Tyr Ala Lys Cys Leu Gln
1265 1270 1275
Lys Ile Asp Lys Asp Asp Val Asn Gly Asn Tyr Glu Lys Ile Asn
1280 1285 1290
Tyr Lys Leu Leu Pro Gly Pro Asn Lys Met Leu Pro Lys Val Phe
1295 1300 1305
Phe Ser Lys Lys Trp Met Ala Tyr Tyr Asn Pro Ser Glu Asp Ile
1310 1315 1320
Gln Lys Ile Tyr Lys Asn Gly Thr Phe Lys Lys Gly Asp Met Phe
1325 1330 1335
Asn Leu Asn Asp Cys His Lys Leu Ile Asp Phe Phe Lys Asp Ser
1340 1345 1350
Ile Ser Arg Tyr Pro Lys Trp Ser Asn Ala Tyr Asp Phe Asn Phe
1355 1360 1365
Ser Glu Thr Glu Lys Tyr Lys Asp Ile Ala Gly Phe Tyr Arg Glu
1370 1375 1380
Val Glu Glu Gln Gly Tyr Lys Val Ser Phe Glu Ser Ala Ser Lys
1385 1390 1395
Lys Glu Val Asp Lys Leu Val Glu Glu Gly Lys Leu Tyr Met Phe
1400 1405 1410
Gln Ile Tyr Asn Lys Asp Phe Ser Asp Lys Ser His Gly Thr Pro
1415 1420 1425
Asn Leu His Thr Met Tyr Phe Lys Leu Leu Phe Asp Glu Asn Asn
1430 1435 1440
His Gly Gln Ile Arg Leu Ser Gly Gly Ala Glu Leu Phe Met Arg
1445 1450 1455
Arg Ala Ser Leu Lys Lys Glu Glu Leu Val Val His Pro Ala Asn
1460 1465 1470
Ser Pro Ile Ala Asn Lys Asn Pro Asp Asn Pro Lys Lys Thr Thr
1475 1480 1485
Thr Leu Ser Tyr Asp Val Tyr Lys Asp Lys Arg Phe Ser Glu Asp
1490 1495 1500
Gln Tyr Glu Leu His Ile Pro Ile Ala Ile Asn Lys Cys Pro Lys
1505 1510 1515
Asn Ile Phe Lys Ile Asn Thr Glu Val Arg Val Leu Leu Lys His
1520 1525 1530
Asp Asp Asn Pro Tyr Val Ile Gly Ile Asp Arg Gly Glu Arg Asn
1535 1540 1545
Leu Leu Tyr Ile Val Val Val Asp Gly Lys Gly Asn Ile Val Glu
1550 1555 1560
Gln Tyr Ser Leu Asn Glu Ile Ile Asn Asn Phe Asn Gly Ile Arg
1565 1570 1575
Ile Lys Thr Asp Tyr His Ser Leu Leu Asp Lys Lys Glu Lys Glu
1580 1585 1590
Arg Phe Glu Ala Arg Gln Asn Trp Thr Ser Ile Glu Asn Ile Lys
1595 1600 1605
Glu Leu Lys Ala Gly Tyr Ile Ser Gln Val Val His Lys Ile Cys
1610 1615 1620
Glu Leu Val Glu Lys Tyr Asp Ala Val Ile Ala Leu Glu Asp Leu
1625 1630 1635
Asn Ser Gly Phe Lys Asn Ser Arg Val Lys Val Glu Lys Gln Val
1640 1645 1650
Tyr Gln Lys Phe Glu Lys Met Leu Ile Asp Lys Leu Asn Tyr Met
1655 1660 1665
Val Asp Lys Lys Ser Asn Pro Cys Ala Thr Gly Gly Ala Leu Lys
1670 1675 1680
Gly Tyr Gln Ile Thr Asn Lys Phe Glu Ser Phe Lys Ser Met Ser
1685 1690 1695
Thr Gln Asn Gly Phe Ile Phe Tyr Ile Pro Ala Trp Leu Thr Ser
1700 1705 1710
Lys Ile Asp Pro Ser Thr Gly Phe Val Asn Leu Leu Lys Thr Lys
1715 1720 1725
Tyr Thr Ser Ile Ala Asp Ser Lys Lys Phe Ile Ser Ser Phe Asp
1730 1735 1740
Arg Ile Met Tyr Val Pro Glu Glu Asp Leu Phe Glu Phe Ala Leu
1745 1750 1755
Asp Tyr Lys Asn Phe Ser Arg Thr Asp Ala Asp Tyr Ile Lys Lys
1760 1765 1770
Trp Lys Leu Tyr Ser Tyr Gly Asn Arg Ile Arg Ile Phe Arg Asn
1775 1780 1785
Pro Lys Lys Asn Asn Val Phe Asp Trp Glu Glu Val Cys Leu Thr
1790 1795 1800
Ser Ala Tyr Lys Glu Leu Phe Asn Lys Tyr Gly Ile Asn Tyr Gln
1805 1810 1815
Gln Gly Asp Ile Arg Ala Leu Leu Cys Glu Gln Ser Asp Lys Ala
1820 1825 1830
Phe Tyr Ser Ser Phe Met Ala Leu Met Ser Leu Met Leu Gln Met
1835 1840 1845
Ala Asn Ser Ile Thr Gly Arg Thr Asp Val Asp Phe Leu Ile Ser
1850 1855 1860
Pro Val Lys Asn Ser Asp Gly Ile Phe Tyr Asp Ser Arg Asn Tyr
1865 1870 1875
Glu Ala Gln Glu Asn Ala Ile Leu Pro Lys Asn Ala Asp Ala Asn
1880 1885 1890
Gly Ala Tyr Asn Ile Ala Arg Lys Val Leu Trp Ala Ile Gly Gln
1895 1900 1905
Phe Lys Lys Ala Glu Asp Glu Lys Leu Asp Lys Val Lys Ile Ala
1910 1915 1920
Ile Ser Asn Lys Glu Trp Leu Glu Tyr Ala Gln Thr Ser Val Lys
1925 1930 1935
His Ser Gly Gly Ser Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu
1940 1945 1950
Pro Lys Lys Lys Arg Lys Val Gly Ser Gly
1955 1960
<210> 72
<211> 1979
<212> PRT
<213> 人工的
<220>
<223> 合成的REDRAW编辑器
<400> 72
Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys
1 5 10 15
Arg Lys Val Thr Leu Asn Ile Glu Asp Glu Tyr Arg Leu His Glu Thr
20 25 30
Ser Lys Glu Pro Asp Val Ser Leu Gly Ser Thr Trp Leu Ser Asp Phe
35 40 45
Pro Gln Ala Trp Ala Glu Thr Gly Gly Met Gly Leu Ala Val Arg Gln
50 55 60
Ala Pro Leu Ile Ile Pro Leu Lys Ala Thr Ser Thr Pro Val Ser Ile
65 70 75 80
Lys Gln Tyr Pro Met Ser Gln Glu Ala Arg Leu Gly Ile Lys Pro His
85 90 95
Ile Gln Arg Leu Leu Asp Gln Gly Ile Leu Val Pro Cys Gln Ser Pro
100 105 110
Trp Asn Thr Pro Leu Leu Pro Val Lys Lys Pro Gly Thr Asn Asp Tyr
115 120 125
Arg Pro Val Gln Asp Leu Arg Glu Val Asn Lys Arg Val Glu Asp Ile
130 135 140
His Pro Thr Val Pro Asn Pro Tyr Asn Leu Leu Ser Gly Leu Pro Pro
145 150 155 160
Ser His Gln Trp Tyr Thr Val Leu Asp Leu Lys Asp Ala Phe Phe Cys
165 170 175
Leu Arg Leu His Pro Thr Ser Gln Pro Leu Phe Ala Phe Glu Trp Arg
180 185 190
Asp Pro Glu Met Gly Ile Ser Gly Gln Leu Thr Trp Thr Arg Leu Pro
195 200 205
Gln Gly Phe Lys Asn Ser Pro Thr Leu Phe Asn Glu Ala Leu His Arg
210 215 220
Asp Leu Ala Asp Phe Arg Ile Gln His Pro Asp Leu Ile Leu Leu Gln
225 230 235 240
Tyr Val Asp Asp Leu Leu Leu Ala Ala Thr Ser Glu Leu Asp Cys Gln
245 250 255
Gln Gly Thr Arg Ala Leu Leu Gln Thr Leu Gly Asn Leu Gly Tyr Arg
260 265 270
Ala Ser Ala Lys Lys Ala Gln Ile Cys Gln Lys Gln Val Lys Tyr Leu
275 280 285
Gly Tyr Leu Leu Lys Glu Gly Gln Arg Trp Leu Thr Glu Ala Arg Lys
290 295 300
Glu Thr Val Met Gly Gln Pro Thr Pro Lys Thr Pro Arg Gln Leu Arg
305 310 315 320
Glu Phe Leu Gly Lys Ala Gly Phe Cys Arg Leu Phe Ile Pro Gly Phe
325 330 335
Ala Glu Met Ala Ala Pro Leu Tyr Pro Leu Thr Lys Pro Gly Thr Leu
340 345 350
Phe Asn Trp Gly Pro Asp Gln Gln Lys Ala Tyr Gln Glu Ile Lys Gln
355 360 365
Ala Leu Leu Thr Ala Pro Ala Leu Gly Leu Pro Asp Leu Thr Lys Pro
370 375 380
Phe Glu Leu Phe Val Asp Glu Lys Gln Gly Tyr Ala Lys Gly Val Leu
385 390 395 400
Thr Gln Lys Leu Gly Pro Trp Arg Arg Pro Val Ala Tyr Leu Ser Lys
405 410 415
Lys Leu Asp Pro Val Ala Ala Gly Trp Pro Pro Cys Leu Arg Met Val
420 425 430
Ala Ala Ile Ala Val Leu Thr Lys Asp Ala Gly Lys Leu Thr Met Gly
435 440 445
Gln Pro Leu Val Ile Leu Ala Pro His Ala Val Glu Ala Leu Val Lys
450 455 460
Gln Pro Pro Asp Arg Trp Leu Ser Asn Ala Arg Met Thr His Tyr Gln
465 470 475 480
Ala Leu Leu Leu Asp Thr Asp Arg Val Gln Phe Gly Pro Val Val Ala
485 490 495
Leu Asn Pro Ala Thr Leu Leu Pro Leu Pro Glu Glu Gly Leu Gln His
500 505 510
Asn Cys Leu Asp Ile Leu Ala Glu Ala His Gly Thr Arg Pro Asp Leu
515 520 525
Thr Asp Gln Pro Leu Pro Asp Ala Asp His Thr Trp Tyr Thr Asp Gly
530 535 540
Ser Ser Leu Leu Gln Glu Gly Gln Arg Lys Ala Gly Ala Ala Val Thr
545 550 555 560
Thr Glu Thr Glu Val Ile Trp Ala Lys Ala Leu Pro Ala Gly Thr Ser
565 570 575
Ala Gln Arg Ala Glu Leu Ile Ala Leu Thr Gln Ala Leu Lys Met Ala
580 585 590
Glu Gly Lys Lys Leu Asn Val Tyr Thr Asp Ser Arg Tyr Ala Phe Ala
595 600 605
Thr Ala His Ile His Gly Glu Ile Tyr Arg Arg Arg Gly Trp Leu Thr
610 615 620
Ser Glu Gly Lys Glu Ile Lys Asn Lys Asp Glu Ile Leu Ala Leu Leu
625 630 635 640
Lys Ala Leu Phe Leu Pro Lys Arg Leu Ser Ile Ile His Cys Pro Gly
645 650 655
His Gln Lys Gly His Ser Ala Glu Ala Arg Gly Asn Arg Met Ala Asp
660 665 670
Gln Ala Ala Arg Lys Ala Ala Ile Thr Glu Thr Pro Asp Thr Ser Thr
675 680 685
Leu Leu Ile Glu Asn Ser Ser Pro Ser Gly Gly Ser Ser Gly Gly Ser
690 695 700
Ser Gly Ser Glu Thr Pro Gly Thr Ser Glu Ser Ala Thr Pro Glu Ser
705 710 715 720
Ser Gly Gly Ser Ser Gly Gly Ser Ser Lys Leu Glu Lys Phe Thr Asn
725 730 735
Cys Tyr Ser Leu Ser Lys Thr Leu Arg Phe Lys Ala Ile Pro Val Gly
740 745 750
Lys Thr Gln Glu Asn Ile Asp Asn Lys Arg Leu Leu Val Glu Asp Glu
755 760 765
Lys Arg Ala Glu Asp Tyr Lys Gly Val Lys Lys Leu Leu Asp Arg Tyr
770 775 780
Tyr Leu Ser Phe Ile Asn Asp Val Leu His Ser Ile Lys Leu Lys Asn
785 790 795 800
Leu Asn Asn Tyr Ile Ser Leu Phe Arg Lys Lys Thr Arg Thr Glu Lys
805 810 815
Glu Asn Lys Glu Leu Glu Asn Leu Glu Ile Asn Leu Arg Lys Glu Ile
820 825 830
Ala Lys Ala Phe Lys Gly Asn Glu Gly Tyr Lys Ser Leu Phe Lys Lys
835 840 845
Asp Ile Ile Glu Thr Ile Leu Pro Glu Phe Leu Asp Asp Lys Asp Glu
850 855 860
Ile Ala Leu Val Asn Ser Phe Asn Gly Phe Thr Thr Ala Phe Thr Gly
865 870 875 880
Phe Phe Asp Asn Arg Glu Asn Met Phe Ser Glu Glu Ala Lys Ser Thr
885 890 895
Ser Ile Ala Phe Arg Cys Ile Asn Glu Asn Leu Thr Arg Tyr Ile Ser
900 905 910
Asn Met Asp Ile Phe Glu Lys Val Asp Ala Ile Phe Asp Lys His Glu
915 920 925
Val Gln Glu Ile Lys Glu Lys Ile Leu Asn Ser Asp Tyr Asp Val Glu
930 935 940
Asp Phe Phe Glu Gly Glu Phe Phe Asn Phe Val Leu Thr Gln Glu Gly
945 950 955 960
Ile Asp Val Tyr Asn Ala Ile Ile Gly Gly Phe Val Thr Glu Ser Gly
965 970 975
Glu Lys Ile Lys Gly Leu Asn Glu Tyr Ile Asn Leu Tyr Asn Gln Lys
980 985 990
Thr Lys Gln Lys Leu Pro Lys Phe Lys Pro Leu Tyr Lys Gln Val Leu
995 1000 1005
Ser Asp Arg Glu Ser Leu Ser Phe Tyr Gly Glu Gly Tyr Thr Ser
1010 1015 1020
Asp Glu Glu Val Leu Glu Val Phe Arg Asn Thr Leu Asn Lys Asn
1025 1030 1035
Ser Glu Ile Phe Ser Ser Ile Lys Lys Leu Glu Lys Leu Phe Lys
1040 1045 1050
Asn Phe Asp Glu Tyr Ser Ser Ala Gly Ile Phe Val Lys Asn Gly
1055 1060 1065
Pro Ala Ile Ser Thr Ile Ser Lys Asp Ile Phe Gly Glu Trp Asn
1070 1075 1080
Val Ile Arg Asp Lys Trp Asn Ala Glu Tyr Asp Asp Ile His Leu
1085 1090 1095
Lys Lys Lys Ala Val Val Thr Glu Lys Tyr Glu Asp Asp Arg Arg
1100 1105 1110
Lys Ser Phe Lys Lys Ile Gly Ser Phe Ser Leu Glu Gln Leu Gln
1115 1120 1125
Glu Tyr Ala Asp Ala Asp Leu Ser Val Val Glu Lys Leu Lys Glu
1130 1135 1140
Ile Ile Ile Gln Lys Val Asp Glu Ile Tyr Lys Val Tyr Gly Ser
1145 1150 1155
Ser Glu Lys Leu Phe Asp Ala Asp Phe Val Leu Glu Lys Ser Leu
1160 1165 1170
Lys Lys Asn Asp Ala Val Val Ala Ile Met Lys Asp Leu Leu Asp
1175 1180 1185
Ser Val Lys Ser Phe Glu Asn Tyr Ile Lys Ala Phe Phe Gly Glu
1190 1195 1200
Gly Lys Glu Thr Asn Arg Asp Glu Ser Phe Tyr Gly Asp Phe Val
1205 1210 1215
Leu Ala Tyr Asp Ile Leu Leu Lys Val Asp His Ile Tyr Asp Ala
1220 1225 1230
Ile Arg Asn Tyr Val Thr Gln Lys Pro Tyr Ser Lys Asp Lys Phe
1235 1240 1245
Lys Leu Tyr Phe Gln Asn Pro Gln Phe Met Gly Gly Trp Asp Lys
1250 1255 1260
Asp Lys Glu Thr Asp Tyr Arg Ala Thr Ile Leu Arg Tyr Gly Ser
1265 1270 1275
Lys Tyr Tyr Leu Ala Ile Met Asp Lys Lys Tyr Ala Lys Cys Leu
1280 1285 1290
Gln Lys Ile Asp Lys Asp Asp Val Asn Gly Asn Tyr Glu Lys Ile
1295 1300 1305
Asn Tyr Lys Leu Leu Pro Gly Pro Asn Lys Met Leu Pro Lys Val
1310 1315 1320
Phe Phe Ser Lys Lys Trp Met Ala Tyr Tyr Asn Pro Ser Glu Asp
1325 1330 1335
Ile Gln Lys Ile Tyr Lys Asn Gly Thr Phe Lys Lys Gly Asp Met
1340 1345 1350
Phe Asn Leu Asn Asp Cys His Lys Leu Ile Asp Phe Phe Lys Asp
1355 1360 1365
Ser Ile Ser Arg Tyr Pro Lys Trp Ser Asn Ala Tyr Asp Phe Asn
1370 1375 1380
Phe Ser Glu Thr Glu Lys Tyr Lys Asp Ile Ala Gly Phe Tyr Arg
1385 1390 1395
Glu Val Glu Glu Gln Gly Tyr Lys Val Ser Phe Glu Ser Ala Ser
1400 1405 1410
Lys Lys Glu Val Asp Lys Leu Val Glu Glu Gly Lys Leu Tyr Met
1415 1420 1425
Phe Gln Ile Tyr Asn Lys Asp Phe Ser Asp Lys Ser His Gly Thr
1430 1435 1440
Pro Asn Leu His Thr Met Tyr Phe Lys Leu Leu Phe Asp Glu Asn
1445 1450 1455
Asn His Gly Gln Ile Arg Leu Ser Gly Gly Ala Glu Leu Phe Met
1460 1465 1470
Arg Arg Ala Ser Leu Lys Lys Glu Glu Leu Val Val His Pro Ala
1475 1480 1485
Asn Ser Pro Ile Ala Asn Lys Asn Pro Asp Asn Pro Lys Lys Thr
1490 1495 1500
Thr Thr Leu Ser Tyr Asp Val Tyr Lys Asp Lys Arg Phe Ser Glu
1505 1510 1515
Asp Gln Tyr Glu Leu His Ile Pro Ile Ala Ile Asn Lys Cys Pro
1520 1525 1530
Lys Asn Ile Phe Lys Ile Asn Thr Glu Val Arg Val Leu Leu Lys
1535 1540 1545
His Asp Asp Asn Pro Tyr Val Ile Gly Ile Asp Arg Gly Glu Arg
1550 1555 1560
Asn Leu Leu Tyr Ile Val Val Val Asp Gly Lys Gly Asn Ile Val
1565 1570 1575
Glu Gln Tyr Ser Leu Asn Glu Ile Ile Asn Asn Phe Asn Gly Ile
1580 1585 1590
Arg Ile Lys Thr Asp Tyr His Ser Leu Leu Asp Lys Lys Glu Lys
1595 1600 1605
Glu Arg Phe Glu Ala Arg Gln Asn Trp Thr Ser Ile Glu Asn Ile
1610 1615 1620
Lys Glu Leu Lys Ala Gly Tyr Ile Ser Gln Val Val His Lys Ile
1625 1630 1635
Cys Glu Leu Val Glu Lys Tyr Asp Ala Val Ile Ala Leu Glu Asp
1640 1645 1650
Leu Asn Ser Gly Phe Lys Asn Ser Arg Val Lys Val Glu Lys Gln
1655 1660 1665
Val Tyr Gln Lys Phe Glu Lys Met Leu Ile Asp Lys Leu Asn Tyr
1670 1675 1680
Met Val Asp Lys Lys Ser Asn Pro Cys Ala Thr Gly Gly Ala Leu
1685 1690 1695
Lys Gly Tyr Gln Ile Thr Asn Lys Phe Glu Ser Phe Lys Ser Met
1700 1705 1710
Ser Thr Gln Asn Gly Phe Ile Phe Tyr Ile Pro Ala Trp Leu Thr
1715 1720 1725
Ser Lys Ile Asp Pro Ser Thr Gly Phe Val Asn Leu Leu Lys Thr
1730 1735 1740
Lys Tyr Thr Ser Ile Ala Asp Ser Lys Lys Phe Ile Ser Ser Phe
1745 1750 1755
Asp Arg Ile Met Tyr Val Pro Glu Glu Asp Leu Phe Glu Phe Ala
1760 1765 1770
Leu Asp Tyr Lys Asn Phe Ser Arg Thr Asp Ala Asp Tyr Ile Lys
1775 1780 1785
Lys Trp Lys Leu Tyr Ser Tyr Gly Asn Arg Ile Arg Ile Phe Arg
1790 1795 1800
Asn Pro Lys Lys Asn Asn Val Phe Asp Trp Glu Glu Val Cys Leu
1805 1810 1815
Thr Ser Ala Tyr Lys Glu Leu Phe Asn Lys Tyr Gly Ile Asn Tyr
1820 1825 1830
Gln Gln Gly Asp Ile Arg Ala Leu Leu Cys Glu Gln Ser Asp Lys
1835 1840 1845
Ala Phe Tyr Ser Ser Phe Met Ala Leu Met Ser Leu Met Leu Gln
1850 1855 1860
Met Ala Asn Ser Ile Thr Gly Arg Thr Asp Val Asp Phe Leu Ile
1865 1870 1875
Ser Pro Val Lys Asn Ser Asp Gly Ile Phe Tyr Asp Ser Arg Asn
1880 1885 1890
Tyr Glu Ala Gln Glu Asn Ala Ile Leu Pro Lys Asn Ala Asp Ala
1895 1900 1905
Asn Gly Ala Tyr Asn Ile Ala Arg Lys Val Leu Trp Ala Ile Gly
1910 1915 1920
Gln Phe Lys Lys Ala Glu Asp Glu Lys Leu Asp Lys Val Lys Ile
1925 1930 1935
Ala Ile Ser Asn Lys Glu Trp Leu Glu Tyr Ala Gln Thr Ser Val
1940 1945 1950
Lys His Ser Gly Gly Ser Lys Arg Thr Ala Asp Gly Ser Glu Phe
1955 1960 1965
Glu Pro Lys Lys Lys Arg Lys Val Gly Ser Gly
1970 1975
<210> 73
<211> 1963
<212> PRT
<213> 人工的
<220>
<223> 合成的REDRAW编辑器
<400> 73
Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys
1 5 10 15
Arg Lys Val Ser Lys Leu Glu Lys Phe Thr Asn Cys Tyr Ser Leu Ser
20 25 30
Lys Thr Leu Arg Phe Lys Ala Ile Pro Val Gly Lys Thr Gln Glu Asn
35 40 45
Ile Asp Asn Lys Arg Leu Leu Val Glu Asp Glu Lys Arg Ala Glu Asp
50 55 60
Tyr Lys Gly Val Lys Lys Leu Leu Asp Arg Tyr Tyr Leu Ser Phe Ile
65 70 75 80
Asn Asp Val Leu His Ser Ile Lys Leu Lys Asn Leu Asn Asn Tyr Ile
85 90 95
Ser Leu Phe Arg Lys Lys Thr Arg Thr Glu Lys Glu Asn Lys Glu Leu
100 105 110
Glu Asn Leu Glu Ile Asn Leu Arg Lys Glu Ile Ala Lys Ala Phe Lys
115 120 125
Gly Asn Glu Gly Tyr Lys Ser Leu Phe Lys Lys Asp Ile Ile Glu Thr
130 135 140
Ile Leu Pro Glu Phe Leu Asp Asp Lys Asp Glu Ile Ala Leu Val Asn
145 150 155 160
Ser Phe Asn Gly Phe Thr Thr Ala Phe Thr Gly Phe Phe Asp Asn Arg
165 170 175
Glu Asn Met Phe Ser Glu Glu Ala Lys Ser Thr Ser Ile Ala Phe Arg
180 185 190
Cys Ile Asn Glu Asn Leu Thr Arg Tyr Ile Ser Asn Met Asp Ile Phe
195 200 205
Glu Lys Val Asp Ala Ile Phe Asp Lys His Glu Val Gln Glu Ile Lys
210 215 220
Glu Lys Ile Leu Asn Ser Asp Tyr Asp Val Glu Asp Phe Phe Glu Gly
225 230 235 240
Glu Phe Phe Asn Phe Val Leu Thr Gln Glu Gly Ile Asp Val Tyr Asn
245 250 255
Ala Ile Ile Gly Gly Phe Val Thr Glu Ser Gly Glu Lys Ile Lys Gly
260 265 270
Leu Asn Glu Tyr Ile Asn Leu Tyr Asn Gln Lys Thr Lys Gln Lys Leu
275 280 285
Pro Lys Phe Lys Pro Leu Tyr Lys Gln Val Leu Ser Asp Arg Glu Ser
290 295 300
Leu Ser Phe Tyr Gly Glu Gly Tyr Thr Ser Asp Glu Glu Val Leu Glu
305 310 315 320
Val Phe Arg Asn Thr Leu Asn Lys Asn Ser Glu Ile Phe Ser Ser Ile
325 330 335
Lys Lys Leu Glu Lys Leu Phe Lys Asn Phe Asp Glu Tyr Ser Ser Ala
340 345 350
Gly Ile Phe Val Lys Asn Gly Pro Ala Ile Ser Thr Ile Ser Lys Asp
355 360 365
Ile Phe Gly Glu Trp Asn Val Ile Arg Asp Lys Trp Asn Ala Glu Tyr
370 375 380
Asp Asp Ile His Leu Lys Lys Lys Ala Val Val Thr Glu Lys Tyr Glu
385 390 395 400
Asp Asp Arg Arg Lys Ser Phe Lys Lys Ile Gly Ser Phe Ser Leu Glu
405 410 415
Gln Leu Gln Glu Tyr Ala Asp Ala Asp Leu Ser Val Val Glu Lys Leu
420 425 430
Lys Glu Ile Ile Ile Gln Lys Val Asp Glu Ile Tyr Lys Val Tyr Gly
435 440 445
Ser Ser Glu Lys Leu Phe Asp Ala Asp Phe Val Leu Glu Lys Ser Leu
450 455 460
Lys Lys Asn Asp Ala Val Val Ala Ile Met Lys Asp Leu Leu Asp Ser
465 470 475 480
Val Lys Ser Phe Glu Asn Tyr Ile Lys Ala Phe Phe Gly Glu Gly Lys
485 490 495
Glu Thr Asn Arg Asp Glu Ser Phe Tyr Gly Asp Phe Val Leu Ala Tyr
500 505 510
Asp Ile Leu Leu Lys Val Asp His Ile Tyr Asp Ala Ile Arg Asn Tyr
515 520 525
Val Thr Gln Lys Pro Tyr Ser Lys Asp Lys Phe Lys Leu Tyr Phe Gln
530 535 540
Asn Pro Gln Phe Met Gly Gly Trp Asp Lys Asp Lys Glu Thr Asp Tyr
545 550 555 560
Arg Ala Thr Ile Leu Arg Tyr Gly Ser Lys Tyr Tyr Leu Ala Ile Met
565 570 575
Asp Lys Lys Tyr Ala Lys Cys Leu Gln Lys Ile Asp Lys Asp Asp Val
580 585 590
Asn Gly Asn Tyr Glu Lys Ile Asn Tyr Lys Leu Leu Pro Gly Pro Asn
595 600 605
Lys Met Leu Pro Lys Val Phe Phe Ser Lys Lys Trp Met Ala Tyr Tyr
610 615 620
Asn Pro Ser Glu Asp Ile Gln Lys Ile Tyr Lys Asn Gly Thr Phe Lys
625 630 635 640
Lys Gly Asp Met Phe Asn Leu Asn Asp Cys His Lys Leu Ile Asp Phe
645 650 655
Phe Lys Asp Ser Ile Ser Arg Tyr Pro Lys Trp Ser Asn Ala Tyr Asp
660 665 670
Phe Asn Phe Ser Glu Thr Glu Lys Tyr Lys Asp Ile Ala Gly Phe Tyr
675 680 685
Arg Glu Val Glu Glu Gln Gly Tyr Lys Val Ser Phe Glu Ser Ala Ser
690 695 700
Lys Lys Glu Val Asp Lys Leu Val Glu Glu Gly Lys Leu Tyr Met Phe
705 710 715 720
Gln Ile Tyr Asn Lys Asp Phe Ser Asp Lys Ser His Gly Thr Pro Asn
725 730 735
Leu His Thr Met Tyr Phe Lys Leu Leu Phe Asp Glu Asn Asn His Gly
740 745 750
Gln Ile Arg Leu Ser Gly Gly Ala Glu Leu Phe Met Arg Arg Ala Ser
755 760 765
Leu Lys Lys Glu Glu Leu Val Val His Pro Ala Asn Ser Pro Ile Ala
770 775 780
Asn Lys Asn Pro Asp Asn Pro Lys Lys Thr Thr Thr Leu Ser Tyr Asp
785 790 795 800
Val Tyr Lys Asp Lys Arg Phe Ser Glu Asp Gln Tyr Glu Leu His Ile
805 810 815
Pro Ile Ala Ile Asn Lys Cys Pro Lys Asn Ile Phe Lys Ile Asn Thr
820 825 830
Glu Val Arg Val Leu Leu Lys His Asp Asp Asn Pro Tyr Val Ile Gly
835 840 845
Ile Asp Arg Gly Glu Arg Asn Leu Leu Tyr Ile Val Val Val Asp Gly
850 855 860
Lys Gly Asn Ile Val Glu Gln Tyr Ser Leu Asn Glu Ile Ile Asn Asn
865 870 875 880
Phe Asn Gly Ile Arg Ile Lys Thr Asp Tyr His Ser Leu Leu Asp Lys
885 890 895
Lys Glu Lys Glu Arg Phe Glu Ala Arg Gln Asn Trp Thr Ser Ile Glu
900 905 910
Asn Ile Lys Glu Leu Lys Ala Gly Tyr Ile Ser Gln Val Val His Lys
915 920 925
Ile Cys Glu Leu Val Glu Lys Tyr Asp Ala Val Ile Ala Leu Glu Asp
930 935 940
Leu Asn Ser Gly Phe Lys Asn Ser Arg Val Lys Val Glu Lys Gln Val
945 950 955 960
Tyr Gln Lys Phe Glu Lys Met Leu Ile Asp Lys Leu Asn Tyr Met Val
965 970 975
Asp Lys Lys Ser Asn Pro Cys Ala Thr Gly Gly Ala Leu Lys Gly Tyr
980 985 990
Gln Ile Thr Asn Lys Phe Glu Ser Phe Lys Ser Met Ser Thr Gln Asn
995 1000 1005
Gly Phe Ile Phe Tyr Ile Pro Ala Trp Leu Thr Ser Lys Ile Asp
1010 1015 1020
Pro Ser Thr Gly Phe Val Asn Leu Leu Lys Thr Lys Tyr Thr Ser
1025 1030 1035
Ile Ala Asp Ser Lys Lys Phe Ile Ser Ser Phe Asp Arg Ile Met
1040 1045 1050
Tyr Val Pro Glu Glu Asp Leu Phe Glu Phe Ala Leu Asp Tyr Lys
1055 1060 1065
Asn Phe Ser Arg Thr Asp Ala Asp Tyr Ile Lys Lys Trp Lys Leu
1070 1075 1080
Tyr Ser Tyr Gly Asn Arg Ile Arg Ile Phe Arg Asn Pro Lys Lys
1085 1090 1095
Asn Asn Val Phe Asp Trp Glu Glu Val Cys Leu Thr Ser Ala Tyr
1100 1105 1110
Lys Glu Leu Phe Asn Lys Tyr Gly Ile Asn Tyr Gln Gln Gly Asp
1115 1120 1125
Ile Arg Ala Leu Leu Cys Glu Gln Ser Asp Lys Ala Phe Tyr Ser
1130 1135 1140
Ser Phe Met Ala Leu Met Ser Leu Met Leu Gln Met Ala Asn Ser
1145 1150 1155
Ile Thr Gly Arg Thr Asp Val Asp Phe Leu Ile Ser Pro Val Lys
1160 1165 1170
Asn Ser Asp Gly Ile Phe Tyr Asp Ser Arg Asn Tyr Glu Ala Gln
1175 1180 1185
Glu Asn Ala Ile Leu Pro Lys Asn Ala Asp Ala Asn Gly Ala Tyr
1190 1195 1200
Asn Ile Ala Arg Lys Val Leu Trp Ala Ile Gly Gln Phe Lys Lys
1205 1210 1215
Ala Glu Asp Glu Lys Leu Asp Lys Val Lys Ile Ala Ile Ser Asn
1220 1225 1230
Lys Glu Trp Leu Glu Tyr Ala Gln Thr Ser Val Lys His Ser Gly
1235 1240 1245
Ser Glu Thr Pro Gly Thr Ser Glu Ser Ala Thr Pro Glu Ser Thr
1250 1255 1260
Leu Asn Ile Glu Asp Glu Tyr Arg Leu His Glu Thr Ser Lys Glu
1265 1270 1275
Pro Asp Val Ser Leu Gly Ser Thr Trp Leu Ser Asp Phe Pro Gln
1280 1285 1290
Ala Trp Ala Glu Thr Gly Gly Met Gly Leu Ala Val Arg Gln Ala
1295 1300 1305
Pro Leu Ile Ile Pro Leu Lys Ala Thr Ser Thr Pro Val Ser Ile
1310 1315 1320
Lys Gln Tyr Pro Met Ser Gln Glu Ala Arg Leu Gly Ile Lys Pro
1325 1330 1335
His Ile Gln Arg Leu Leu Asp Gln Gly Ile Leu Val Pro Cys Gln
1340 1345 1350
Ser Pro Trp Asn Thr Pro Leu Leu Pro Val Lys Lys Pro Gly Thr
1355 1360 1365
Asn Asp Tyr Arg Pro Val Gln Asp Leu Arg Glu Val Asn Lys Arg
1370 1375 1380
Val Glu Asp Ile His Pro Thr Val Pro Asn Pro Tyr Asn Leu Leu
1385 1390 1395
Ser Gly Leu Pro Pro Ser His Gln Trp Tyr Thr Val Leu Asp Leu
1400 1405 1410
Lys Asp Ala Phe Phe Cys Leu Arg Leu His Pro Thr Ser Gln Pro
1415 1420 1425
Leu Phe Ala Phe Glu Trp Arg Asp Pro Glu Met Gly Ile Ser Gly
1430 1435 1440
Gln Leu Thr Trp Thr Arg Leu Pro Gln Gly Phe Lys Asn Ser Pro
1445 1450 1455
Thr Leu Phe Asn Glu Ala Leu His Arg Asp Leu Ala Asp Phe Arg
1460 1465 1470
Ile Gln His Pro Asp Leu Ile Leu Leu Gln Tyr Val Asp Asp Leu
1475 1480 1485
Leu Leu Ala Ala Thr Ser Glu Leu Asp Cys Gln Gln Gly Thr Arg
1490 1495 1500
Ala Leu Leu Gln Thr Leu Gly Asn Leu Gly Tyr Arg Ala Ser Ala
1505 1510 1515
Lys Lys Ala Gln Ile Cys Gln Lys Gln Val Lys Tyr Leu Gly Tyr
1520 1525 1530
Leu Leu Lys Glu Gly Gln Arg Trp Leu Thr Glu Ala Arg Lys Glu
1535 1540 1545
Thr Val Met Gly Gln Pro Thr Pro Lys Thr Pro Arg Gln Leu Arg
1550 1555 1560
Glu Phe Leu Gly Lys Ala Gly Phe Cys Arg Leu Phe Ile Pro Gly
1565 1570 1575
Phe Ala Glu Met Ala Ala Pro Leu Tyr Pro Leu Thr Lys Pro Gly
1580 1585 1590
Thr Leu Phe Asn Trp Gly Pro Asp Gln Gln Lys Ala Tyr Gln Glu
1595 1600 1605
Ile Lys Gln Ala Leu Leu Thr Ala Pro Ala Leu Gly Leu Pro Asp
1610 1615 1620
Leu Thr Lys Pro Phe Glu Leu Phe Val Asp Glu Lys Gln Gly Tyr
1625 1630 1635
Ala Lys Gly Val Leu Thr Gln Lys Leu Gly Pro Trp Arg Arg Pro
1640 1645 1650
Val Ala Tyr Leu Ser Lys Lys Leu Asp Pro Val Ala Ala Gly Trp
1655 1660 1665
Pro Pro Cys Leu Arg Met Val Ala Ala Ile Ala Val Leu Thr Lys
1670 1675 1680
Asp Ala Gly Lys Leu Thr Met Gly Gln Pro Leu Val Ile Leu Ala
1685 1690 1695
Pro His Ala Val Glu Ala Leu Val Lys Gln Pro Pro Asp Arg Trp
1700 1705 1710
Leu Ser Asn Ala Arg Met Thr His Tyr Gln Ala Leu Leu Leu Asp
1715 1720 1725
Thr Asp Arg Val Gln Phe Gly Pro Val Val Ala Leu Asn Pro Ala
1730 1735 1740
Thr Leu Leu Pro Leu Pro Glu Glu Gly Leu Gln His Asn Cys Leu
1745 1750 1755
Asp Ile Leu Ala Glu Ala His Gly Thr Arg Pro Asp Leu Thr Asp
1760 1765 1770
Gln Pro Leu Pro Asp Ala Asp His Thr Trp Tyr Thr Asp Gly Ser
1775 1780 1785
Ser Leu Leu Gln Glu Gly Gln Arg Lys Ala Gly Ala Ala Val Thr
1790 1795 1800
Thr Glu Thr Glu Val Ile Trp Ala Lys Ala Leu Pro Ala Gly Thr
1805 1810 1815
Ser Ala Gln Arg Ala Glu Leu Ile Ala Leu Thr Gln Ala Leu Lys
1820 1825 1830
Met Ala Glu Gly Lys Lys Leu Asn Val Tyr Thr Asp Ser Arg Tyr
1835 1840 1845
Ala Phe Ala Thr Ala His Ile His Gly Glu Ile Tyr Arg Arg Arg
1850 1855 1860
Gly Trp Leu Thr Ser Glu Gly Lys Glu Ile Lys Asn Lys Asp Glu
1865 1870 1875
Ile Leu Ala Leu Leu Lys Ala Leu Phe Leu Pro Lys Arg Leu Ser
1880 1885 1890
Ile Ile His Cys Pro Gly His Gln Lys Gly His Ser Ala Glu Ala
1895 1900 1905
Arg Gly Asn Arg Met Ala Asp Gln Ala Ala Arg Lys Ala Ala Ile
1910 1915 1920
Thr Glu Thr Pro Asp Thr Ser Thr Leu Leu Ile Glu Asn Ser Ser
1925 1930 1935
Pro Ser Gly Gly Ser Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu
1940 1945 1950
Pro Lys Lys Lys Arg Lys Val Gly Ser Gly
1955 1960
<210> 74
<211> 1979
<212> PRT
<213> 人工的
<220>
<223> 合成的REDRAW编辑器
<400> 74
Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys
1 5 10 15
Arg Lys Val Ser Lys Leu Glu Lys Phe Thr Asn Cys Tyr Ser Leu Ser
20 25 30
Lys Thr Leu Arg Phe Lys Ala Ile Pro Val Gly Lys Thr Gln Glu Asn
35 40 45
Ile Asp Asn Lys Arg Leu Leu Val Glu Asp Glu Lys Arg Ala Glu Asp
50 55 60
Tyr Lys Gly Val Lys Lys Leu Leu Asp Arg Tyr Tyr Leu Ser Phe Ile
65 70 75 80
Asn Asp Val Leu His Ser Ile Lys Leu Lys Asn Leu Asn Asn Tyr Ile
85 90 95
Ser Leu Phe Arg Lys Lys Thr Arg Thr Glu Lys Glu Asn Lys Glu Leu
100 105 110
Glu Asn Leu Glu Ile Asn Leu Arg Lys Glu Ile Ala Lys Ala Phe Lys
115 120 125
Gly Asn Glu Gly Tyr Lys Ser Leu Phe Lys Lys Asp Ile Ile Glu Thr
130 135 140
Ile Leu Pro Glu Phe Leu Asp Asp Lys Asp Glu Ile Ala Leu Val Asn
145 150 155 160
Ser Phe Asn Gly Phe Thr Thr Ala Phe Thr Gly Phe Phe Asp Asn Arg
165 170 175
Glu Asn Met Phe Ser Glu Glu Ala Lys Ser Thr Ser Ile Ala Phe Arg
180 185 190
Cys Ile Asn Glu Asn Leu Thr Arg Tyr Ile Ser Asn Met Asp Ile Phe
195 200 205
Glu Lys Val Asp Ala Ile Phe Asp Lys His Glu Val Gln Glu Ile Lys
210 215 220
Glu Lys Ile Leu Asn Ser Asp Tyr Asp Val Glu Asp Phe Phe Glu Gly
225 230 235 240
Glu Phe Phe Asn Phe Val Leu Thr Gln Glu Gly Ile Asp Val Tyr Asn
245 250 255
Ala Ile Ile Gly Gly Phe Val Thr Glu Ser Gly Glu Lys Ile Lys Gly
260 265 270
Leu Asn Glu Tyr Ile Asn Leu Tyr Asn Gln Lys Thr Lys Gln Lys Leu
275 280 285
Pro Lys Phe Lys Pro Leu Tyr Lys Gln Val Leu Ser Asp Arg Glu Ser
290 295 300
Leu Ser Phe Tyr Gly Glu Gly Tyr Thr Ser Asp Glu Glu Val Leu Glu
305 310 315 320
Val Phe Arg Asn Thr Leu Asn Lys Asn Ser Glu Ile Phe Ser Ser Ile
325 330 335
Lys Lys Leu Glu Lys Leu Phe Lys Asn Phe Asp Glu Tyr Ser Ser Ala
340 345 350
Gly Ile Phe Val Lys Asn Gly Pro Ala Ile Ser Thr Ile Ser Lys Asp
355 360 365
Ile Phe Gly Glu Trp Asn Val Ile Arg Asp Lys Trp Asn Ala Glu Tyr
370 375 380
Asp Asp Ile His Leu Lys Lys Lys Ala Val Val Thr Glu Lys Tyr Glu
385 390 395 400
Asp Asp Arg Arg Lys Ser Phe Lys Lys Ile Gly Ser Phe Ser Leu Glu
405 410 415
Gln Leu Gln Glu Tyr Ala Asp Ala Asp Leu Ser Val Val Glu Lys Leu
420 425 430
Lys Glu Ile Ile Ile Gln Lys Val Asp Glu Ile Tyr Lys Val Tyr Gly
435 440 445
Ser Ser Glu Lys Leu Phe Asp Ala Asp Phe Val Leu Glu Lys Ser Leu
450 455 460
Lys Lys Asn Asp Ala Val Val Ala Ile Met Lys Asp Leu Leu Asp Ser
465 470 475 480
Val Lys Ser Phe Glu Asn Tyr Ile Lys Ala Phe Phe Gly Glu Gly Lys
485 490 495
Glu Thr Asn Arg Asp Glu Ser Phe Tyr Gly Asp Phe Val Leu Ala Tyr
500 505 510
Asp Ile Leu Leu Lys Val Asp His Ile Tyr Asp Ala Ile Arg Asn Tyr
515 520 525
Val Thr Gln Lys Pro Tyr Ser Lys Asp Lys Phe Lys Leu Tyr Phe Gln
530 535 540
Asn Pro Gln Phe Met Gly Gly Trp Asp Lys Asp Lys Glu Thr Asp Tyr
545 550 555 560
Arg Ala Thr Ile Leu Arg Tyr Gly Ser Lys Tyr Tyr Leu Ala Ile Met
565 570 575
Asp Lys Lys Tyr Ala Lys Cys Leu Gln Lys Ile Asp Lys Asp Asp Val
580 585 590
Asn Gly Asn Tyr Glu Lys Ile Asn Tyr Lys Leu Leu Pro Gly Pro Asn
595 600 605
Lys Met Leu Pro Lys Val Phe Phe Ser Lys Lys Trp Met Ala Tyr Tyr
610 615 620
Asn Pro Ser Glu Asp Ile Gln Lys Ile Tyr Lys Asn Gly Thr Phe Lys
625 630 635 640
Lys Gly Asp Met Phe Asn Leu Asn Asp Cys His Lys Leu Ile Asp Phe
645 650 655
Phe Lys Asp Ser Ile Ser Arg Tyr Pro Lys Trp Ser Asn Ala Tyr Asp
660 665 670
Phe Asn Phe Ser Glu Thr Glu Lys Tyr Lys Asp Ile Ala Gly Phe Tyr
675 680 685
Arg Glu Val Glu Glu Gln Gly Tyr Lys Val Ser Phe Glu Ser Ala Ser
690 695 700
Lys Lys Glu Val Asp Lys Leu Val Glu Glu Gly Lys Leu Tyr Met Phe
705 710 715 720
Gln Ile Tyr Asn Lys Asp Phe Ser Asp Lys Ser His Gly Thr Pro Asn
725 730 735
Leu His Thr Met Tyr Phe Lys Leu Leu Phe Asp Glu Asn Asn His Gly
740 745 750
Gln Ile Arg Leu Ser Gly Gly Ala Glu Leu Phe Met Arg Arg Ala Ser
755 760 765
Leu Lys Lys Glu Glu Leu Val Val His Pro Ala Asn Ser Pro Ile Ala
770 775 780
Asn Lys Asn Pro Asp Asn Pro Lys Lys Thr Thr Thr Leu Ser Tyr Asp
785 790 795 800
Val Tyr Lys Asp Lys Arg Phe Ser Glu Asp Gln Tyr Glu Leu His Ile
805 810 815
Pro Ile Ala Ile Asn Lys Cys Pro Lys Asn Ile Phe Lys Ile Asn Thr
820 825 830
Glu Val Arg Val Leu Leu Lys His Asp Asp Asn Pro Tyr Val Ile Gly
835 840 845
Ile Asp Arg Gly Glu Arg Asn Leu Leu Tyr Ile Val Val Val Asp Gly
850 855 860
Lys Gly Asn Ile Val Glu Gln Tyr Ser Leu Asn Glu Ile Ile Asn Asn
865 870 875 880
Phe Asn Gly Ile Arg Ile Lys Thr Asp Tyr His Ser Leu Leu Asp Lys
885 890 895
Lys Glu Lys Glu Arg Phe Glu Ala Arg Gln Asn Trp Thr Ser Ile Glu
900 905 910
Asn Ile Lys Glu Leu Lys Ala Gly Tyr Ile Ser Gln Val Val His Lys
915 920 925
Ile Cys Glu Leu Val Glu Lys Tyr Asp Ala Val Ile Ala Leu Glu Asp
930 935 940
Leu Asn Ser Gly Phe Lys Asn Ser Arg Val Lys Val Glu Lys Gln Val
945 950 955 960
Tyr Gln Lys Phe Glu Lys Met Leu Ile Asp Lys Leu Asn Tyr Met Val
965 970 975
Asp Lys Lys Ser Asn Pro Cys Ala Thr Gly Gly Ala Leu Lys Gly Tyr
980 985 990
Gln Ile Thr Asn Lys Phe Glu Ser Phe Lys Ser Met Ser Thr Gln Asn
995 1000 1005
Gly Phe Ile Phe Tyr Ile Pro Ala Trp Leu Thr Ser Lys Ile Asp
1010 1015 1020
Pro Ser Thr Gly Phe Val Asn Leu Leu Lys Thr Lys Tyr Thr Ser
1025 1030 1035
Ile Ala Asp Ser Lys Lys Phe Ile Ser Ser Phe Asp Arg Ile Met
1040 1045 1050
Tyr Val Pro Glu Glu Asp Leu Phe Glu Phe Ala Leu Asp Tyr Lys
1055 1060 1065
Asn Phe Ser Arg Thr Asp Ala Asp Tyr Ile Lys Lys Trp Lys Leu
1070 1075 1080
Tyr Ser Tyr Gly Asn Arg Ile Arg Ile Phe Arg Asn Pro Lys Lys
1085 1090 1095
Asn Asn Val Phe Asp Trp Glu Glu Val Cys Leu Thr Ser Ala Tyr
1100 1105 1110
Lys Glu Leu Phe Asn Lys Tyr Gly Ile Asn Tyr Gln Gln Gly Asp
1115 1120 1125
Ile Arg Ala Leu Leu Cys Glu Gln Ser Asp Lys Ala Phe Tyr Ser
1130 1135 1140
Ser Phe Met Ala Leu Met Ser Leu Met Leu Gln Met Ala Asn Ser
1145 1150 1155
Ile Thr Gly Arg Thr Asp Val Asp Phe Leu Ile Ser Pro Val Lys
1160 1165 1170
Asn Ser Asp Gly Ile Phe Tyr Asp Ser Arg Asn Tyr Glu Ala Gln
1175 1180 1185
Glu Asn Ala Ile Leu Pro Lys Asn Ala Asp Ala Asn Gly Ala Tyr
1190 1195 1200
Asn Ile Ala Arg Lys Val Leu Trp Ala Ile Gly Gln Phe Lys Lys
1205 1210 1215
Ala Glu Asp Glu Lys Leu Asp Lys Val Lys Ile Ala Ile Ser Asn
1220 1225 1230
Lys Glu Trp Leu Glu Tyr Ala Gln Thr Ser Val Lys His Ser Gly
1235 1240 1245
Gly Ser Ser Gly Gly Ser Ser Gly Ser Glu Thr Pro Gly Thr Ser
1250 1255 1260
Glu Ser Ala Thr Pro Glu Ser Ser Gly Gly Ser Ser Gly Gly Ser
1265 1270 1275
Thr Leu Asn Ile Glu Asp Glu Tyr Arg Leu His Glu Thr Ser Lys
1280 1285 1290
Glu Pro Asp Val Ser Leu Gly Ser Thr Trp Leu Ser Asp Phe Pro
1295 1300 1305
Gln Ala Trp Ala Glu Thr Gly Gly Met Gly Leu Ala Val Arg Gln
1310 1315 1320
Ala Pro Leu Ile Ile Pro Leu Lys Ala Thr Ser Thr Pro Val Ser
1325 1330 1335
Ile Lys Gln Tyr Pro Met Ser Gln Glu Ala Arg Leu Gly Ile Lys
1340 1345 1350
Pro His Ile Gln Arg Leu Leu Asp Gln Gly Ile Leu Val Pro Cys
1355 1360 1365
Gln Ser Pro Trp Asn Thr Pro Leu Leu Pro Val Lys Lys Pro Gly
1370 1375 1380
Thr Asn Asp Tyr Arg Pro Val Gln Asp Leu Arg Glu Val Asn Lys
1385 1390 1395
Arg Val Glu Asp Ile His Pro Thr Val Pro Asn Pro Tyr Asn Leu
1400 1405 1410
Leu Ser Gly Leu Pro Pro Ser His Gln Trp Tyr Thr Val Leu Asp
1415 1420 1425
Leu Lys Asp Ala Phe Phe Cys Leu Arg Leu His Pro Thr Ser Gln
1430 1435 1440
Pro Leu Phe Ala Phe Glu Trp Arg Asp Pro Glu Met Gly Ile Ser
1445 1450 1455
Gly Gln Leu Thr Trp Thr Arg Leu Pro Gln Gly Phe Lys Asn Ser
1460 1465 1470
Pro Thr Leu Phe Asn Glu Ala Leu His Arg Asp Leu Ala Asp Phe
1475 1480 1485
Arg Ile Gln His Pro Asp Leu Ile Leu Leu Gln Tyr Val Asp Asp
1490 1495 1500
Leu Leu Leu Ala Ala Thr Ser Glu Leu Asp Cys Gln Gln Gly Thr
1505 1510 1515
Arg Ala Leu Leu Gln Thr Leu Gly Asn Leu Gly Tyr Arg Ala Ser
1520 1525 1530
Ala Lys Lys Ala Gln Ile Cys Gln Lys Gln Val Lys Tyr Leu Gly
1535 1540 1545
Tyr Leu Leu Lys Glu Gly Gln Arg Trp Leu Thr Glu Ala Arg Lys
1550 1555 1560
Glu Thr Val Met Gly Gln Pro Thr Pro Lys Thr Pro Arg Gln Leu
1565 1570 1575
Arg Glu Phe Leu Gly Lys Ala Gly Phe Cys Arg Leu Phe Ile Pro
1580 1585 1590
Gly Phe Ala Glu Met Ala Ala Pro Leu Tyr Pro Leu Thr Lys Pro
1595 1600 1605
Gly Thr Leu Phe Asn Trp Gly Pro Asp Gln Gln Lys Ala Tyr Gln
1610 1615 1620
Glu Ile Lys Gln Ala Leu Leu Thr Ala Pro Ala Leu Gly Leu Pro
1625 1630 1635
Asp Leu Thr Lys Pro Phe Glu Leu Phe Val Asp Glu Lys Gln Gly
1640 1645 1650
Tyr Ala Lys Gly Val Leu Thr Gln Lys Leu Gly Pro Trp Arg Arg
1655 1660 1665
Pro Val Ala Tyr Leu Ser Lys Lys Leu Asp Pro Val Ala Ala Gly
1670 1675 1680
Trp Pro Pro Cys Leu Arg Met Val Ala Ala Ile Ala Val Leu Thr
1685 1690 1695
Lys Asp Ala Gly Lys Leu Thr Met Gly Gln Pro Leu Val Ile Leu
1700 1705 1710
Ala Pro His Ala Val Glu Ala Leu Val Lys Gln Pro Pro Asp Arg
1715 1720 1725
Trp Leu Ser Asn Ala Arg Met Thr His Tyr Gln Ala Leu Leu Leu
1730 1735 1740
Asp Thr Asp Arg Val Gln Phe Gly Pro Val Val Ala Leu Asn Pro
1745 1750 1755
Ala Thr Leu Leu Pro Leu Pro Glu Glu Gly Leu Gln His Asn Cys
1760 1765 1770
Leu Asp Ile Leu Ala Glu Ala His Gly Thr Arg Pro Asp Leu Thr
1775 1780 1785
Asp Gln Pro Leu Pro Asp Ala Asp His Thr Trp Tyr Thr Asp Gly
1790 1795 1800
Ser Ser Leu Leu Gln Glu Gly Gln Arg Lys Ala Gly Ala Ala Val
1805 1810 1815
Thr Thr Glu Thr Glu Val Ile Trp Ala Lys Ala Leu Pro Ala Gly
1820 1825 1830
Thr Ser Ala Gln Arg Ala Glu Leu Ile Ala Leu Thr Gln Ala Leu
1835 1840 1845
Lys Met Ala Glu Gly Lys Lys Leu Asn Val Tyr Thr Asp Ser Arg
1850 1855 1860
Tyr Ala Phe Ala Thr Ala His Ile His Gly Glu Ile Tyr Arg Arg
1865 1870 1875
Arg Gly Trp Leu Thr Ser Glu Gly Lys Glu Ile Lys Asn Lys Asp
1880 1885 1890
Glu Ile Leu Ala Leu Leu Lys Ala Leu Phe Leu Pro Lys Arg Leu
1895 1900 1905
Ser Ile Ile His Cys Pro Gly His Gln Lys Gly His Ser Ala Glu
1910 1915 1920
Ala Arg Gly Asn Arg Met Ala Asp Gln Ala Ala Arg Lys Ala Ala
1925 1930 1935
Ile Thr Glu Thr Pro Asp Thr Ser Thr Leu Leu Ile Glu Asn Ser
1940 1945 1950
Ser Pro Ser Gly Gly Ser Lys Arg Thr Ala Asp Gly Ser Glu Phe
1955 1960 1965
Glu Pro Lys Lys Lys Arg Lys Val Gly Ser Gly
1970 1975
<210> 75
<211> 1963
<212> PRT
<213> 人工的
<220>
<223> 合成的REDRAW编辑器
<400> 75
Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys
1 5 10 15
Arg Lys Val Thr Leu Asn Ile Glu Asp Glu Tyr Arg Leu His Glu Thr
20 25 30
Ser Lys Glu Pro Asp Val Ser Leu Gly Ser Thr Trp Leu Ser Asp Phe
35 40 45
Pro Gln Ala Trp Ala Glu Thr Gly Gly Met Gly Leu Ala Val Arg Gln
50 55 60
Ala Pro Leu Ile Ile Pro Leu Lys Ala Thr Ser Thr Pro Val Ser Ile
65 70 75 80
Lys Gln Tyr Pro Met Ser Gln Glu Ala Arg Leu Gly Ile Lys Pro His
85 90 95
Ile Gln Arg Leu Leu Asp Gln Gly Ile Leu Val Pro Cys Gln Ser Pro
100 105 110
Trp Asn Thr Pro Leu Leu Pro Val Lys Lys Pro Gly Thr Asn Asp Tyr
115 120 125
Arg Pro Val Gln Asp Leu Arg Glu Val Asn Lys Arg Val Glu Asp Ile
130 135 140
His Pro Thr Val Pro Asn Pro Tyr Asn Leu Leu Ser Gly Leu Pro Pro
145 150 155 160
Ser His Gln Trp Tyr Thr Val Leu Asp Leu Lys Asp Ala Phe Phe Cys
165 170 175
Leu Arg Leu His Pro Thr Ser Gln Pro Leu Phe Ala Phe Glu Trp Arg
180 185 190
Asp Pro Glu Met Gly Ile Ser Gly Gln Leu Thr Trp Thr Arg Leu Pro
195 200 205
Gln Gly Phe Lys Asn Ser Pro Thr Leu Phe Asn Glu Ala Leu His Arg
210 215 220
Asp Leu Ala Asp Phe Arg Ile Gln His Pro Asp Leu Ile Leu Leu Gln
225 230 235 240
Tyr Val Asp Asp Leu Leu Leu Ala Ala Thr Ser Glu Leu Asp Cys Gln
245 250 255
Gln Gly Thr Arg Ala Leu Leu Gln Thr Leu Gly Asn Leu Gly Tyr Arg
260 265 270
Ala Ser Ala Lys Lys Ala Gln Ile Cys Gln Lys Gln Val Lys Tyr Leu
275 280 285
Gly Tyr Leu Leu Lys Glu Gly Gln Arg Trp Leu Thr Glu Ala Arg Lys
290 295 300
Glu Thr Val Met Gly Gln Pro Thr Pro Lys Thr Pro Arg Gln Leu Arg
305 310 315 320
Glu Phe Leu Gly Lys Ala Gly Phe Cys Arg Leu Phe Ile Pro Gly Phe
325 330 335
Ala Glu Met Ala Ala Pro Leu Tyr Pro Leu Thr Lys Pro Gly Thr Leu
340 345 350
Phe Asn Trp Gly Pro Asp Gln Gln Lys Ala Tyr Gln Glu Ile Lys Gln
355 360 365
Ala Leu Leu Thr Ala Pro Ala Leu Gly Leu Pro Asp Leu Thr Lys Pro
370 375 380
Phe Glu Leu Phe Val Asp Glu Lys Gln Gly Tyr Ala Lys Gly Val Leu
385 390 395 400
Thr Gln Lys Leu Gly Pro Trp Arg Arg Pro Val Ala Tyr Leu Ser Lys
405 410 415
Lys Leu Asp Pro Val Ala Ala Gly Trp Pro Pro Cys Leu Arg Met Val
420 425 430
Ala Ala Ile Ala Val Leu Thr Lys Asp Ala Gly Lys Leu Thr Met Gly
435 440 445
Gln Pro Leu Val Ile Leu Ala Pro His Ala Val Glu Ala Leu Val Lys
450 455 460
Gln Pro Pro Asp Arg Trp Leu Ser Asn Ala Arg Met Thr His Tyr Gln
465 470 475 480
Ala Leu Leu Leu Asp Thr Asp Arg Val Gln Phe Gly Pro Val Val Ala
485 490 495
Leu Asn Pro Ala Thr Leu Leu Pro Leu Pro Glu Glu Gly Leu Gln His
500 505 510
Asn Cys Leu Asp Ile Leu Ala Glu Ala His Gly Thr Arg Pro Asp Leu
515 520 525
Thr Asp Gln Pro Leu Pro Asp Ala Asp His Thr Trp Tyr Thr Asp Gly
530 535 540
Ser Ser Leu Leu Gln Glu Gly Gln Arg Lys Ala Gly Ala Ala Val Thr
545 550 555 560
Thr Glu Thr Glu Val Ile Trp Ala Lys Ala Leu Pro Ala Gly Thr Ser
565 570 575
Ala Gln Arg Ala Glu Leu Ile Ala Leu Thr Gln Ala Leu Lys Met Ala
580 585 590
Glu Gly Lys Lys Leu Asn Val Tyr Thr Asp Ser Arg Tyr Ala Phe Ala
595 600 605
Thr Ala His Ile His Gly Glu Ile Tyr Arg Arg Arg Gly Trp Leu Thr
610 615 620
Ser Glu Gly Lys Glu Ile Lys Asn Lys Asp Glu Ile Leu Ala Leu Leu
625 630 635 640
Lys Ala Leu Phe Leu Pro Lys Arg Leu Ser Ile Ile His Cys Pro Gly
645 650 655
His Gln Lys Gly His Ser Ala Glu Ala Arg Gly Asn Arg Met Ala Asp
660 665 670
Gln Ala Ala Arg Lys Ala Ala Ile Thr Glu Thr Pro Asp Thr Ser Thr
675 680 685
Leu Leu Ile Glu Asn Ser Ser Pro Ser Gly Ser Glu Thr Pro Gly Thr
690 695 700
Ser Glu Ser Ala Thr Pro Glu Ser Ser Lys Leu Glu Lys Phe Thr Asn
705 710 715 720
Cys Tyr Ser Leu Ser Lys Thr Leu Arg Phe Lys Ala Ile Pro Val Gly
725 730 735
Lys Thr Gln Glu Asn Ile Asp Asn Lys Arg Leu Leu Val Glu Asp Glu
740 745 750
Lys Arg Ala Glu Asp Tyr Lys Gly Val Lys Lys Leu Leu Asp Arg Tyr
755 760 765
Tyr Leu Ser Phe Ile Asn Asp Val Leu His Ser Ile Lys Leu Lys Asn
770 775 780
Leu Asn Asn Tyr Ile Ser Leu Phe Arg Lys Lys Thr Arg Thr Glu Lys
785 790 795 800
Glu Asn Lys Glu Leu Glu Asn Leu Glu Ile Asn Leu Arg Lys Glu Ile
805 810 815
Ala Lys Ala Phe Lys Gly Asn Glu Gly Tyr Lys Ser Leu Phe Lys Lys
820 825 830
Asp Ile Ile Glu Thr Ile Leu Pro Glu Phe Leu Asp Asp Lys Asp Glu
835 840 845
Ile Ala Leu Val Asn Ser Phe Asn Gly Phe Thr Thr Ala Phe Thr Gly
850 855 860
Phe Phe Asp Asn Arg Glu Asn Met Phe Ser Glu Glu Ala Lys Ser Thr
865 870 875 880
Ser Ile Ala Phe Arg Cys Ile Asn Glu Asn Leu Thr Arg Tyr Ile Ser
885 890 895
Asn Met Asp Ile Phe Glu Lys Val Asp Ala Ile Phe Asp Lys His Glu
900 905 910
Val Gln Glu Ile Lys Glu Lys Ile Leu Asn Ser Asp Tyr Asp Val Glu
915 920 925
Asp Phe Phe Glu Gly Glu Phe Phe Asn Phe Val Leu Thr Gln Glu Gly
930 935 940
Ile Asp Val Tyr Asn Ala Ile Ile Gly Gly Phe Val Thr Glu Ser Gly
945 950 955 960
Glu Lys Ile Lys Gly Leu Asn Glu Tyr Ile Asn Leu Tyr Asn Gln Lys
965 970 975
Thr Lys Gln Lys Leu Pro Lys Phe Lys Pro Leu Tyr Lys Gln Val Leu
980 985 990
Ser Asp Arg Glu Ser Leu Ser Phe Tyr Gly Glu Gly Tyr Thr Ser Asp
995 1000 1005
Glu Glu Val Leu Glu Val Phe Arg Asn Thr Leu Asn Lys Asn Ser
1010 1015 1020
Glu Ile Phe Ser Ser Ile Lys Lys Leu Glu Lys Leu Phe Lys Asn
1025 1030 1035
Phe Asp Glu Tyr Ser Ser Ala Gly Ile Phe Val Lys Asn Gly Pro
1040 1045 1050
Ala Ile Ser Thr Ile Ser Lys Asp Ile Phe Gly Glu Trp Asn Val
1055 1060 1065
Ile Arg Asp Lys Trp Asn Ala Glu Tyr Asp Asp Ile His Leu Lys
1070 1075 1080
Lys Lys Ala Val Val Thr Glu Lys Tyr Glu Asp Asp Arg Arg Lys
1085 1090 1095
Ser Phe Lys Lys Ile Gly Ser Phe Ser Leu Glu Gln Leu Gln Glu
1100 1105 1110
Tyr Ala Asp Ala Asp Leu Ser Val Val Glu Lys Leu Lys Glu Ile
1115 1120 1125
Ile Ile Gln Lys Val Asp Glu Ile Tyr Lys Val Tyr Gly Ser Ser
1130 1135 1140
Glu Lys Leu Phe Asp Ala Asp Phe Val Leu Glu Lys Ser Leu Lys
1145 1150 1155
Lys Asn Asp Ala Val Val Ala Ile Met Lys Asp Leu Leu Asp Ser
1160 1165 1170
Val Lys Ser Phe Glu Asn Tyr Ile Lys Ala Phe Phe Gly Glu Gly
1175 1180 1185
Lys Glu Thr Asn Arg Asp Glu Ser Phe Tyr Gly Asp Phe Val Leu
1190 1195 1200
Ala Tyr Asp Ile Leu Leu Lys Val Asp His Ile Tyr Asp Ala Ile
1205 1210 1215
Arg Asn Tyr Val Thr Gln Lys Pro Tyr Ser Lys Asp Lys Phe Lys
1220 1225 1230
Leu Tyr Phe Gln Asn Pro Gln Phe Met Arg Gly Trp Asp Lys Asp
1235 1240 1245
Lys Glu Thr Asp Tyr Arg Ala Thr Ile Leu Arg Tyr Gly Ser Lys
1250 1255 1260
Tyr Tyr Leu Ala Ile Met Asp Lys Lys Tyr Ala Lys Cys Leu Gln
1265 1270 1275
Lys Ile Asp Lys Asp Asp Val Asn Gly Asn Tyr Glu Lys Ile Asn
1280 1285 1290
Tyr Lys Leu Leu Pro Gly Pro Asn Lys Met Leu Pro Arg Val Phe
1295 1300 1305
Phe Ser Lys Lys Trp Met Ala Tyr Tyr Asn Pro Ser Glu Asp Ile
1310 1315 1320
Gln Lys Ile Tyr Lys Asn Gly Thr Phe Lys Lys Gly Asp Met Phe
1325 1330 1335
Asn Leu Asn Asp Cys His Lys Leu Ile Asp Phe Phe Lys Asp Ser
1340 1345 1350
Ile Ser Arg Tyr Pro Lys Trp Ser Asn Ala Tyr Asp Phe Asn Phe
1355 1360 1365
Ser Glu Thr Glu Lys Tyr Lys Asp Ile Ala Gly Phe Tyr Arg Glu
1370 1375 1380
Val Glu Glu Gln Gly Tyr Lys Val Ser Phe Glu Ser Ala Ser Lys
1385 1390 1395
Lys Glu Val Asp Lys Leu Val Glu Glu Gly Lys Leu Tyr Met Phe
1400 1405 1410
Gln Ile Tyr Asn Lys Asp Phe Ser Asp Lys Ser His Gly Thr Pro
1415 1420 1425
Asn Leu His Thr Met Tyr Phe Lys Leu Leu Phe Asp Glu Asn Asn
1430 1435 1440
His Gly Gln Ile Arg Leu Ser Gly Gly Ala Glu Leu Phe Met Arg
1445 1450 1455
Arg Ala Ser Leu Lys Lys Glu Glu Leu Val Val Ala Pro Ala Asn
1460 1465 1470
Ser Pro Ile Ala Asn Lys Asn Pro Asp Asn Pro Lys Lys Thr Thr
1475 1480 1485
Thr Leu Ser Tyr Asp Val Tyr Lys Asp Lys Arg Phe Ser Glu Asp
1490 1495 1500
Gln Tyr Glu Leu His Ile Pro Ile Ala Ile Asn Lys Cys Pro Lys
1505 1510 1515
Asn Ile Phe Lys Ile Asn Thr Glu Val Arg Val Leu Leu Lys His
1520 1525 1530
Asp Asp Asn Pro Tyr Val Ile Gly Ile Asp Arg Gly Glu Arg Asn
1535 1540 1545
Leu Leu Tyr Ile Val Val Val Asp Gly Lys Gly Asn Ile Val Glu
1550 1555 1560
Gln Tyr Ser Leu Asn Glu Ile Ile Asn Asn Phe Asn Gly Ile Arg
1565 1570 1575
Ile Lys Thr Asp Tyr His Ser Leu Leu Asp Lys Lys Glu Lys Glu
1580 1585 1590
Arg Phe Glu Ala Arg Gln Asn Trp Thr Ser Ile Glu Asn Ile Lys
1595 1600 1605
Glu Leu Lys Ala Gly Tyr Ile Ser Gln Val Val His Lys Ile Cys
1610 1615 1620
Glu Leu Val Glu Lys Tyr Asp Ala Val Ile Ala Leu Glu Asp Leu
1625 1630 1635
Asn Ser Gly Phe Lys Asn Ser Arg Val Lys Val Glu Lys Gln Val
1640 1645 1650
Tyr Gln Lys Phe Glu Lys Met Leu Ile Asp Lys Leu Asn Tyr Met
1655 1660 1665
Val Asp Lys Lys Ser Asn Pro Cys Ala Thr Gly Gly Ala Leu Lys
1670 1675 1680
Gly Tyr Gln Ile Thr Asn Lys Phe Glu Ser Phe Lys Ser Met Ser
1685 1690 1695
Thr Gln Asn Gly Phe Ile Phe Tyr Ile Pro Ala Trp Leu Thr Ser
1700 1705 1710
Lys Ile Asp Pro Ser Thr Gly Phe Val Asn Leu Leu Lys Thr Lys
1715 1720 1725
Tyr Thr Ser Ile Ala Asp Ser Lys Lys Phe Ile Ser Ser Phe Asp
1730 1735 1740
Arg Ile Met Tyr Val Pro Glu Glu Asp Leu Phe Glu Phe Ala Leu
1745 1750 1755
Asp Tyr Lys Asn Phe Ser Arg Thr Asp Ala Asp Tyr Ile Lys Lys
1760 1765 1770
Trp Lys Leu Tyr Ser Tyr Gly Asn Arg Ile Arg Ile Phe Arg Asn
1775 1780 1785
Pro Lys Lys Asn Asn Val Phe Asp Trp Glu Glu Val Cys Leu Thr
1790 1795 1800
Ser Ala Tyr Lys Glu Leu Phe Asn Lys Tyr Gly Ile Asn Tyr Gln
1805 1810 1815
Gln Gly Asp Ile Arg Ala Leu Leu Cys Glu Gln Ser Asp Lys Ala
1820 1825 1830
Phe Tyr Ser Ser Phe Met Ala Leu Met Ser Leu Met Leu Gln Met
1835 1840 1845
Ala Asn Ser Ile Thr Gly Arg Thr Asp Val Asp Phe Leu Ile Ser
1850 1855 1860
Pro Val Lys Asn Ser Asp Gly Ile Phe Tyr Asp Ser Arg Asn Tyr
1865 1870 1875
Glu Ala Gln Glu Asn Ala Ile Leu Pro Lys Asn Ala Asp Ala Asn
1880 1885 1890
Gly Ala Tyr Asn Ile Ala Arg Lys Val Leu Trp Ala Ile Gly Gln
1895 1900 1905
Phe Lys Lys Ala Glu Asp Glu Lys Leu Asp Lys Val Lys Ile Ala
1910 1915 1920
Ile Ser Asn Lys Glu Trp Leu Glu Tyr Ala Gln Thr Ser Val Lys
1925 1930 1935
His Ser Gly Gly Ser Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu
1940 1945 1950
Pro Lys Lys Lys Arg Lys Val Gly Ser Gly
1955 1960
<210> 76
<211> 1979
<212> PRT
<213> 人工的
<220>
<223> 合成的REDRAW编辑器
<400> 76
Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys
1 5 10 15
Arg Lys Val Thr Leu Asn Ile Glu Asp Glu Tyr Arg Leu His Glu Thr
20 25 30
Ser Lys Glu Pro Asp Val Ser Leu Gly Ser Thr Trp Leu Ser Asp Phe
35 40 45
Pro Gln Ala Trp Ala Glu Thr Gly Gly Met Gly Leu Ala Val Arg Gln
50 55 60
Ala Pro Leu Ile Ile Pro Leu Lys Ala Thr Ser Thr Pro Val Ser Ile
65 70 75 80
Lys Gln Tyr Pro Met Ser Gln Glu Ala Arg Leu Gly Ile Lys Pro His
85 90 95
Ile Gln Arg Leu Leu Asp Gln Gly Ile Leu Val Pro Cys Gln Ser Pro
100 105 110
Trp Asn Thr Pro Leu Leu Pro Val Lys Lys Pro Gly Thr Asn Asp Tyr
115 120 125
Arg Pro Val Gln Asp Leu Arg Glu Val Asn Lys Arg Val Glu Asp Ile
130 135 140
His Pro Thr Val Pro Asn Pro Tyr Asn Leu Leu Ser Gly Leu Pro Pro
145 150 155 160
Ser His Gln Trp Tyr Thr Val Leu Asp Leu Lys Asp Ala Phe Phe Cys
165 170 175
Leu Arg Leu His Pro Thr Ser Gln Pro Leu Phe Ala Phe Glu Trp Arg
180 185 190
Asp Pro Glu Met Gly Ile Ser Gly Gln Leu Thr Trp Thr Arg Leu Pro
195 200 205
Gln Gly Phe Lys Asn Ser Pro Thr Leu Phe Asn Glu Ala Leu His Arg
210 215 220
Asp Leu Ala Asp Phe Arg Ile Gln His Pro Asp Leu Ile Leu Leu Gln
225 230 235 240
Tyr Val Asp Asp Leu Leu Leu Ala Ala Thr Ser Glu Leu Asp Cys Gln
245 250 255
Gln Gly Thr Arg Ala Leu Leu Gln Thr Leu Gly Asn Leu Gly Tyr Arg
260 265 270
Ala Ser Ala Lys Lys Ala Gln Ile Cys Gln Lys Gln Val Lys Tyr Leu
275 280 285
Gly Tyr Leu Leu Lys Glu Gly Gln Arg Trp Leu Thr Glu Ala Arg Lys
290 295 300
Glu Thr Val Met Gly Gln Pro Thr Pro Lys Thr Pro Arg Gln Leu Arg
305 310 315 320
Glu Phe Leu Gly Lys Ala Gly Phe Cys Arg Leu Phe Ile Pro Gly Phe
325 330 335
Ala Glu Met Ala Ala Pro Leu Tyr Pro Leu Thr Lys Pro Gly Thr Leu
340 345 350
Phe Asn Trp Gly Pro Asp Gln Gln Lys Ala Tyr Gln Glu Ile Lys Gln
355 360 365
Ala Leu Leu Thr Ala Pro Ala Leu Gly Leu Pro Asp Leu Thr Lys Pro
370 375 380
Phe Glu Leu Phe Val Asp Glu Lys Gln Gly Tyr Ala Lys Gly Val Leu
385 390 395 400
Thr Gln Lys Leu Gly Pro Trp Arg Arg Pro Val Ala Tyr Leu Ser Lys
405 410 415
Lys Leu Asp Pro Val Ala Ala Gly Trp Pro Pro Cys Leu Arg Met Val
420 425 430
Ala Ala Ile Ala Val Leu Thr Lys Asp Ala Gly Lys Leu Thr Met Gly
435 440 445
Gln Pro Leu Val Ile Leu Ala Pro His Ala Val Glu Ala Leu Val Lys
450 455 460
Gln Pro Pro Asp Arg Trp Leu Ser Asn Ala Arg Met Thr His Tyr Gln
465 470 475 480
Ala Leu Leu Leu Asp Thr Asp Arg Val Gln Phe Gly Pro Val Val Ala
485 490 495
Leu Asn Pro Ala Thr Leu Leu Pro Leu Pro Glu Glu Gly Leu Gln His
500 505 510
Asn Cys Leu Asp Ile Leu Ala Glu Ala His Gly Thr Arg Pro Asp Leu
515 520 525
Thr Asp Gln Pro Leu Pro Asp Ala Asp His Thr Trp Tyr Thr Asp Gly
530 535 540
Ser Ser Leu Leu Gln Glu Gly Gln Arg Lys Ala Gly Ala Ala Val Thr
545 550 555 560
Thr Glu Thr Glu Val Ile Trp Ala Lys Ala Leu Pro Ala Gly Thr Ser
565 570 575
Ala Gln Arg Ala Glu Leu Ile Ala Leu Thr Gln Ala Leu Lys Met Ala
580 585 590
Glu Gly Lys Lys Leu Asn Val Tyr Thr Asp Ser Arg Tyr Ala Phe Ala
595 600 605
Thr Ala His Ile His Gly Glu Ile Tyr Arg Arg Arg Gly Trp Leu Thr
610 615 620
Ser Glu Gly Lys Glu Ile Lys Asn Lys Asp Glu Ile Leu Ala Leu Leu
625 630 635 640
Lys Ala Leu Phe Leu Pro Lys Arg Leu Ser Ile Ile His Cys Pro Gly
645 650 655
His Gln Lys Gly His Ser Ala Glu Ala Arg Gly Asn Arg Met Ala Asp
660 665 670
Gln Ala Ala Arg Lys Ala Ala Ile Thr Glu Thr Pro Asp Thr Ser Thr
675 680 685
Leu Leu Ile Glu Asn Ser Ser Pro Ser Gly Gly Ser Ser Gly Gly Ser
690 695 700
Ser Gly Ser Glu Thr Pro Gly Thr Ser Glu Ser Ala Thr Pro Glu Ser
705 710 715 720
Ser Gly Gly Ser Ser Gly Gly Ser Ser Lys Leu Glu Lys Phe Thr Asn
725 730 735
Cys Tyr Ser Leu Ser Lys Thr Leu Arg Phe Lys Ala Ile Pro Val Gly
740 745 750
Lys Thr Gln Glu Asn Ile Asp Asn Lys Arg Leu Leu Val Glu Asp Glu
755 760 765
Lys Arg Ala Glu Asp Tyr Lys Gly Val Lys Lys Leu Leu Asp Arg Tyr
770 775 780
Tyr Leu Ser Phe Ile Asn Asp Val Leu His Ser Ile Lys Leu Lys Asn
785 790 795 800
Leu Asn Asn Tyr Ile Ser Leu Phe Arg Lys Lys Thr Arg Thr Glu Lys
805 810 815
Glu Asn Lys Glu Leu Glu Asn Leu Glu Ile Asn Leu Arg Lys Glu Ile
820 825 830
Ala Lys Ala Phe Lys Gly Asn Glu Gly Tyr Lys Ser Leu Phe Lys Lys
835 840 845
Asp Ile Ile Glu Thr Ile Leu Pro Glu Phe Leu Asp Asp Lys Asp Glu
850 855 860
Ile Ala Leu Val Asn Ser Phe Asn Gly Phe Thr Thr Ala Phe Thr Gly
865 870 875 880
Phe Phe Asp Asn Arg Glu Asn Met Phe Ser Glu Glu Ala Lys Ser Thr
885 890 895
Ser Ile Ala Phe Arg Cys Ile Asn Glu Asn Leu Thr Arg Tyr Ile Ser
900 905 910
Asn Met Asp Ile Phe Glu Lys Val Asp Ala Ile Phe Asp Lys His Glu
915 920 925
Val Gln Glu Ile Lys Glu Lys Ile Leu Asn Ser Asp Tyr Asp Val Glu
930 935 940
Asp Phe Phe Glu Gly Glu Phe Phe Asn Phe Val Leu Thr Gln Glu Gly
945 950 955 960
Ile Asp Val Tyr Asn Ala Ile Ile Gly Gly Phe Val Thr Glu Ser Gly
965 970 975
Glu Lys Ile Lys Gly Leu Asn Glu Tyr Ile Asn Leu Tyr Asn Gln Lys
980 985 990
Thr Lys Gln Lys Leu Pro Lys Phe Lys Pro Leu Tyr Lys Gln Val Leu
995 1000 1005
Ser Asp Arg Glu Ser Leu Ser Phe Tyr Gly Glu Gly Tyr Thr Ser
1010 1015 1020
Asp Glu Glu Val Leu Glu Val Phe Arg Asn Thr Leu Asn Lys Asn
1025 1030 1035
Ser Glu Ile Phe Ser Ser Ile Lys Lys Leu Glu Lys Leu Phe Lys
1040 1045 1050
Asn Phe Asp Glu Tyr Ser Ser Ala Gly Ile Phe Val Lys Asn Gly
1055 1060 1065
Pro Ala Ile Ser Thr Ile Ser Lys Asp Ile Phe Gly Glu Trp Asn
1070 1075 1080
Val Ile Arg Asp Lys Trp Asn Ala Glu Tyr Asp Asp Ile His Leu
1085 1090 1095
Lys Lys Lys Ala Val Val Thr Glu Lys Tyr Glu Asp Asp Arg Arg
1100 1105 1110
Lys Ser Phe Lys Lys Ile Gly Ser Phe Ser Leu Glu Gln Leu Gln
1115 1120 1125
Glu Tyr Ala Asp Ala Asp Leu Ser Val Val Glu Lys Leu Lys Glu
1130 1135 1140
Ile Ile Ile Gln Lys Val Asp Glu Ile Tyr Lys Val Tyr Gly Ser
1145 1150 1155
Ser Glu Lys Leu Phe Asp Ala Asp Phe Val Leu Glu Lys Ser Leu
1160 1165 1170
Lys Lys Asn Asp Ala Val Val Ala Ile Met Lys Asp Leu Leu Asp
1175 1180 1185
Ser Val Lys Ser Phe Glu Asn Tyr Ile Lys Ala Phe Phe Gly Glu
1190 1195 1200
Gly Lys Glu Thr Asn Arg Asp Glu Ser Phe Tyr Gly Asp Phe Val
1205 1210 1215
Leu Ala Tyr Asp Ile Leu Leu Lys Val Asp His Ile Tyr Asp Ala
1220 1225 1230
Ile Arg Asn Tyr Val Thr Gln Lys Pro Tyr Ser Lys Asp Lys Phe
1235 1240 1245
Lys Leu Tyr Phe Gln Asn Pro Gln Phe Met Arg Gly Trp Asp Lys
1250 1255 1260
Asp Lys Glu Thr Asp Tyr Arg Ala Thr Ile Leu Arg Tyr Gly Ser
1265 1270 1275
Lys Tyr Tyr Leu Ala Ile Met Asp Lys Lys Tyr Ala Lys Cys Leu
1280 1285 1290
Gln Lys Ile Asp Lys Asp Asp Val Asn Gly Asn Tyr Glu Lys Ile
1295 1300 1305
Asn Tyr Lys Leu Leu Pro Gly Pro Asn Lys Met Leu Pro Arg Val
1310 1315 1320
Phe Phe Ser Lys Lys Trp Met Ala Tyr Tyr Asn Pro Ser Glu Asp
1325 1330 1335
Ile Gln Lys Ile Tyr Lys Asn Gly Thr Phe Lys Lys Gly Asp Met
1340 1345 1350
Phe Asn Leu Asn Asp Cys His Lys Leu Ile Asp Phe Phe Lys Asp
1355 1360 1365
Ser Ile Ser Arg Tyr Pro Lys Trp Ser Asn Ala Tyr Asp Phe Asn
1370 1375 1380
Phe Ser Glu Thr Glu Lys Tyr Lys Asp Ile Ala Gly Phe Tyr Arg
1385 1390 1395
Glu Val Glu Glu Gln Gly Tyr Lys Val Ser Phe Glu Ser Ala Ser
1400 1405 1410
Lys Lys Glu Val Asp Lys Leu Val Glu Glu Gly Lys Leu Tyr Met
1415 1420 1425
Phe Gln Ile Tyr Asn Lys Asp Phe Ser Asp Lys Ser His Gly Thr
1430 1435 1440
Pro Asn Leu His Thr Met Tyr Phe Lys Leu Leu Phe Asp Glu Asn
1445 1450 1455
Asn His Gly Gln Ile Arg Leu Ser Gly Gly Ala Glu Leu Phe Met
1460 1465 1470
Arg Arg Ala Ser Leu Lys Lys Glu Glu Leu Val Val Ala Pro Ala
1475 1480 1485
Asn Ser Pro Ile Ala Asn Lys Asn Pro Asp Asn Pro Lys Lys Thr
1490 1495 1500
Thr Thr Leu Ser Tyr Asp Val Tyr Lys Asp Lys Arg Phe Ser Glu
1505 1510 1515
Asp Gln Tyr Glu Leu His Ile Pro Ile Ala Ile Asn Lys Cys Pro
1520 1525 1530
Lys Asn Ile Phe Lys Ile Asn Thr Glu Val Arg Val Leu Leu Lys
1535 1540 1545
His Asp Asp Asn Pro Tyr Val Ile Gly Ile Asp Arg Gly Glu Arg
1550 1555 1560
Asn Leu Leu Tyr Ile Val Val Val Asp Gly Lys Gly Asn Ile Val
1565 1570 1575
Glu Gln Tyr Ser Leu Asn Glu Ile Ile Asn Asn Phe Asn Gly Ile
1580 1585 1590
Arg Ile Lys Thr Asp Tyr His Ser Leu Leu Asp Lys Lys Glu Lys
1595 1600 1605
Glu Arg Phe Glu Ala Arg Gln Asn Trp Thr Ser Ile Glu Asn Ile
1610 1615 1620
Lys Glu Leu Lys Ala Gly Tyr Ile Ser Gln Val Val His Lys Ile
1625 1630 1635
Cys Glu Leu Val Glu Lys Tyr Asp Ala Val Ile Ala Leu Glu Asp
1640 1645 1650
Leu Asn Ser Gly Phe Lys Asn Ser Arg Val Lys Val Glu Lys Gln
1655 1660 1665
Val Tyr Gln Lys Phe Glu Lys Met Leu Ile Asp Lys Leu Asn Tyr
1670 1675 1680
Met Val Asp Lys Lys Ser Asn Pro Cys Ala Thr Gly Gly Ala Leu
1685 1690 1695
Lys Gly Tyr Gln Ile Thr Asn Lys Phe Glu Ser Phe Lys Ser Met
1700 1705 1710
Ser Thr Gln Asn Gly Phe Ile Phe Tyr Ile Pro Ala Trp Leu Thr
1715 1720 1725
Ser Lys Ile Asp Pro Ser Thr Gly Phe Val Asn Leu Leu Lys Thr
1730 1735 1740
Lys Tyr Thr Ser Ile Ala Asp Ser Lys Lys Phe Ile Ser Ser Phe
1745 1750 1755
Asp Arg Ile Met Tyr Val Pro Glu Glu Asp Leu Phe Glu Phe Ala
1760 1765 1770
Leu Asp Tyr Lys Asn Phe Ser Arg Thr Asp Ala Asp Tyr Ile Lys
1775 1780 1785
Lys Trp Lys Leu Tyr Ser Tyr Gly Asn Arg Ile Arg Ile Phe Arg
1790 1795 1800
Asn Pro Lys Lys Asn Asn Val Phe Asp Trp Glu Glu Val Cys Leu
1805 1810 1815
Thr Ser Ala Tyr Lys Glu Leu Phe Asn Lys Tyr Gly Ile Asn Tyr
1820 1825 1830
Gln Gln Gly Asp Ile Arg Ala Leu Leu Cys Glu Gln Ser Asp Lys
1835 1840 1845
Ala Phe Tyr Ser Ser Phe Met Ala Leu Met Ser Leu Met Leu Gln
1850 1855 1860
Met Ala Asn Ser Ile Thr Gly Arg Thr Asp Val Asp Phe Leu Ile
1865 1870 1875
Ser Pro Val Lys Asn Ser Asp Gly Ile Phe Tyr Asp Ser Arg Asn
1880 1885 1890
Tyr Glu Ala Gln Glu Asn Ala Ile Leu Pro Lys Asn Ala Asp Ala
1895 1900 1905
Asn Gly Ala Tyr Asn Ile Ala Arg Lys Val Leu Trp Ala Ile Gly
1910 1915 1920
Gln Phe Lys Lys Ala Glu Asp Glu Lys Leu Asp Lys Val Lys Ile
1925 1930 1935
Ala Ile Ser Asn Lys Glu Trp Leu Glu Tyr Ala Gln Thr Ser Val
1940 1945 1950
Lys His Ser Gly Gly Ser Lys Arg Thr Ala Asp Gly Ser Glu Phe
1955 1960 1965
Glu Pro Lys Lys Lys Arg Lys Val Gly Ser Gly
1970 1975
<210> 77
<211> 1963
<212> PRT
<213> 人工的
<220>
<223> 合成的REDRAW编辑器
<400> 77
Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys
1 5 10 15
Arg Lys Val Ser Lys Leu Glu Lys Phe Thr Asn Cys Tyr Ser Leu Ser
20 25 30
Lys Thr Leu Arg Phe Lys Ala Ile Pro Val Gly Lys Thr Gln Glu Asn
35 40 45
Ile Asp Asn Lys Arg Leu Leu Val Glu Asp Glu Lys Arg Ala Glu Asp
50 55 60
Tyr Lys Gly Val Lys Lys Leu Leu Asp Arg Tyr Tyr Leu Ser Phe Ile
65 70 75 80
Asn Asp Val Leu His Ser Ile Lys Leu Lys Asn Leu Asn Asn Tyr Ile
85 90 95
Ser Leu Phe Arg Lys Lys Thr Arg Thr Glu Lys Glu Asn Lys Glu Leu
100 105 110
Glu Asn Leu Glu Ile Asn Leu Arg Lys Glu Ile Ala Lys Ala Phe Lys
115 120 125
Gly Asn Glu Gly Tyr Lys Ser Leu Phe Lys Lys Asp Ile Ile Glu Thr
130 135 140
Ile Leu Pro Glu Phe Leu Asp Asp Lys Asp Glu Ile Ala Leu Val Asn
145 150 155 160
Ser Phe Asn Gly Phe Thr Thr Ala Phe Thr Gly Phe Phe Asp Asn Arg
165 170 175
Glu Asn Met Phe Ser Glu Glu Ala Lys Ser Thr Ser Ile Ala Phe Arg
180 185 190
Cys Ile Asn Glu Asn Leu Thr Arg Tyr Ile Ser Asn Met Asp Ile Phe
195 200 205
Glu Lys Val Asp Ala Ile Phe Asp Lys His Glu Val Gln Glu Ile Lys
210 215 220
Glu Lys Ile Leu Asn Ser Asp Tyr Asp Val Glu Asp Phe Phe Glu Gly
225 230 235 240
Glu Phe Phe Asn Phe Val Leu Thr Gln Glu Gly Ile Asp Val Tyr Asn
245 250 255
Ala Ile Ile Gly Gly Phe Val Thr Glu Ser Gly Glu Lys Ile Lys Gly
260 265 270
Leu Asn Glu Tyr Ile Asn Leu Tyr Asn Gln Lys Thr Lys Gln Lys Leu
275 280 285
Pro Lys Phe Lys Pro Leu Tyr Lys Gln Val Leu Ser Asp Arg Glu Ser
290 295 300
Leu Ser Phe Tyr Gly Glu Gly Tyr Thr Ser Asp Glu Glu Val Leu Glu
305 310 315 320
Val Phe Arg Asn Thr Leu Asn Lys Asn Ser Glu Ile Phe Ser Ser Ile
325 330 335
Lys Lys Leu Glu Lys Leu Phe Lys Asn Phe Asp Glu Tyr Ser Ser Ala
340 345 350
Gly Ile Phe Val Lys Asn Gly Pro Ala Ile Ser Thr Ile Ser Lys Asp
355 360 365
Ile Phe Gly Glu Trp Asn Val Ile Arg Asp Lys Trp Asn Ala Glu Tyr
370 375 380
Asp Asp Ile His Leu Lys Lys Lys Ala Val Val Thr Glu Lys Tyr Glu
385 390 395 400
Asp Asp Arg Arg Lys Ser Phe Lys Lys Ile Gly Ser Phe Ser Leu Glu
405 410 415
Gln Leu Gln Glu Tyr Ala Asp Ala Asp Leu Ser Val Val Glu Lys Leu
420 425 430
Lys Glu Ile Ile Ile Gln Lys Val Asp Glu Ile Tyr Lys Val Tyr Gly
435 440 445
Ser Ser Glu Lys Leu Phe Asp Ala Asp Phe Val Leu Glu Lys Ser Leu
450 455 460
Lys Lys Asn Asp Ala Val Val Ala Ile Met Lys Asp Leu Leu Asp Ser
465 470 475 480
Val Lys Ser Phe Glu Asn Tyr Ile Lys Ala Phe Phe Gly Glu Gly Lys
485 490 495
Glu Thr Asn Arg Asp Glu Ser Phe Tyr Gly Asp Phe Val Leu Ala Tyr
500 505 510
Asp Ile Leu Leu Lys Val Asp His Ile Tyr Asp Ala Ile Arg Asn Tyr
515 520 525
Val Thr Gln Lys Pro Tyr Ser Lys Asp Lys Phe Lys Leu Tyr Phe Gln
530 535 540
Asn Pro Gln Phe Met Arg Gly Trp Asp Lys Asp Lys Glu Thr Asp Tyr
545 550 555 560
Arg Ala Thr Ile Leu Arg Tyr Gly Ser Lys Tyr Tyr Leu Ala Ile Met
565 570 575
Asp Lys Lys Tyr Ala Lys Cys Leu Gln Lys Ile Asp Lys Asp Asp Val
580 585 590
Asn Gly Asn Tyr Glu Lys Ile Asn Tyr Lys Leu Leu Pro Gly Pro Asn
595 600 605
Lys Met Leu Pro Arg Val Phe Phe Ser Lys Lys Trp Met Ala Tyr Tyr
610 615 620
Asn Pro Ser Glu Asp Ile Gln Lys Ile Tyr Lys Asn Gly Thr Phe Lys
625 630 635 640
Lys Gly Asp Met Phe Asn Leu Asn Asp Cys His Lys Leu Ile Asp Phe
645 650 655
Phe Lys Asp Ser Ile Ser Arg Tyr Pro Lys Trp Ser Asn Ala Tyr Asp
660 665 670
Phe Asn Phe Ser Glu Thr Glu Lys Tyr Lys Asp Ile Ala Gly Phe Tyr
675 680 685
Arg Glu Val Glu Glu Gln Gly Tyr Lys Val Ser Phe Glu Ser Ala Ser
690 695 700
Lys Lys Glu Val Asp Lys Leu Val Glu Glu Gly Lys Leu Tyr Met Phe
705 710 715 720
Gln Ile Tyr Asn Lys Asp Phe Ser Asp Lys Ser His Gly Thr Pro Asn
725 730 735
Leu His Thr Met Tyr Phe Lys Leu Leu Phe Asp Glu Asn Asn His Gly
740 745 750
Gln Ile Arg Leu Ser Gly Gly Ala Glu Leu Phe Met Arg Arg Ala Ser
755 760 765
Leu Lys Lys Glu Glu Leu Val Val Ala Pro Ala Asn Ser Pro Ile Ala
770 775 780
Asn Lys Asn Pro Asp Asn Pro Lys Lys Thr Thr Thr Leu Ser Tyr Asp
785 790 795 800
Val Tyr Lys Asp Lys Arg Phe Ser Glu Asp Gln Tyr Glu Leu His Ile
805 810 815
Pro Ile Ala Ile Asn Lys Cys Pro Lys Asn Ile Phe Lys Ile Asn Thr
820 825 830
Glu Val Arg Val Leu Leu Lys His Asp Asp Asn Pro Tyr Val Ile Gly
835 840 845
Ile Asp Arg Gly Glu Arg Asn Leu Leu Tyr Ile Val Val Val Asp Gly
850 855 860
Lys Gly Asn Ile Val Glu Gln Tyr Ser Leu Asn Glu Ile Ile Asn Asn
865 870 875 880
Phe Asn Gly Ile Arg Ile Lys Thr Asp Tyr His Ser Leu Leu Asp Lys
885 890 895
Lys Glu Lys Glu Arg Phe Glu Ala Arg Gln Asn Trp Thr Ser Ile Glu
900 905 910
Asn Ile Lys Glu Leu Lys Ala Gly Tyr Ile Ser Gln Val Val His Lys
915 920 925
Ile Cys Glu Leu Val Glu Lys Tyr Asp Ala Val Ile Ala Leu Glu Asp
930 935 940
Leu Asn Ser Gly Phe Lys Asn Ser Arg Val Lys Val Glu Lys Gln Val
945 950 955 960
Tyr Gln Lys Phe Glu Lys Met Leu Ile Asp Lys Leu Asn Tyr Met Val
965 970 975
Asp Lys Lys Ser Asn Pro Cys Ala Thr Gly Gly Ala Leu Lys Gly Tyr
980 985 990
Gln Ile Thr Asn Lys Phe Glu Ser Phe Lys Ser Met Ser Thr Gln Asn
995 1000 1005
Gly Phe Ile Phe Tyr Ile Pro Ala Trp Leu Thr Ser Lys Ile Asp
1010 1015 1020
Pro Ser Thr Gly Phe Val Asn Leu Leu Lys Thr Lys Tyr Thr Ser
1025 1030 1035
Ile Ala Asp Ser Lys Lys Phe Ile Ser Ser Phe Asp Arg Ile Met
1040 1045 1050
Tyr Val Pro Glu Glu Asp Leu Phe Glu Phe Ala Leu Asp Tyr Lys
1055 1060 1065
Asn Phe Ser Arg Thr Asp Ala Asp Tyr Ile Lys Lys Trp Lys Leu
1070 1075 1080
Tyr Ser Tyr Gly Asn Arg Ile Arg Ile Phe Arg Asn Pro Lys Lys
1085 1090 1095
Asn Asn Val Phe Asp Trp Glu Glu Val Cys Leu Thr Ser Ala Tyr
1100 1105 1110
Lys Glu Leu Phe Asn Lys Tyr Gly Ile Asn Tyr Gln Gln Gly Asp
1115 1120 1125
Ile Arg Ala Leu Leu Cys Glu Gln Ser Asp Lys Ala Phe Tyr Ser
1130 1135 1140
Ser Phe Met Ala Leu Met Ser Leu Met Leu Gln Met Ala Asn Ser
1145 1150 1155
Ile Thr Gly Arg Thr Asp Val Asp Phe Leu Ile Ser Pro Val Lys
1160 1165 1170
Asn Ser Asp Gly Ile Phe Tyr Asp Ser Arg Asn Tyr Glu Ala Gln
1175 1180 1185
Glu Asn Ala Ile Leu Pro Lys Asn Ala Asp Ala Asn Gly Ala Tyr
1190 1195 1200
Asn Ile Ala Arg Lys Val Leu Trp Ala Ile Gly Gln Phe Lys Lys
1205 1210 1215
Ala Glu Asp Glu Lys Leu Asp Lys Val Lys Ile Ala Ile Ser Asn
1220 1225 1230
Lys Glu Trp Leu Glu Tyr Ala Gln Thr Ser Val Lys His Ser Gly
1235 1240 1245
Ser Glu Thr Pro Gly Thr Ser Glu Ser Ala Thr Pro Glu Ser Thr
1250 1255 1260
Leu Asn Ile Glu Asp Glu Tyr Arg Leu His Glu Thr Ser Lys Glu
1265 1270 1275
Pro Asp Val Ser Leu Gly Ser Thr Trp Leu Ser Asp Phe Pro Gln
1280 1285 1290
Ala Trp Ala Glu Thr Gly Gly Met Gly Leu Ala Val Arg Gln Ala
1295 1300 1305
Pro Leu Ile Ile Pro Leu Lys Ala Thr Ser Thr Pro Val Ser Ile
1310 1315 1320
Lys Gln Tyr Pro Met Ser Gln Glu Ala Arg Leu Gly Ile Lys Pro
1325 1330 1335
His Ile Gln Arg Leu Leu Asp Gln Gly Ile Leu Val Pro Cys Gln
1340 1345 1350
Ser Pro Trp Asn Thr Pro Leu Leu Pro Val Lys Lys Pro Gly Thr
1355 1360 1365
Asn Asp Tyr Arg Pro Val Gln Asp Leu Arg Glu Val Asn Lys Arg
1370 1375 1380
Val Glu Asp Ile His Pro Thr Val Pro Asn Pro Tyr Asn Leu Leu
1385 1390 1395
Ser Gly Leu Pro Pro Ser His Gln Trp Tyr Thr Val Leu Asp Leu
1400 1405 1410
Lys Asp Ala Phe Phe Cys Leu Arg Leu His Pro Thr Ser Gln Pro
1415 1420 1425
Leu Phe Ala Phe Glu Trp Arg Asp Pro Glu Met Gly Ile Ser Gly
1430 1435 1440
Gln Leu Thr Trp Thr Arg Leu Pro Gln Gly Phe Lys Asn Ser Pro
1445 1450 1455
Thr Leu Phe Asn Glu Ala Leu His Arg Asp Leu Ala Asp Phe Arg
1460 1465 1470
Ile Gln His Pro Asp Leu Ile Leu Leu Gln Tyr Val Asp Asp Leu
1475 1480 1485
Leu Leu Ala Ala Thr Ser Glu Leu Asp Cys Gln Gln Gly Thr Arg
1490 1495 1500
Ala Leu Leu Gln Thr Leu Gly Asn Leu Gly Tyr Arg Ala Ser Ala
1505 1510 1515
Lys Lys Ala Gln Ile Cys Gln Lys Gln Val Lys Tyr Leu Gly Tyr
1520 1525 1530
Leu Leu Lys Glu Gly Gln Arg Trp Leu Thr Glu Ala Arg Lys Glu
1535 1540 1545
Thr Val Met Gly Gln Pro Thr Pro Lys Thr Pro Arg Gln Leu Arg
1550 1555 1560
Glu Phe Leu Gly Lys Ala Gly Phe Cys Arg Leu Phe Ile Pro Gly
1565 1570 1575
Phe Ala Glu Met Ala Ala Pro Leu Tyr Pro Leu Thr Lys Pro Gly
1580 1585 1590
Thr Leu Phe Asn Trp Gly Pro Asp Gln Gln Lys Ala Tyr Gln Glu
1595 1600 1605
Ile Lys Gln Ala Leu Leu Thr Ala Pro Ala Leu Gly Leu Pro Asp
1610 1615 1620
Leu Thr Lys Pro Phe Glu Leu Phe Val Asp Glu Lys Gln Gly Tyr
1625 1630 1635
Ala Lys Gly Val Leu Thr Gln Lys Leu Gly Pro Trp Arg Arg Pro
1640 1645 1650
Val Ala Tyr Leu Ser Lys Lys Leu Asp Pro Val Ala Ala Gly Trp
1655 1660 1665
Pro Pro Cys Leu Arg Met Val Ala Ala Ile Ala Val Leu Thr Lys
1670 1675 1680
Asp Ala Gly Lys Leu Thr Met Gly Gln Pro Leu Val Ile Leu Ala
1685 1690 1695
Pro His Ala Val Glu Ala Leu Val Lys Gln Pro Pro Asp Arg Trp
1700 1705 1710
Leu Ser Asn Ala Arg Met Thr His Tyr Gln Ala Leu Leu Leu Asp
1715 1720 1725
Thr Asp Arg Val Gln Phe Gly Pro Val Val Ala Leu Asn Pro Ala
1730 1735 1740
Thr Leu Leu Pro Leu Pro Glu Glu Gly Leu Gln His Asn Cys Leu
1745 1750 1755
Asp Ile Leu Ala Glu Ala His Gly Thr Arg Pro Asp Leu Thr Asp
1760 1765 1770
Gln Pro Leu Pro Asp Ala Asp His Thr Trp Tyr Thr Asp Gly Ser
1775 1780 1785
Ser Leu Leu Gln Glu Gly Gln Arg Lys Ala Gly Ala Ala Val Thr
1790 1795 1800
Thr Glu Thr Glu Val Ile Trp Ala Lys Ala Leu Pro Ala Gly Thr
1805 1810 1815
Ser Ala Gln Arg Ala Glu Leu Ile Ala Leu Thr Gln Ala Leu Lys
1820 1825 1830
Met Ala Glu Gly Lys Lys Leu Asn Val Tyr Thr Asp Ser Arg Tyr
1835 1840 1845
Ala Phe Ala Thr Ala His Ile His Gly Glu Ile Tyr Arg Arg Arg
1850 1855 1860
Gly Trp Leu Thr Ser Glu Gly Lys Glu Ile Lys Asn Lys Asp Glu
1865 1870 1875
Ile Leu Ala Leu Leu Lys Ala Leu Phe Leu Pro Lys Arg Leu Ser
1880 1885 1890
Ile Ile His Cys Pro Gly His Gln Lys Gly His Ser Ala Glu Ala
1895 1900 1905
Arg Gly Asn Arg Met Ala Asp Gln Ala Ala Arg Lys Ala Ala Ile
1910 1915 1920
Thr Glu Thr Pro Asp Thr Ser Thr Leu Leu Ile Glu Asn Ser Ser
1925 1930 1935
Pro Ser Gly Gly Ser Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu
1940 1945 1950
Pro Lys Lys Lys Arg Lys Val Gly Ser Gly
1955 1960
<210> 78
<211> 1979
<212> PRT
<213> 人工的
<220>
<223> 合成的REDRAW编辑器
<400> 78
Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys
1 5 10 15
Arg Lys Val Ser Lys Leu Glu Lys Phe Thr Asn Cys Tyr Ser Leu Ser
20 25 30
Lys Thr Leu Arg Phe Lys Ala Ile Pro Val Gly Lys Thr Gln Glu Asn
35 40 45
Ile Asp Asn Lys Arg Leu Leu Val Glu Asp Glu Lys Arg Ala Glu Asp
50 55 60
Tyr Lys Gly Val Lys Lys Leu Leu Asp Arg Tyr Tyr Leu Ser Phe Ile
65 70 75 80
Asn Asp Val Leu His Ser Ile Lys Leu Lys Asn Leu Asn Asn Tyr Ile
85 90 95
Ser Leu Phe Arg Lys Lys Thr Arg Thr Glu Lys Glu Asn Lys Glu Leu
100 105 110
Glu Asn Leu Glu Ile Asn Leu Arg Lys Glu Ile Ala Lys Ala Phe Lys
115 120 125
Gly Asn Glu Gly Tyr Lys Ser Leu Phe Lys Lys Asp Ile Ile Glu Thr
130 135 140
Ile Leu Pro Glu Phe Leu Asp Asp Lys Asp Glu Ile Ala Leu Val Asn
145 150 155 160
Ser Phe Asn Gly Phe Thr Thr Ala Phe Thr Gly Phe Phe Asp Asn Arg
165 170 175
Glu Asn Met Phe Ser Glu Glu Ala Lys Ser Thr Ser Ile Ala Phe Arg
180 185 190
Cys Ile Asn Glu Asn Leu Thr Arg Tyr Ile Ser Asn Met Asp Ile Phe
195 200 205
Glu Lys Val Asp Ala Ile Phe Asp Lys His Glu Val Gln Glu Ile Lys
210 215 220
Glu Lys Ile Leu Asn Ser Asp Tyr Asp Val Glu Asp Phe Phe Glu Gly
225 230 235 240
Glu Phe Phe Asn Phe Val Leu Thr Gln Glu Gly Ile Asp Val Tyr Asn
245 250 255
Ala Ile Ile Gly Gly Phe Val Thr Glu Ser Gly Glu Lys Ile Lys Gly
260 265 270
Leu Asn Glu Tyr Ile Asn Leu Tyr Asn Gln Lys Thr Lys Gln Lys Leu
275 280 285
Pro Lys Phe Lys Pro Leu Tyr Lys Gln Val Leu Ser Asp Arg Glu Ser
290 295 300
Leu Ser Phe Tyr Gly Glu Gly Tyr Thr Ser Asp Glu Glu Val Leu Glu
305 310 315 320
Val Phe Arg Asn Thr Leu Asn Lys Asn Ser Glu Ile Phe Ser Ser Ile
325 330 335
Lys Lys Leu Glu Lys Leu Phe Lys Asn Phe Asp Glu Tyr Ser Ser Ala
340 345 350
Gly Ile Phe Val Lys Asn Gly Pro Ala Ile Ser Thr Ile Ser Lys Asp
355 360 365
Ile Phe Gly Glu Trp Asn Val Ile Arg Asp Lys Trp Asn Ala Glu Tyr
370 375 380
Asp Asp Ile His Leu Lys Lys Lys Ala Val Val Thr Glu Lys Tyr Glu
385 390 395 400
Asp Asp Arg Arg Lys Ser Phe Lys Lys Ile Gly Ser Phe Ser Leu Glu
405 410 415
Gln Leu Gln Glu Tyr Ala Asp Ala Asp Leu Ser Val Val Glu Lys Leu
420 425 430
Lys Glu Ile Ile Ile Gln Lys Val Asp Glu Ile Tyr Lys Val Tyr Gly
435 440 445
Ser Ser Glu Lys Leu Phe Asp Ala Asp Phe Val Leu Glu Lys Ser Leu
450 455 460
Lys Lys Asn Asp Ala Val Val Ala Ile Met Lys Asp Leu Leu Asp Ser
465 470 475 480
Val Lys Ser Phe Glu Asn Tyr Ile Lys Ala Phe Phe Gly Glu Gly Lys
485 490 495
Glu Thr Asn Arg Asp Glu Ser Phe Tyr Gly Asp Phe Val Leu Ala Tyr
500 505 510
Asp Ile Leu Leu Lys Val Asp His Ile Tyr Asp Ala Ile Arg Asn Tyr
515 520 525
Val Thr Gln Lys Pro Tyr Ser Lys Asp Lys Phe Lys Leu Tyr Phe Gln
530 535 540
Asn Pro Gln Phe Met Arg Gly Trp Asp Lys Asp Lys Glu Thr Asp Tyr
545 550 555 560
Arg Ala Thr Ile Leu Arg Tyr Gly Ser Lys Tyr Tyr Leu Ala Ile Met
565 570 575
Asp Lys Lys Tyr Ala Lys Cys Leu Gln Lys Ile Asp Lys Asp Asp Val
580 585 590
Asn Gly Asn Tyr Glu Lys Ile Asn Tyr Lys Leu Leu Pro Gly Pro Asn
595 600 605
Lys Met Leu Pro Arg Val Phe Phe Ser Lys Lys Trp Met Ala Tyr Tyr
610 615 620
Asn Pro Ser Glu Asp Ile Gln Lys Ile Tyr Lys Asn Gly Thr Phe Lys
625 630 635 640
Lys Gly Asp Met Phe Asn Leu Asn Asp Cys His Lys Leu Ile Asp Phe
645 650 655
Phe Lys Asp Ser Ile Ser Arg Tyr Pro Lys Trp Ser Asn Ala Tyr Asp
660 665 670
Phe Asn Phe Ser Glu Thr Glu Lys Tyr Lys Asp Ile Ala Gly Phe Tyr
675 680 685
Arg Glu Val Glu Glu Gln Gly Tyr Lys Val Ser Phe Glu Ser Ala Ser
690 695 700
Lys Lys Glu Val Asp Lys Leu Val Glu Glu Gly Lys Leu Tyr Met Phe
705 710 715 720
Gln Ile Tyr Asn Lys Asp Phe Ser Asp Lys Ser His Gly Thr Pro Asn
725 730 735
Leu His Thr Met Tyr Phe Lys Leu Leu Phe Asp Glu Asn Asn His Gly
740 745 750
Gln Ile Arg Leu Ser Gly Gly Ala Glu Leu Phe Met Arg Arg Ala Ser
755 760 765
Leu Lys Lys Glu Glu Leu Val Val Ala Pro Ala Asn Ser Pro Ile Ala
770 775 780
Asn Lys Asn Pro Asp Asn Pro Lys Lys Thr Thr Thr Leu Ser Tyr Asp
785 790 795 800
Val Tyr Lys Asp Lys Arg Phe Ser Glu Asp Gln Tyr Glu Leu His Ile
805 810 815
Pro Ile Ala Ile Asn Lys Cys Pro Lys Asn Ile Phe Lys Ile Asn Thr
820 825 830
Glu Val Arg Val Leu Leu Lys His Asp Asp Asn Pro Tyr Val Ile Gly
835 840 845
Ile Asp Arg Gly Glu Arg Asn Leu Leu Tyr Ile Val Val Val Asp Gly
850 855 860
Lys Gly Asn Ile Val Glu Gln Tyr Ser Leu Asn Glu Ile Ile Asn Asn
865 870 875 880
Phe Asn Gly Ile Arg Ile Lys Thr Asp Tyr His Ser Leu Leu Asp Lys
885 890 895
Lys Glu Lys Glu Arg Phe Glu Ala Arg Gln Asn Trp Thr Ser Ile Glu
900 905 910
Asn Ile Lys Glu Leu Lys Ala Gly Tyr Ile Ser Gln Val Val His Lys
915 920 925
Ile Cys Glu Leu Val Glu Lys Tyr Asp Ala Val Ile Ala Leu Glu Asp
930 935 940
Leu Asn Ser Gly Phe Lys Asn Ser Arg Val Lys Val Glu Lys Gln Val
945 950 955 960
Tyr Gln Lys Phe Glu Lys Met Leu Ile Asp Lys Leu Asn Tyr Met Val
965 970 975
Asp Lys Lys Ser Asn Pro Cys Ala Thr Gly Gly Ala Leu Lys Gly Tyr
980 985 990
Gln Ile Thr Asn Lys Phe Glu Ser Phe Lys Ser Met Ser Thr Gln Asn
995 1000 1005
Gly Phe Ile Phe Tyr Ile Pro Ala Trp Leu Thr Ser Lys Ile Asp
1010 1015 1020
Pro Ser Thr Gly Phe Val Asn Leu Leu Lys Thr Lys Tyr Thr Ser
1025 1030 1035
Ile Ala Asp Ser Lys Lys Phe Ile Ser Ser Phe Asp Arg Ile Met
1040 1045 1050
Tyr Val Pro Glu Glu Asp Leu Phe Glu Phe Ala Leu Asp Tyr Lys
1055 1060 1065
Asn Phe Ser Arg Thr Asp Ala Asp Tyr Ile Lys Lys Trp Lys Leu
1070 1075 1080
Tyr Ser Tyr Gly Asn Arg Ile Arg Ile Phe Arg Asn Pro Lys Lys
1085 1090 1095
Asn Asn Val Phe Asp Trp Glu Glu Val Cys Leu Thr Ser Ala Tyr
1100 1105 1110
Lys Glu Leu Phe Asn Lys Tyr Gly Ile Asn Tyr Gln Gln Gly Asp
1115 1120 1125
Ile Arg Ala Leu Leu Cys Glu Gln Ser Asp Lys Ala Phe Tyr Ser
1130 1135 1140
Ser Phe Met Ala Leu Met Ser Leu Met Leu Gln Met Ala Asn Ser
1145 1150 1155
Ile Thr Gly Arg Thr Asp Val Asp Phe Leu Ile Ser Pro Val Lys
1160 1165 1170
Asn Ser Asp Gly Ile Phe Tyr Asp Ser Arg Asn Tyr Glu Ala Gln
1175 1180 1185
Glu Asn Ala Ile Leu Pro Lys Asn Ala Asp Ala Asn Gly Ala Tyr
1190 1195 1200
Asn Ile Ala Arg Lys Val Leu Trp Ala Ile Gly Gln Phe Lys Lys
1205 1210 1215
Ala Glu Asp Glu Lys Leu Asp Lys Val Lys Ile Ala Ile Ser Asn
1220 1225 1230
Lys Glu Trp Leu Glu Tyr Ala Gln Thr Ser Val Lys His Ser Gly
1235 1240 1245
Gly Ser Ser Gly Gly Ser Ser Gly Ser Glu Thr Pro Gly Thr Ser
1250 1255 1260
Glu Ser Ala Thr Pro Glu Ser Ser Gly Gly Ser Ser Gly Gly Ser
1265 1270 1275
Thr Leu Asn Ile Glu Asp Glu Tyr Arg Leu His Glu Thr Ser Lys
1280 1285 1290
Glu Pro Asp Val Ser Leu Gly Ser Thr Trp Leu Ser Asp Phe Pro
1295 1300 1305
Gln Ala Trp Ala Glu Thr Gly Gly Met Gly Leu Ala Val Arg Gln
1310 1315 1320
Ala Pro Leu Ile Ile Pro Leu Lys Ala Thr Ser Thr Pro Val Ser
1325 1330 1335
Ile Lys Gln Tyr Pro Met Ser Gln Glu Ala Arg Leu Gly Ile Lys
1340 1345 1350
Pro His Ile Gln Arg Leu Leu Asp Gln Gly Ile Leu Val Pro Cys
1355 1360 1365
Gln Ser Pro Trp Asn Thr Pro Leu Leu Pro Val Lys Lys Pro Gly
1370 1375 1380
Thr Asn Asp Tyr Arg Pro Val Gln Asp Leu Arg Glu Val Asn Lys
1385 1390 1395
Arg Val Glu Asp Ile His Pro Thr Val Pro Asn Pro Tyr Asn Leu
1400 1405 1410
Leu Ser Gly Leu Pro Pro Ser His Gln Trp Tyr Thr Val Leu Asp
1415 1420 1425
Leu Lys Asp Ala Phe Phe Cys Leu Arg Leu His Pro Thr Ser Gln
1430 1435 1440
Pro Leu Phe Ala Phe Glu Trp Arg Asp Pro Glu Met Gly Ile Ser
1445 1450 1455
Gly Gln Leu Thr Trp Thr Arg Leu Pro Gln Gly Phe Lys Asn Ser
1460 1465 1470
Pro Thr Leu Phe Asn Glu Ala Leu His Arg Asp Leu Ala Asp Phe
1475 1480 1485
Arg Ile Gln His Pro Asp Leu Ile Leu Leu Gln Tyr Val Asp Asp
1490 1495 1500
Leu Leu Leu Ala Ala Thr Ser Glu Leu Asp Cys Gln Gln Gly Thr
1505 1510 1515
Arg Ala Leu Leu Gln Thr Leu Gly Asn Leu Gly Tyr Arg Ala Ser
1520 1525 1530
Ala Lys Lys Ala Gln Ile Cys Gln Lys Gln Val Lys Tyr Leu Gly
1535 1540 1545
Tyr Leu Leu Lys Glu Gly Gln Arg Trp Leu Thr Glu Ala Arg Lys
1550 1555 1560
Glu Thr Val Met Gly Gln Pro Thr Pro Lys Thr Pro Arg Gln Leu
1565 1570 1575
Arg Glu Phe Leu Gly Lys Ala Gly Phe Cys Arg Leu Phe Ile Pro
1580 1585 1590
Gly Phe Ala Glu Met Ala Ala Pro Leu Tyr Pro Leu Thr Lys Pro
1595 1600 1605
Gly Thr Leu Phe Asn Trp Gly Pro Asp Gln Gln Lys Ala Tyr Gln
1610 1615 1620
Glu Ile Lys Gln Ala Leu Leu Thr Ala Pro Ala Leu Gly Leu Pro
1625 1630 1635
Asp Leu Thr Lys Pro Phe Glu Leu Phe Val Asp Glu Lys Gln Gly
1640 1645 1650
Tyr Ala Lys Gly Val Leu Thr Gln Lys Leu Gly Pro Trp Arg Arg
1655 1660 1665
Pro Val Ala Tyr Leu Ser Lys Lys Leu Asp Pro Val Ala Ala Gly
1670 1675 1680
Trp Pro Pro Cys Leu Arg Met Val Ala Ala Ile Ala Val Leu Thr
1685 1690 1695
Lys Asp Ala Gly Lys Leu Thr Met Gly Gln Pro Leu Val Ile Leu
1700 1705 1710
Ala Pro His Ala Val Glu Ala Leu Val Lys Gln Pro Pro Asp Arg
1715 1720 1725
Trp Leu Ser Asn Ala Arg Met Thr His Tyr Gln Ala Leu Leu Leu
1730 1735 1740
Asp Thr Asp Arg Val Gln Phe Gly Pro Val Val Ala Leu Asn Pro
1745 1750 1755
Ala Thr Leu Leu Pro Leu Pro Glu Glu Gly Leu Gln His Asn Cys
1760 1765 1770
Leu Asp Ile Leu Ala Glu Ala His Gly Thr Arg Pro Asp Leu Thr
1775 1780 1785
Asp Gln Pro Leu Pro Asp Ala Asp His Thr Trp Tyr Thr Asp Gly
1790 1795 1800
Ser Ser Leu Leu Gln Glu Gly Gln Arg Lys Ala Gly Ala Ala Val
1805 1810 1815
Thr Thr Glu Thr Glu Val Ile Trp Ala Lys Ala Leu Pro Ala Gly
1820 1825 1830
Thr Ser Ala Gln Arg Ala Glu Leu Ile Ala Leu Thr Gln Ala Leu
1835 1840 1845
Lys Met Ala Glu Gly Lys Lys Leu Asn Val Tyr Thr Asp Ser Arg
1850 1855 1860
Tyr Ala Phe Ala Thr Ala His Ile His Gly Glu Ile Tyr Arg Arg
1865 1870 1875
Arg Gly Trp Leu Thr Ser Glu Gly Lys Glu Ile Lys Asn Lys Asp
1880 1885 1890
Glu Ile Leu Ala Leu Leu Lys Ala Leu Phe Leu Pro Lys Arg Leu
1895 1900 1905
Ser Ile Ile His Cys Pro Gly His Gln Lys Gly His Ser Ala Glu
1910 1915 1920
Ala Arg Gly Asn Arg Met Ala Asp Gln Ala Ala Arg Lys Ala Ala
1925 1930 1935
Ile Thr Glu Thr Pro Asp Thr Ser Thr Leu Leu Ile Glu Asn Ser
1940 1945 1950
Ser Pro Ser Gly Gly Ser Lys Arg Thr Ala Asp Gly Ser Glu Phe
1955 1960 1965
Glu Pro Lys Lys Lys Arg Lys Val Gly Ser Gly
1970 1975
<210> 79
<211> 1963
<212> PRT
<213> 人工的
<220>
<223> 合成的REDRAW编辑器
<400> 79
Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys
1 5 10 15
Arg Lys Val Thr Leu Asn Ile Glu Asp Glu Tyr Arg Leu His Glu Thr
20 25 30
Ser Lys Glu Pro Asp Val Ser Leu Gly Ser Thr Trp Leu Ser Asp Phe
35 40 45
Pro Gln Ala Trp Ala Glu Thr Gly Gly Met Gly Leu Ala Val Arg Gln
50 55 60
Ala Pro Leu Ile Ile Pro Leu Lys Ala Thr Ser Thr Pro Val Ser Ile
65 70 75 80
Lys Gln Tyr Pro Met Ser Gln Glu Ala Arg Leu Gly Ile Lys Pro His
85 90 95
Ile Gln Arg Leu Leu Asp Gln Gly Ile Leu Val Pro Cys Gln Ser Pro
100 105 110
Trp Asn Thr Pro Leu Leu Pro Val Lys Lys Pro Gly Thr Asn Asp Tyr
115 120 125
Arg Pro Val Gln Asp Leu Arg Glu Val Asn Lys Arg Val Glu Asp Ile
130 135 140
His Pro Thr Val Pro Asn Pro Tyr Asn Leu Leu Ser Gly Leu Pro Pro
145 150 155 160
Ser His Gln Trp Tyr Thr Val Leu Asp Leu Lys Asp Ala Phe Phe Cys
165 170 175
Leu Arg Leu His Pro Thr Ser Gln Pro Leu Phe Ala Phe Glu Trp Arg
180 185 190
Asp Pro Glu Met Gly Ile Ser Gly Gln Leu Thr Trp Thr Arg Leu Pro
195 200 205
Gln Gly Phe Lys Asn Ser Pro Thr Leu Phe Asn Glu Ala Leu His Arg
210 215 220
Asp Leu Ala Asp Phe Arg Ile Gln His Pro Asp Leu Ile Leu Leu Gln
225 230 235 240
Tyr Val Asp Asp Leu Leu Leu Ala Ala Thr Ser Glu Leu Asp Cys Gln
245 250 255
Gln Gly Thr Arg Ala Leu Leu Gln Thr Leu Gly Asn Leu Gly Tyr Arg
260 265 270
Ala Ser Ala Lys Lys Ala Gln Ile Cys Gln Lys Gln Val Lys Tyr Leu
275 280 285
Gly Tyr Leu Leu Lys Glu Gly Gln Arg Trp Leu Thr Glu Ala Arg Lys
290 295 300
Glu Thr Val Met Gly Gln Pro Thr Pro Lys Thr Pro Arg Gln Leu Arg
305 310 315 320
Glu Phe Leu Gly Lys Ala Gly Phe Cys Arg Leu Phe Ile Pro Gly Phe
325 330 335
Ala Glu Met Ala Ala Pro Leu Tyr Pro Leu Thr Lys Pro Gly Thr Leu
340 345 350
Phe Asn Trp Gly Pro Asp Gln Gln Lys Ala Tyr Gln Glu Ile Lys Gln
355 360 365
Ala Leu Leu Thr Ala Pro Ala Leu Gly Leu Pro Asp Leu Thr Lys Pro
370 375 380
Phe Glu Leu Phe Val Asp Glu Lys Gln Gly Tyr Ala Lys Gly Val Leu
385 390 395 400
Thr Gln Lys Leu Gly Pro Trp Arg Arg Pro Val Ala Tyr Leu Ser Lys
405 410 415
Lys Leu Asp Pro Val Ala Ala Gly Trp Pro Pro Cys Leu Arg Met Val
420 425 430
Ala Ala Ile Ala Val Leu Thr Lys Asp Ala Gly Lys Leu Thr Met Gly
435 440 445
Gln Pro Leu Val Ile Leu Ala Pro His Ala Val Glu Ala Leu Val Lys
450 455 460
Gln Pro Pro Asp Arg Trp Leu Ser Asn Ala Arg Met Thr His Tyr Gln
465 470 475 480
Ala Leu Leu Leu Asp Thr Asp Arg Val Gln Phe Gly Pro Val Val Ala
485 490 495
Leu Asn Pro Ala Thr Leu Leu Pro Leu Pro Glu Glu Gly Leu Gln His
500 505 510
Asn Cys Leu Asp Ile Leu Ala Glu Ala His Gly Thr Arg Pro Asp Leu
515 520 525
Thr Asp Gln Pro Leu Pro Asp Ala Asp His Thr Trp Tyr Thr Asp Gly
530 535 540
Ser Ser Leu Leu Gln Glu Gly Gln Arg Lys Ala Gly Ala Ala Val Thr
545 550 555 560
Thr Glu Thr Glu Val Ile Trp Ala Lys Ala Leu Pro Ala Gly Thr Ser
565 570 575
Ala Gln Arg Ala Glu Leu Ile Ala Leu Thr Gln Ala Leu Lys Met Ala
580 585 590
Glu Gly Lys Lys Leu Asn Val Tyr Thr Asp Ser Arg Tyr Ala Phe Ala
595 600 605
Thr Ala His Ile His Gly Glu Ile Tyr Arg Arg Arg Gly Trp Leu Thr
610 615 620
Ser Glu Gly Lys Glu Ile Lys Asn Lys Asp Glu Ile Leu Ala Leu Leu
625 630 635 640
Lys Ala Leu Phe Leu Pro Lys Arg Leu Ser Ile Ile His Cys Pro Gly
645 650 655
His Gln Lys Gly His Ser Ala Glu Ala Arg Gly Asn Arg Met Ala Asp
660 665 670
Gln Ala Ala Arg Lys Ala Ala Ile Thr Glu Thr Pro Asp Thr Ser Thr
675 680 685
Leu Leu Ile Glu Asn Ser Ser Pro Ser Gly Ser Glu Thr Pro Gly Thr
690 695 700
Ser Glu Ser Ala Thr Pro Glu Ser Ser Lys Leu Glu Lys Phe Thr Asn
705 710 715 720
Cys Tyr Ser Leu Ser Lys Thr Leu Arg Phe Lys Ala Ile Pro Val Gly
725 730 735
Lys Thr Gln Glu Asn Ile Asp Asn Lys Arg Leu Leu Val Glu Asp Glu
740 745 750
Lys Arg Ala Glu Asp Tyr Lys Gly Val Lys Lys Leu Leu Asp Arg Tyr
755 760 765
Tyr Leu Ser Phe Ile Asn Asp Val Leu His Ser Ile Lys Leu Lys Asn
770 775 780
Leu Asn Asn Tyr Ile Ser Leu Phe Arg Lys Lys Thr Arg Thr Glu Lys
785 790 795 800
Glu Asn Lys Glu Leu Glu Asn Leu Glu Ile Asn Leu Arg Lys Glu Ile
805 810 815
Ala Lys Ala Phe Lys Gly Asn Glu Gly Tyr Lys Ser Leu Phe Lys Lys
820 825 830
Asp Ile Ile Glu Thr Ile Leu Pro Glu Phe Leu Asp Asp Lys Asp Glu
835 840 845
Ile Ala Leu Val Asn Ser Phe Asn Gly Phe Thr Thr Ala Phe Thr Gly
850 855 860
Phe Phe Asp Asn Arg Glu Asn Met Phe Ser Glu Glu Ala Lys Ser Thr
865 870 875 880
Ser Ile Ala Phe Arg Cys Ile Asn Glu Asn Leu Thr Arg Tyr Ile Ser
885 890 895
Asn Met Asp Ile Phe Glu Lys Val Asp Ala Ile Phe Asp Lys His Glu
900 905 910
Val Gln Glu Ile Lys Glu Lys Ile Leu Asn Ser Asp Tyr Asp Val Glu
915 920 925
Asp Phe Phe Glu Gly Glu Phe Phe Asn Phe Val Leu Thr Gln Glu Gly
930 935 940
Ile Asp Val Tyr Asn Ala Ile Ile Gly Gly Phe Val Thr Glu Ser Gly
945 950 955 960
Glu Lys Ile Lys Gly Leu Asn Glu Tyr Ile Asn Leu Tyr Asn Gln Lys
965 970 975
Thr Lys Gln Lys Leu Pro Lys Phe Lys Pro Leu Tyr Lys Gln Val Leu
980 985 990
Ser Asp Arg Glu Ser Leu Ser Phe Tyr Gly Glu Gly Tyr Thr Ser Asp
995 1000 1005
Glu Glu Val Leu Glu Val Phe Arg Asn Thr Leu Asn Lys Asn Ser
1010 1015 1020
Glu Ile Phe Ser Ser Ile Lys Lys Leu Glu Lys Leu Phe Lys Asn
1025 1030 1035
Phe Asp Glu Tyr Ser Ser Ala Gly Ile Phe Val Lys Asn Gly Pro
1040 1045 1050
Ala Ile Ser Thr Ile Ser Lys Asp Ile Phe Gly Glu Trp Asn Val
1055 1060 1065
Ile Arg Asp Lys Trp Asn Ala Glu Tyr Asp Asp Ile His Leu Lys
1070 1075 1080
Lys Lys Ala Val Val Thr Glu Lys Tyr Glu Asp Asp Arg Arg Lys
1085 1090 1095
Ser Phe Lys Lys Ile Gly Ser Phe Ser Leu Glu Gln Leu Gln Glu
1100 1105 1110
Tyr Ala Asp Ala Asp Leu Ser Val Val Glu Lys Leu Lys Glu Ile
1115 1120 1125
Ile Ile Gln Lys Val Asp Glu Ile Tyr Lys Val Tyr Gly Ser Ser
1130 1135 1140
Glu Lys Leu Phe Asp Ala Asp Phe Val Leu Glu Lys Ser Leu Lys
1145 1150 1155
Lys Asn Asp Ala Val Val Ala Ile Met Lys Asp Leu Leu Asp Ser
1160 1165 1170
Val Lys Ser Phe Glu Asn Tyr Ile Lys Ala Phe Phe Gly Glu Gly
1175 1180 1185
Lys Glu Thr Asn Arg Asp Glu Ser Phe Tyr Gly Asp Phe Val Leu
1190 1195 1200
Ala Tyr Asp Ile Leu Leu Lys Val Asp His Ile Tyr Asp Ala Ile
1205 1210 1215
Arg Asn Tyr Val Thr Gln Lys Pro Tyr Ser Lys Asp Lys Phe Lys
1220 1225 1230
Leu Tyr Phe Gln Asn Pro Gln Phe Met Arg Gly Trp Asp Lys Asp
1235 1240 1245
Val Glu Thr Asp Tyr Arg Ala Thr Ile Leu Arg Tyr Gly Ser Lys
1250 1255 1260
Tyr Tyr Leu Ala Ile Met Asp Lys Lys Tyr Ala Lys Cys Leu Gln
1265 1270 1275
Lys Ile Asp Lys Asp Asp Val Asn Gly Asn Tyr Glu Lys Ile Asn
1280 1285 1290
Tyr Lys Leu Leu Pro Gly Pro Asn Lys Met Leu Pro Arg Val Phe
1295 1300 1305
Phe Ser Lys Lys Trp Met Ala Tyr Tyr Asn Pro Ser Glu Asp Ile
1310 1315 1320
Gln Lys Ile Tyr Lys Asn Gly Thr Phe Lys Lys Gly Asp Met Phe
1325 1330 1335
Asn Leu Asn Asp Cys His Lys Leu Ile Asp Phe Phe Lys Asp Ser
1340 1345 1350
Ile Ser Arg Tyr Pro Lys Trp Ser Asn Ala Tyr Asp Phe Asn Phe
1355 1360 1365
Ser Glu Thr Glu Lys Tyr Lys Asp Ile Ala Gly Phe Tyr Arg Glu
1370 1375 1380
Val Glu Glu Gln Gly Tyr Lys Val Ser Phe Glu Ser Ala Ser Lys
1385 1390 1395
Lys Glu Val Asp Lys Leu Val Glu Glu Gly Lys Leu Tyr Met Phe
1400 1405 1410
Gln Ile Tyr Asn Lys Asp Phe Ser Asp Lys Ser His Gly Thr Pro
1415 1420 1425
Asn Leu His Thr Met Tyr Phe Lys Leu Leu Phe Asp Glu Asn Asn
1430 1435 1440
His Gly Gln Ile Arg Leu Ser Gly Gly Ala Glu Leu Phe Met Arg
1445 1450 1455
Arg Ala Ser Leu Lys Lys Glu Glu Leu Val Val Ala Pro Ala Asn
1460 1465 1470
Ser Pro Ile Ala Asn Lys Asn Pro Asp Asn Pro Lys Lys Thr Thr
1475 1480 1485
Thr Leu Ser Tyr Asp Val Tyr Lys Asp Lys Arg Phe Ser Glu Asp
1490 1495 1500
Gln Tyr Glu Leu His Ile Pro Ile Ala Ile Asn Lys Cys Pro Lys
1505 1510 1515
Asn Ile Phe Lys Ile Asn Thr Glu Val Arg Val Leu Leu Lys His
1520 1525 1530
Asp Asp Asn Pro Tyr Val Ile Gly Ile Asp Arg Gly Glu Arg Asn
1535 1540 1545
Leu Leu Tyr Ile Val Val Val Asp Gly Lys Gly Asn Ile Val Glu
1550 1555 1560
Gln Tyr Ser Leu Asn Glu Ile Ile Asn Asn Phe Asn Gly Ile Arg
1565 1570 1575
Ile Lys Thr Asp Tyr His Ser Leu Leu Asp Lys Lys Glu Lys Glu
1580 1585 1590
Arg Phe Glu Ala Arg Gln Asn Trp Thr Ser Ile Glu Asn Ile Lys
1595 1600 1605
Glu Leu Lys Ala Gly Tyr Ile Ser Gln Val Val His Lys Ile Cys
1610 1615 1620
Glu Leu Val Glu Lys Tyr Asp Ala Val Ile Ala Leu Glu Asp Leu
1625 1630 1635
Asn Ser Gly Phe Lys Asn Ser Arg Val Lys Val Glu Lys Gln Val
1640 1645 1650
Tyr Gln Lys Phe Glu Lys Met Leu Ile Asp Lys Leu Asn Tyr Met
1655 1660 1665
Val Asp Lys Lys Ser Asn Pro Cys Ala Thr Gly Gly Ala Leu Lys
1670 1675 1680
Gly Tyr Gln Ile Thr Asn Lys Phe Glu Ser Phe Lys Ser Met Ser
1685 1690 1695
Thr Gln Asn Gly Phe Ile Phe Tyr Ile Pro Ala Trp Leu Thr Ser
1700 1705 1710
Lys Ile Asp Pro Ser Thr Gly Phe Val Asn Leu Leu Lys Thr Lys
1715 1720 1725
Tyr Thr Ser Ile Ala Asp Ser Lys Lys Phe Ile Ser Ser Phe Asp
1730 1735 1740
Arg Ile Met Tyr Val Pro Glu Glu Asp Leu Phe Glu Phe Ala Leu
1745 1750 1755
Asp Tyr Lys Asn Phe Ser Arg Thr Asp Ala Asp Tyr Ile Lys Lys
1760 1765 1770
Trp Lys Leu Tyr Ser Tyr Gly Asn Arg Ile Arg Ile Phe Arg Asn
1775 1780 1785
Pro Lys Lys Asn Asn Val Phe Asp Trp Glu Glu Val Cys Leu Thr
1790 1795 1800
Ser Ala Tyr Lys Glu Leu Phe Asn Lys Tyr Gly Ile Asn Tyr Gln
1805 1810 1815
Gln Gly Asp Ile Arg Ala Leu Leu Cys Glu Gln Ser Asp Lys Ala
1820 1825 1830
Phe Tyr Ser Ser Phe Met Ala Leu Met Ser Leu Met Leu Gln Met
1835 1840 1845
Ala Asn Ser Ile Thr Gly Arg Thr Asp Val Asp Phe Leu Ile Ser
1850 1855 1860
Pro Val Lys Asn Ser Asp Gly Ile Phe Tyr Asp Ser Arg Asn Tyr
1865 1870 1875
Glu Ala Gln Glu Asn Ala Ile Leu Pro Lys Asn Ala Asp Ala Asn
1880 1885 1890
Gly Ala Tyr Asn Ile Ala Arg Lys Val Leu Trp Ala Ile Gly Gln
1895 1900 1905
Phe Lys Lys Ala Glu Asp Glu Lys Leu Asp Lys Val Lys Ile Ala
1910 1915 1920
Ile Ser Asn Lys Glu Trp Leu Glu Tyr Ala Gln Thr Ser Val Lys
1925 1930 1935
His Ser Gly Gly Ser Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu
1940 1945 1950
Pro Lys Lys Lys Arg Lys Val Gly Ser Gly
1955 1960
<210> 80
<211> 1979
<212> PRT
<213> 人工的
<220>
<223> 合成的REDRAW编辑器
<400> 80
Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys
1 5 10 15
Arg Lys Val Thr Leu Asn Ile Glu Asp Glu Tyr Arg Leu His Glu Thr
20 25 30
Ser Lys Glu Pro Asp Val Ser Leu Gly Ser Thr Trp Leu Ser Asp Phe
35 40 45
Pro Gln Ala Trp Ala Glu Thr Gly Gly Met Gly Leu Ala Val Arg Gln
50 55 60
Ala Pro Leu Ile Ile Pro Leu Lys Ala Thr Ser Thr Pro Val Ser Ile
65 70 75 80
Lys Gln Tyr Pro Met Ser Gln Glu Ala Arg Leu Gly Ile Lys Pro His
85 90 95
Ile Gln Arg Leu Leu Asp Gln Gly Ile Leu Val Pro Cys Gln Ser Pro
100 105 110
Trp Asn Thr Pro Leu Leu Pro Val Lys Lys Pro Gly Thr Asn Asp Tyr
115 120 125
Arg Pro Val Gln Asp Leu Arg Glu Val Asn Lys Arg Val Glu Asp Ile
130 135 140
His Pro Thr Val Pro Asn Pro Tyr Asn Leu Leu Ser Gly Leu Pro Pro
145 150 155 160
Ser His Gln Trp Tyr Thr Val Leu Asp Leu Lys Asp Ala Phe Phe Cys
165 170 175
Leu Arg Leu His Pro Thr Ser Gln Pro Leu Phe Ala Phe Glu Trp Arg
180 185 190
Asp Pro Glu Met Gly Ile Ser Gly Gln Leu Thr Trp Thr Arg Leu Pro
195 200 205
Gln Gly Phe Lys Asn Ser Pro Thr Leu Phe Asn Glu Ala Leu His Arg
210 215 220
Asp Leu Ala Asp Phe Arg Ile Gln His Pro Asp Leu Ile Leu Leu Gln
225 230 235 240
Tyr Val Asp Asp Leu Leu Leu Ala Ala Thr Ser Glu Leu Asp Cys Gln
245 250 255
Gln Gly Thr Arg Ala Leu Leu Gln Thr Leu Gly Asn Leu Gly Tyr Arg
260 265 270
Ala Ser Ala Lys Lys Ala Gln Ile Cys Gln Lys Gln Val Lys Tyr Leu
275 280 285
Gly Tyr Leu Leu Lys Glu Gly Gln Arg Trp Leu Thr Glu Ala Arg Lys
290 295 300
Glu Thr Val Met Gly Gln Pro Thr Pro Lys Thr Pro Arg Gln Leu Arg
305 310 315 320
Glu Phe Leu Gly Lys Ala Gly Phe Cys Arg Leu Phe Ile Pro Gly Phe
325 330 335
Ala Glu Met Ala Ala Pro Leu Tyr Pro Leu Thr Lys Pro Gly Thr Leu
340 345 350
Phe Asn Trp Gly Pro Asp Gln Gln Lys Ala Tyr Gln Glu Ile Lys Gln
355 360 365
Ala Leu Leu Thr Ala Pro Ala Leu Gly Leu Pro Asp Leu Thr Lys Pro
370 375 380
Phe Glu Leu Phe Val Asp Glu Lys Gln Gly Tyr Ala Lys Gly Val Leu
385 390 395 400
Thr Gln Lys Leu Gly Pro Trp Arg Arg Pro Val Ala Tyr Leu Ser Lys
405 410 415
Lys Leu Asp Pro Val Ala Ala Gly Trp Pro Pro Cys Leu Arg Met Val
420 425 430
Ala Ala Ile Ala Val Leu Thr Lys Asp Ala Gly Lys Leu Thr Met Gly
435 440 445
Gln Pro Leu Val Ile Leu Ala Pro His Ala Val Glu Ala Leu Val Lys
450 455 460
Gln Pro Pro Asp Arg Trp Leu Ser Asn Ala Arg Met Thr His Tyr Gln
465 470 475 480
Ala Leu Leu Leu Asp Thr Asp Arg Val Gln Phe Gly Pro Val Val Ala
485 490 495
Leu Asn Pro Ala Thr Leu Leu Pro Leu Pro Glu Glu Gly Leu Gln His
500 505 510
Asn Cys Leu Asp Ile Leu Ala Glu Ala His Gly Thr Arg Pro Asp Leu
515 520 525
Thr Asp Gln Pro Leu Pro Asp Ala Asp His Thr Trp Tyr Thr Asp Gly
530 535 540
Ser Ser Leu Leu Gln Glu Gly Gln Arg Lys Ala Gly Ala Ala Val Thr
545 550 555 560
Thr Glu Thr Glu Val Ile Trp Ala Lys Ala Leu Pro Ala Gly Thr Ser
565 570 575
Ala Gln Arg Ala Glu Leu Ile Ala Leu Thr Gln Ala Leu Lys Met Ala
580 585 590
Glu Gly Lys Lys Leu Asn Val Tyr Thr Asp Ser Arg Tyr Ala Phe Ala
595 600 605
Thr Ala His Ile His Gly Glu Ile Tyr Arg Arg Arg Gly Trp Leu Thr
610 615 620
Ser Glu Gly Lys Glu Ile Lys Asn Lys Asp Glu Ile Leu Ala Leu Leu
625 630 635 640
Lys Ala Leu Phe Leu Pro Lys Arg Leu Ser Ile Ile His Cys Pro Gly
645 650 655
His Gln Lys Gly His Ser Ala Glu Ala Arg Gly Asn Arg Met Ala Asp
660 665 670
Gln Ala Ala Arg Lys Ala Ala Ile Thr Glu Thr Pro Asp Thr Ser Thr
675 680 685
Leu Leu Ile Glu Asn Ser Ser Pro Ser Gly Gly Ser Ser Gly Gly Ser
690 695 700
Ser Gly Ser Glu Thr Pro Gly Thr Ser Glu Ser Ala Thr Pro Glu Ser
705 710 715 720
Ser Gly Gly Ser Ser Gly Gly Ser Ser Lys Leu Glu Lys Phe Thr Asn
725 730 735
Cys Tyr Ser Leu Ser Lys Thr Leu Arg Phe Lys Ala Ile Pro Val Gly
740 745 750
Lys Thr Gln Glu Asn Ile Asp Asn Lys Arg Leu Leu Val Glu Asp Glu
755 760 765
Lys Arg Ala Glu Asp Tyr Lys Gly Val Lys Lys Leu Leu Asp Arg Tyr
770 775 780
Tyr Leu Ser Phe Ile Asn Asp Val Leu His Ser Ile Lys Leu Lys Asn
785 790 795 800
Leu Asn Asn Tyr Ile Ser Leu Phe Arg Lys Lys Thr Arg Thr Glu Lys
805 810 815
Glu Asn Lys Glu Leu Glu Asn Leu Glu Ile Asn Leu Arg Lys Glu Ile
820 825 830
Ala Lys Ala Phe Lys Gly Asn Glu Gly Tyr Lys Ser Leu Phe Lys Lys
835 840 845
Asp Ile Ile Glu Thr Ile Leu Pro Glu Phe Leu Asp Asp Lys Asp Glu
850 855 860
Ile Ala Leu Val Asn Ser Phe Asn Gly Phe Thr Thr Ala Phe Thr Gly
865 870 875 880
Phe Phe Asp Asn Arg Glu Asn Met Phe Ser Glu Glu Ala Lys Ser Thr
885 890 895
Ser Ile Ala Phe Arg Cys Ile Asn Glu Asn Leu Thr Arg Tyr Ile Ser
900 905 910
Asn Met Asp Ile Phe Glu Lys Val Asp Ala Ile Phe Asp Lys His Glu
915 920 925
Val Gln Glu Ile Lys Glu Lys Ile Leu Asn Ser Asp Tyr Asp Val Glu
930 935 940
Asp Phe Phe Glu Gly Glu Phe Phe Asn Phe Val Leu Thr Gln Glu Gly
945 950 955 960
Ile Asp Val Tyr Asn Ala Ile Ile Gly Gly Phe Val Thr Glu Ser Gly
965 970 975
Glu Lys Ile Lys Gly Leu Asn Glu Tyr Ile Asn Leu Tyr Asn Gln Lys
980 985 990
Thr Lys Gln Lys Leu Pro Lys Phe Lys Pro Leu Tyr Lys Gln Val Leu
995 1000 1005
Ser Asp Arg Glu Ser Leu Ser Phe Tyr Gly Glu Gly Tyr Thr Ser
1010 1015 1020
Asp Glu Glu Val Leu Glu Val Phe Arg Asn Thr Leu Asn Lys Asn
1025 1030 1035
Ser Glu Ile Phe Ser Ser Ile Lys Lys Leu Glu Lys Leu Phe Lys
1040 1045 1050
Asn Phe Asp Glu Tyr Ser Ser Ala Gly Ile Phe Val Lys Asn Gly
1055 1060 1065
Pro Ala Ile Ser Thr Ile Ser Lys Asp Ile Phe Gly Glu Trp Asn
1070 1075 1080
Val Ile Arg Asp Lys Trp Asn Ala Glu Tyr Asp Asp Ile His Leu
1085 1090 1095
Lys Lys Lys Ala Val Val Thr Glu Lys Tyr Glu Asp Asp Arg Arg
1100 1105 1110
Lys Ser Phe Lys Lys Ile Gly Ser Phe Ser Leu Glu Gln Leu Gln
1115 1120 1125
Glu Tyr Ala Asp Ala Asp Leu Ser Val Val Glu Lys Leu Lys Glu
1130 1135 1140
Ile Ile Ile Gln Lys Val Asp Glu Ile Tyr Lys Val Tyr Gly Ser
1145 1150 1155
Ser Glu Lys Leu Phe Asp Ala Asp Phe Val Leu Glu Lys Ser Leu
1160 1165 1170
Lys Lys Asn Asp Ala Val Val Ala Ile Met Lys Asp Leu Leu Asp
1175 1180 1185
Ser Val Lys Ser Phe Glu Asn Tyr Ile Lys Ala Phe Phe Gly Glu
1190 1195 1200
Gly Lys Glu Thr Asn Arg Asp Glu Ser Phe Tyr Gly Asp Phe Val
1205 1210 1215
Leu Ala Tyr Asp Ile Leu Leu Lys Val Asp His Ile Tyr Asp Ala
1220 1225 1230
Ile Arg Asn Tyr Val Thr Gln Lys Pro Tyr Ser Lys Asp Lys Phe
1235 1240 1245
Lys Leu Tyr Phe Gln Asn Pro Gln Phe Met Arg Gly Trp Asp Lys
1250 1255 1260
Asp Val Glu Thr Asp Tyr Arg Ala Thr Ile Leu Arg Tyr Gly Ser
1265 1270 1275
Lys Tyr Tyr Leu Ala Ile Met Asp Lys Lys Tyr Ala Lys Cys Leu
1280 1285 1290
Gln Lys Ile Asp Lys Asp Asp Val Asn Gly Asn Tyr Glu Lys Ile
1295 1300 1305
Asn Tyr Lys Leu Leu Pro Gly Pro Asn Lys Met Leu Pro Arg Val
1310 1315 1320
Phe Phe Ser Lys Lys Trp Met Ala Tyr Tyr Asn Pro Ser Glu Asp
1325 1330 1335
Ile Gln Lys Ile Tyr Lys Asn Gly Thr Phe Lys Lys Gly Asp Met
1340 1345 1350
Phe Asn Leu Asn Asp Cys His Lys Leu Ile Asp Phe Phe Lys Asp
1355 1360 1365
Ser Ile Ser Arg Tyr Pro Lys Trp Ser Asn Ala Tyr Asp Phe Asn
1370 1375 1380
Phe Ser Glu Thr Glu Lys Tyr Lys Asp Ile Ala Gly Phe Tyr Arg
1385 1390 1395
Glu Val Glu Glu Gln Gly Tyr Lys Val Ser Phe Glu Ser Ala Ser
1400 1405 1410
Lys Lys Glu Val Asp Lys Leu Val Glu Glu Gly Lys Leu Tyr Met
1415 1420 1425
Phe Gln Ile Tyr Asn Lys Asp Phe Ser Asp Lys Ser His Gly Thr
1430 1435 1440
Pro Asn Leu His Thr Met Tyr Phe Lys Leu Leu Phe Asp Glu Asn
1445 1450 1455
Asn His Gly Gln Ile Arg Leu Ser Gly Gly Ala Glu Leu Phe Met
1460 1465 1470
Arg Arg Ala Ser Leu Lys Lys Glu Glu Leu Val Val Ala Pro Ala
1475 1480 1485
Asn Ser Pro Ile Ala Asn Lys Asn Pro Asp Asn Pro Lys Lys Thr
1490 1495 1500
Thr Thr Leu Ser Tyr Asp Val Tyr Lys Asp Lys Arg Phe Ser Glu
1505 1510 1515
Asp Gln Tyr Glu Leu His Ile Pro Ile Ala Ile Asn Lys Cys Pro
1520 1525 1530
Lys Asn Ile Phe Lys Ile Asn Thr Glu Val Arg Val Leu Leu Lys
1535 1540 1545
His Asp Asp Asn Pro Tyr Val Ile Gly Ile Asp Arg Gly Glu Arg
1550 1555 1560
Asn Leu Leu Tyr Ile Val Val Val Asp Gly Lys Gly Asn Ile Val
1565 1570 1575
Glu Gln Tyr Ser Leu Asn Glu Ile Ile Asn Asn Phe Asn Gly Ile
1580 1585 1590
Arg Ile Lys Thr Asp Tyr His Ser Leu Leu Asp Lys Lys Glu Lys
1595 1600 1605
Glu Arg Phe Glu Ala Arg Gln Asn Trp Thr Ser Ile Glu Asn Ile
1610 1615 1620
Lys Glu Leu Lys Ala Gly Tyr Ile Ser Gln Val Val His Lys Ile
1625 1630 1635
Cys Glu Leu Val Glu Lys Tyr Asp Ala Val Ile Ala Leu Glu Asp
1640 1645 1650
Leu Asn Ser Gly Phe Lys Asn Ser Arg Val Lys Val Glu Lys Gln
1655 1660 1665
Val Tyr Gln Lys Phe Glu Lys Met Leu Ile Asp Lys Leu Asn Tyr
1670 1675 1680
Met Val Asp Lys Lys Ser Asn Pro Cys Ala Thr Gly Gly Ala Leu
1685 1690 1695
Lys Gly Tyr Gln Ile Thr Asn Lys Phe Glu Ser Phe Lys Ser Met
1700 1705 1710
Ser Thr Gln Asn Gly Phe Ile Phe Tyr Ile Pro Ala Trp Leu Thr
1715 1720 1725
Ser Lys Ile Asp Pro Ser Thr Gly Phe Val Asn Leu Leu Lys Thr
1730 1735 1740
Lys Tyr Thr Ser Ile Ala Asp Ser Lys Lys Phe Ile Ser Ser Phe
1745 1750 1755
Asp Arg Ile Met Tyr Val Pro Glu Glu Asp Leu Phe Glu Phe Ala
1760 1765 1770
Leu Asp Tyr Lys Asn Phe Ser Arg Thr Asp Ala Asp Tyr Ile Lys
1775 1780 1785
Lys Trp Lys Leu Tyr Ser Tyr Gly Asn Arg Ile Arg Ile Phe Arg
1790 1795 1800
Asn Pro Lys Lys Asn Asn Val Phe Asp Trp Glu Glu Val Cys Leu
1805 1810 1815
Thr Ser Ala Tyr Lys Glu Leu Phe Asn Lys Tyr Gly Ile Asn Tyr
1820 1825 1830
Gln Gln Gly Asp Ile Arg Ala Leu Leu Cys Glu Gln Ser Asp Lys
1835 1840 1845
Ala Phe Tyr Ser Ser Phe Met Ala Leu Met Ser Leu Met Leu Gln
1850 1855 1860
Met Ala Asn Ser Ile Thr Gly Arg Thr Asp Val Asp Phe Leu Ile
1865 1870 1875
Ser Pro Val Lys Asn Ser Asp Gly Ile Phe Tyr Asp Ser Arg Asn
1880 1885 1890
Tyr Glu Ala Gln Glu Asn Ala Ile Leu Pro Lys Asn Ala Asp Ala
1895 1900 1905
Asn Gly Ala Tyr Asn Ile Ala Arg Lys Val Leu Trp Ala Ile Gly
1910 1915 1920
Gln Phe Lys Lys Ala Glu Asp Glu Lys Leu Asp Lys Val Lys Ile
1925 1930 1935
Ala Ile Ser Asn Lys Glu Trp Leu Glu Tyr Ala Gln Thr Ser Val
1940 1945 1950
Lys His Ser Gly Gly Ser Lys Arg Thr Ala Asp Gly Ser Glu Phe
1955 1960 1965
Glu Pro Lys Lys Lys Arg Lys Val Gly Ser Gly
1970 1975
<210> 81
<211> 1963
<212> PRT
<213> 人工的
<220>
<223> 合成的REDRAW编辑器
<400> 81
Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys
1 5 10 15
Arg Lys Val Ser Lys Leu Glu Lys Phe Thr Asn Cys Tyr Ser Leu Ser
20 25 30
Lys Thr Leu Arg Phe Lys Ala Ile Pro Val Gly Lys Thr Gln Glu Asn
35 40 45
Ile Asp Asn Lys Arg Leu Leu Val Glu Asp Glu Lys Arg Ala Glu Asp
50 55 60
Tyr Lys Gly Val Lys Lys Leu Leu Asp Arg Tyr Tyr Leu Ser Phe Ile
65 70 75 80
Asn Asp Val Leu His Ser Ile Lys Leu Lys Asn Leu Asn Asn Tyr Ile
85 90 95
Ser Leu Phe Arg Lys Lys Thr Arg Thr Glu Lys Glu Asn Lys Glu Leu
100 105 110
Glu Asn Leu Glu Ile Asn Leu Arg Lys Glu Ile Ala Lys Ala Phe Lys
115 120 125
Gly Asn Glu Gly Tyr Lys Ser Leu Phe Lys Lys Asp Ile Ile Glu Thr
130 135 140
Ile Leu Pro Glu Phe Leu Asp Asp Lys Asp Glu Ile Ala Leu Val Asn
145 150 155 160
Ser Phe Asn Gly Phe Thr Thr Ala Phe Thr Gly Phe Phe Asp Asn Arg
165 170 175
Glu Asn Met Phe Ser Glu Glu Ala Lys Ser Thr Ser Ile Ala Phe Arg
180 185 190
Cys Ile Asn Glu Asn Leu Thr Arg Tyr Ile Ser Asn Met Asp Ile Phe
195 200 205
Glu Lys Val Asp Ala Ile Phe Asp Lys His Glu Val Gln Glu Ile Lys
210 215 220
Glu Lys Ile Leu Asn Ser Asp Tyr Asp Val Glu Asp Phe Phe Glu Gly
225 230 235 240
Glu Phe Phe Asn Phe Val Leu Thr Gln Glu Gly Ile Asp Val Tyr Asn
245 250 255
Ala Ile Ile Gly Gly Phe Val Thr Glu Ser Gly Glu Lys Ile Lys Gly
260 265 270
Leu Asn Glu Tyr Ile Asn Leu Tyr Asn Gln Lys Thr Lys Gln Lys Leu
275 280 285
Pro Lys Phe Lys Pro Leu Tyr Lys Gln Val Leu Ser Asp Arg Glu Ser
290 295 300
Leu Ser Phe Tyr Gly Glu Gly Tyr Thr Ser Asp Glu Glu Val Leu Glu
305 310 315 320
Val Phe Arg Asn Thr Leu Asn Lys Asn Ser Glu Ile Phe Ser Ser Ile
325 330 335
Lys Lys Leu Glu Lys Leu Phe Lys Asn Phe Asp Glu Tyr Ser Ser Ala
340 345 350
Gly Ile Phe Val Lys Asn Gly Pro Ala Ile Ser Thr Ile Ser Lys Asp
355 360 365
Ile Phe Gly Glu Trp Asn Val Ile Arg Asp Lys Trp Asn Ala Glu Tyr
370 375 380
Asp Asp Ile His Leu Lys Lys Lys Ala Val Val Thr Glu Lys Tyr Glu
385 390 395 400
Asp Asp Arg Arg Lys Ser Phe Lys Lys Ile Gly Ser Phe Ser Leu Glu
405 410 415
Gln Leu Gln Glu Tyr Ala Asp Ala Asp Leu Ser Val Val Glu Lys Leu
420 425 430
Lys Glu Ile Ile Ile Gln Lys Val Asp Glu Ile Tyr Lys Val Tyr Gly
435 440 445
Ser Ser Glu Lys Leu Phe Asp Ala Asp Phe Val Leu Glu Lys Ser Leu
450 455 460
Lys Lys Asn Asp Ala Val Val Ala Ile Met Lys Asp Leu Leu Asp Ser
465 470 475 480
Val Lys Ser Phe Glu Asn Tyr Ile Lys Ala Phe Phe Gly Glu Gly Lys
485 490 495
Glu Thr Asn Arg Asp Glu Ser Phe Tyr Gly Asp Phe Val Leu Ala Tyr
500 505 510
Asp Ile Leu Leu Lys Val Asp His Ile Tyr Asp Ala Ile Arg Asn Tyr
515 520 525
Val Thr Gln Lys Pro Tyr Ser Lys Asp Lys Phe Lys Leu Tyr Phe Gln
530 535 540
Asn Pro Gln Phe Met Arg Gly Trp Asp Lys Asp Val Glu Thr Asp Tyr
545 550 555 560
Arg Ala Thr Ile Leu Arg Tyr Gly Ser Lys Tyr Tyr Leu Ala Ile Met
565 570 575
Asp Lys Lys Tyr Ala Lys Cys Leu Gln Lys Ile Asp Lys Asp Asp Val
580 585 590
Asn Gly Asn Tyr Glu Lys Ile Asn Tyr Lys Leu Leu Pro Gly Pro Asn
595 600 605
Lys Met Leu Pro Arg Val Phe Phe Ser Lys Lys Trp Met Ala Tyr Tyr
610 615 620
Asn Pro Ser Glu Asp Ile Gln Lys Ile Tyr Lys Asn Gly Thr Phe Lys
625 630 635 640
Lys Gly Asp Met Phe Asn Leu Asn Asp Cys His Lys Leu Ile Asp Phe
645 650 655
Phe Lys Asp Ser Ile Ser Arg Tyr Pro Lys Trp Ser Asn Ala Tyr Asp
660 665 670
Phe Asn Phe Ser Glu Thr Glu Lys Tyr Lys Asp Ile Ala Gly Phe Tyr
675 680 685
Arg Glu Val Glu Glu Gln Gly Tyr Lys Val Ser Phe Glu Ser Ala Ser
690 695 700
Lys Lys Glu Val Asp Lys Leu Val Glu Glu Gly Lys Leu Tyr Met Phe
705 710 715 720
Gln Ile Tyr Asn Lys Asp Phe Ser Asp Lys Ser His Gly Thr Pro Asn
725 730 735
Leu His Thr Met Tyr Phe Lys Leu Leu Phe Asp Glu Asn Asn His Gly
740 745 750
Gln Ile Arg Leu Ser Gly Gly Ala Glu Leu Phe Met Arg Arg Ala Ser
755 760 765
Leu Lys Lys Glu Glu Leu Val Val Ala Pro Ala Asn Ser Pro Ile Ala
770 775 780
Asn Lys Asn Pro Asp Asn Pro Lys Lys Thr Thr Thr Leu Ser Tyr Asp
785 790 795 800
Val Tyr Lys Asp Lys Arg Phe Ser Glu Asp Gln Tyr Glu Leu His Ile
805 810 815
Pro Ile Ala Ile Asn Lys Cys Pro Lys Asn Ile Phe Lys Ile Asn Thr
820 825 830
Glu Val Arg Val Leu Leu Lys His Asp Asp Asn Pro Tyr Val Ile Gly
835 840 845
Ile Asp Arg Gly Glu Arg Asn Leu Leu Tyr Ile Val Val Val Asp Gly
850 855 860
Lys Gly Asn Ile Val Glu Gln Tyr Ser Leu Asn Glu Ile Ile Asn Asn
865 870 875 880
Phe Asn Gly Ile Arg Ile Lys Thr Asp Tyr His Ser Leu Leu Asp Lys
885 890 895
Lys Glu Lys Glu Arg Phe Glu Ala Arg Gln Asn Trp Thr Ser Ile Glu
900 905 910
Asn Ile Lys Glu Leu Lys Ala Gly Tyr Ile Ser Gln Val Val His Lys
915 920 925
Ile Cys Glu Leu Val Glu Lys Tyr Asp Ala Val Ile Ala Leu Glu Asp
930 935 940
Leu Asn Ser Gly Phe Lys Asn Ser Arg Val Lys Val Glu Lys Gln Val
945 950 955 960
Tyr Gln Lys Phe Glu Lys Met Leu Ile Asp Lys Leu Asn Tyr Met Val
965 970 975
Asp Lys Lys Ser Asn Pro Cys Ala Thr Gly Gly Ala Leu Lys Gly Tyr
980 985 990
Gln Ile Thr Asn Lys Phe Glu Ser Phe Lys Ser Met Ser Thr Gln Asn
995 1000 1005
Gly Phe Ile Phe Tyr Ile Pro Ala Trp Leu Thr Ser Lys Ile Asp
1010 1015 1020
Pro Ser Thr Gly Phe Val Asn Leu Leu Lys Thr Lys Tyr Thr Ser
1025 1030 1035
Ile Ala Asp Ser Lys Lys Phe Ile Ser Ser Phe Asp Arg Ile Met
1040 1045 1050
Tyr Val Pro Glu Glu Asp Leu Phe Glu Phe Ala Leu Asp Tyr Lys
1055 1060 1065
Asn Phe Ser Arg Thr Asp Ala Asp Tyr Ile Lys Lys Trp Lys Leu
1070 1075 1080
Tyr Ser Tyr Gly Asn Arg Ile Arg Ile Phe Arg Asn Pro Lys Lys
1085 1090 1095
Asn Asn Val Phe Asp Trp Glu Glu Val Cys Leu Thr Ser Ala Tyr
1100 1105 1110
Lys Glu Leu Phe Asn Lys Tyr Gly Ile Asn Tyr Gln Gln Gly Asp
1115 1120 1125
Ile Arg Ala Leu Leu Cys Glu Gln Ser Asp Lys Ala Phe Tyr Ser
1130 1135 1140
Ser Phe Met Ala Leu Met Ser Leu Met Leu Gln Met Ala Asn Ser
1145 1150 1155
Ile Thr Gly Arg Thr Asp Val Asp Phe Leu Ile Ser Pro Val Lys
1160 1165 1170
Asn Ser Asp Gly Ile Phe Tyr Asp Ser Arg Asn Tyr Glu Ala Gln
1175 1180 1185
Glu Asn Ala Ile Leu Pro Lys Asn Ala Asp Ala Asn Gly Ala Tyr
1190 1195 1200
Asn Ile Ala Arg Lys Val Leu Trp Ala Ile Gly Gln Phe Lys Lys
1205 1210 1215
Ala Glu Asp Glu Lys Leu Asp Lys Val Lys Ile Ala Ile Ser Asn
1220 1225 1230
Lys Glu Trp Leu Glu Tyr Ala Gln Thr Ser Val Lys His Ser Gly
1235 1240 1245
Ser Glu Thr Pro Gly Thr Ser Glu Ser Ala Thr Pro Glu Ser Thr
1250 1255 1260
Leu Asn Ile Glu Asp Glu Tyr Arg Leu His Glu Thr Ser Lys Glu
1265 1270 1275
Pro Asp Val Ser Leu Gly Ser Thr Trp Leu Ser Asp Phe Pro Gln
1280 1285 1290
Ala Trp Ala Glu Thr Gly Gly Met Gly Leu Ala Val Arg Gln Ala
1295 1300 1305
Pro Leu Ile Ile Pro Leu Lys Ala Thr Ser Thr Pro Val Ser Ile
1310 1315 1320
Lys Gln Tyr Pro Met Ser Gln Glu Ala Arg Leu Gly Ile Lys Pro
1325 1330 1335
His Ile Gln Arg Leu Leu Asp Gln Gly Ile Leu Val Pro Cys Gln
1340 1345 1350
Ser Pro Trp Asn Thr Pro Leu Leu Pro Val Lys Lys Pro Gly Thr
1355 1360 1365
Asn Asp Tyr Arg Pro Val Gln Asp Leu Arg Glu Val Asn Lys Arg
1370 1375 1380
Val Glu Asp Ile His Pro Thr Val Pro Asn Pro Tyr Asn Leu Leu
1385 1390 1395
Ser Gly Leu Pro Pro Ser His Gln Trp Tyr Thr Val Leu Asp Leu
1400 1405 1410
Lys Asp Ala Phe Phe Cys Leu Arg Leu His Pro Thr Ser Gln Pro
1415 1420 1425
Leu Phe Ala Phe Glu Trp Arg Asp Pro Glu Met Gly Ile Ser Gly
1430 1435 1440
Gln Leu Thr Trp Thr Arg Leu Pro Gln Gly Phe Lys Asn Ser Pro
1445 1450 1455
Thr Leu Phe Asn Glu Ala Leu His Arg Asp Leu Ala Asp Phe Arg
1460 1465 1470
Ile Gln His Pro Asp Leu Ile Leu Leu Gln Tyr Val Asp Asp Leu
1475 1480 1485
Leu Leu Ala Ala Thr Ser Glu Leu Asp Cys Gln Gln Gly Thr Arg
1490 1495 1500
Ala Leu Leu Gln Thr Leu Gly Asn Leu Gly Tyr Arg Ala Ser Ala
1505 1510 1515
Lys Lys Ala Gln Ile Cys Gln Lys Gln Val Lys Tyr Leu Gly Tyr
1520 1525 1530
Leu Leu Lys Glu Gly Gln Arg Trp Leu Thr Glu Ala Arg Lys Glu
1535 1540 1545
Thr Val Met Gly Gln Pro Thr Pro Lys Thr Pro Arg Gln Leu Arg
1550 1555 1560
Glu Phe Leu Gly Lys Ala Gly Phe Cys Arg Leu Phe Ile Pro Gly
1565 1570 1575
Phe Ala Glu Met Ala Ala Pro Leu Tyr Pro Leu Thr Lys Pro Gly
1580 1585 1590
Thr Leu Phe Asn Trp Gly Pro Asp Gln Gln Lys Ala Tyr Gln Glu
1595 1600 1605
Ile Lys Gln Ala Leu Leu Thr Ala Pro Ala Leu Gly Leu Pro Asp
1610 1615 1620
Leu Thr Lys Pro Phe Glu Leu Phe Val Asp Glu Lys Gln Gly Tyr
1625 1630 1635
Ala Lys Gly Val Leu Thr Gln Lys Leu Gly Pro Trp Arg Arg Pro
1640 1645 1650
Val Ala Tyr Leu Ser Lys Lys Leu Asp Pro Val Ala Ala Gly Trp
1655 1660 1665
Pro Pro Cys Leu Arg Met Val Ala Ala Ile Ala Val Leu Thr Lys
1670 1675 1680
Asp Ala Gly Lys Leu Thr Met Gly Gln Pro Leu Val Ile Leu Ala
1685 1690 1695
Pro His Ala Val Glu Ala Leu Val Lys Gln Pro Pro Asp Arg Trp
1700 1705 1710
Leu Ser Asn Ala Arg Met Thr His Tyr Gln Ala Leu Leu Leu Asp
1715 1720 1725
Thr Asp Arg Val Gln Phe Gly Pro Val Val Ala Leu Asn Pro Ala
1730 1735 1740
Thr Leu Leu Pro Leu Pro Glu Glu Gly Leu Gln His Asn Cys Leu
1745 1750 1755
Asp Ile Leu Ala Glu Ala His Gly Thr Arg Pro Asp Leu Thr Asp
1760 1765 1770
Gln Pro Leu Pro Asp Ala Asp His Thr Trp Tyr Thr Asp Gly Ser
1775 1780 1785
Ser Leu Leu Gln Glu Gly Gln Arg Lys Ala Gly Ala Ala Val Thr
1790 1795 1800
Thr Glu Thr Glu Val Ile Trp Ala Lys Ala Leu Pro Ala Gly Thr
1805 1810 1815
Ser Ala Gln Arg Ala Glu Leu Ile Ala Leu Thr Gln Ala Leu Lys
1820 1825 1830
Met Ala Glu Gly Lys Lys Leu Asn Val Tyr Thr Asp Ser Arg Tyr
1835 1840 1845
Ala Phe Ala Thr Ala His Ile His Gly Glu Ile Tyr Arg Arg Arg
1850 1855 1860
Gly Trp Leu Thr Ser Glu Gly Lys Glu Ile Lys Asn Lys Asp Glu
1865 1870 1875
Ile Leu Ala Leu Leu Lys Ala Leu Phe Leu Pro Lys Arg Leu Ser
1880 1885 1890
Ile Ile His Cys Pro Gly His Gln Lys Gly His Ser Ala Glu Ala
1895 1900 1905
Arg Gly Asn Arg Met Ala Asp Gln Ala Ala Arg Lys Ala Ala Ile
1910 1915 1920
Thr Glu Thr Pro Asp Thr Ser Thr Leu Leu Ile Glu Asn Ser Ser
1925 1930 1935
Pro Ser Gly Gly Ser Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu
1940 1945 1950
Pro Lys Lys Lys Arg Lys Val Gly Ser Gly
1955 1960
<210> 82
<211> 1979
<212> PRT
<213> 人工的
<220>
<223> 合成的REDRAW编辑器
<400> 82
Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys
1 5 10 15
Arg Lys Val Ser Lys Leu Glu Lys Phe Thr Asn Cys Tyr Ser Leu Ser
20 25 30
Lys Thr Leu Arg Phe Lys Ala Ile Pro Val Gly Lys Thr Gln Glu Asn
35 40 45
Ile Asp Asn Lys Arg Leu Leu Val Glu Asp Glu Lys Arg Ala Glu Asp
50 55 60
Tyr Lys Gly Val Lys Lys Leu Leu Asp Arg Tyr Tyr Leu Ser Phe Ile
65 70 75 80
Asn Asp Val Leu His Ser Ile Lys Leu Lys Asn Leu Asn Asn Tyr Ile
85 90 95
Ser Leu Phe Arg Lys Lys Thr Arg Thr Glu Lys Glu Asn Lys Glu Leu
100 105 110
Glu Asn Leu Glu Ile Asn Leu Arg Lys Glu Ile Ala Lys Ala Phe Lys
115 120 125
Gly Asn Glu Gly Tyr Lys Ser Leu Phe Lys Lys Asp Ile Ile Glu Thr
130 135 140
Ile Leu Pro Glu Phe Leu Asp Asp Lys Asp Glu Ile Ala Leu Val Asn
145 150 155 160
Ser Phe Asn Gly Phe Thr Thr Ala Phe Thr Gly Phe Phe Asp Asn Arg
165 170 175
Glu Asn Met Phe Ser Glu Glu Ala Lys Ser Thr Ser Ile Ala Phe Arg
180 185 190
Cys Ile Asn Glu Asn Leu Thr Arg Tyr Ile Ser Asn Met Asp Ile Phe
195 200 205
Glu Lys Val Asp Ala Ile Phe Asp Lys His Glu Val Gln Glu Ile Lys
210 215 220
Glu Lys Ile Leu Asn Ser Asp Tyr Asp Val Glu Asp Phe Phe Glu Gly
225 230 235 240
Glu Phe Phe Asn Phe Val Leu Thr Gln Glu Gly Ile Asp Val Tyr Asn
245 250 255
Ala Ile Ile Gly Gly Phe Val Thr Glu Ser Gly Glu Lys Ile Lys Gly
260 265 270
Leu Asn Glu Tyr Ile Asn Leu Tyr Asn Gln Lys Thr Lys Gln Lys Leu
275 280 285
Pro Lys Phe Lys Pro Leu Tyr Lys Gln Val Leu Ser Asp Arg Glu Ser
290 295 300
Leu Ser Phe Tyr Gly Glu Gly Tyr Thr Ser Asp Glu Glu Val Leu Glu
305 310 315 320
Val Phe Arg Asn Thr Leu Asn Lys Asn Ser Glu Ile Phe Ser Ser Ile
325 330 335
Lys Lys Leu Glu Lys Leu Phe Lys Asn Phe Asp Glu Tyr Ser Ser Ala
340 345 350
Gly Ile Phe Val Lys Asn Gly Pro Ala Ile Ser Thr Ile Ser Lys Asp
355 360 365
Ile Phe Gly Glu Trp Asn Val Ile Arg Asp Lys Trp Asn Ala Glu Tyr
370 375 380
Asp Asp Ile His Leu Lys Lys Lys Ala Val Val Thr Glu Lys Tyr Glu
385 390 395 400
Asp Asp Arg Arg Lys Ser Phe Lys Lys Ile Gly Ser Phe Ser Leu Glu
405 410 415
Gln Leu Gln Glu Tyr Ala Asp Ala Asp Leu Ser Val Val Glu Lys Leu
420 425 430
Lys Glu Ile Ile Ile Gln Lys Val Asp Glu Ile Tyr Lys Val Tyr Gly
435 440 445
Ser Ser Glu Lys Leu Phe Asp Ala Asp Phe Val Leu Glu Lys Ser Leu
450 455 460
Lys Lys Asn Asp Ala Val Val Ala Ile Met Lys Asp Leu Leu Asp Ser
465 470 475 480
Val Lys Ser Phe Glu Asn Tyr Ile Lys Ala Phe Phe Gly Glu Gly Lys
485 490 495
Glu Thr Asn Arg Asp Glu Ser Phe Tyr Gly Asp Phe Val Leu Ala Tyr
500 505 510
Asp Ile Leu Leu Lys Val Asp His Ile Tyr Asp Ala Ile Arg Asn Tyr
515 520 525
Val Thr Gln Lys Pro Tyr Ser Lys Asp Lys Phe Lys Leu Tyr Phe Gln
530 535 540
Asn Pro Gln Phe Met Arg Gly Trp Asp Lys Asp Val Glu Thr Asp Tyr
545 550 555 560
Arg Ala Thr Ile Leu Arg Tyr Gly Ser Lys Tyr Tyr Leu Ala Ile Met
565 570 575
Asp Lys Lys Tyr Ala Lys Cys Leu Gln Lys Ile Asp Lys Asp Asp Val
580 585 590
Asn Gly Asn Tyr Glu Lys Ile Asn Tyr Lys Leu Leu Pro Gly Pro Asn
595 600 605
Lys Met Leu Pro Arg Val Phe Phe Ser Lys Lys Trp Met Ala Tyr Tyr
610 615 620
Asn Pro Ser Glu Asp Ile Gln Lys Ile Tyr Lys Asn Gly Thr Phe Lys
625 630 635 640
Lys Gly Asp Met Phe Asn Leu Asn Asp Cys His Lys Leu Ile Asp Phe
645 650 655
Phe Lys Asp Ser Ile Ser Arg Tyr Pro Lys Trp Ser Asn Ala Tyr Asp
660 665 670
Phe Asn Phe Ser Glu Thr Glu Lys Tyr Lys Asp Ile Ala Gly Phe Tyr
675 680 685
Arg Glu Val Glu Glu Gln Gly Tyr Lys Val Ser Phe Glu Ser Ala Ser
690 695 700
Lys Lys Glu Val Asp Lys Leu Val Glu Glu Gly Lys Leu Tyr Met Phe
705 710 715 720
Gln Ile Tyr Asn Lys Asp Phe Ser Asp Lys Ser His Gly Thr Pro Asn
725 730 735
Leu His Thr Met Tyr Phe Lys Leu Leu Phe Asp Glu Asn Asn His Gly
740 745 750
Gln Ile Arg Leu Ser Gly Gly Ala Glu Leu Phe Met Arg Arg Ala Ser
755 760 765
Leu Lys Lys Glu Glu Leu Val Val Ala Pro Ala Asn Ser Pro Ile Ala
770 775 780
Asn Lys Asn Pro Asp Asn Pro Lys Lys Thr Thr Thr Leu Ser Tyr Asp
785 790 795 800
Val Tyr Lys Asp Lys Arg Phe Ser Glu Asp Gln Tyr Glu Leu His Ile
805 810 815
Pro Ile Ala Ile Asn Lys Cys Pro Lys Asn Ile Phe Lys Ile Asn Thr
820 825 830
Glu Val Arg Val Leu Leu Lys His Asp Asp Asn Pro Tyr Val Ile Gly
835 840 845
Ile Asp Arg Gly Glu Arg Asn Leu Leu Tyr Ile Val Val Val Asp Gly
850 855 860
Lys Gly Asn Ile Val Glu Gln Tyr Ser Leu Asn Glu Ile Ile Asn Asn
865 870 875 880
Phe Asn Gly Ile Arg Ile Lys Thr Asp Tyr His Ser Leu Leu Asp Lys
885 890 895
Lys Glu Lys Glu Arg Phe Glu Ala Arg Gln Asn Trp Thr Ser Ile Glu
900 905 910
Asn Ile Lys Glu Leu Lys Ala Gly Tyr Ile Ser Gln Val Val His Lys
915 920 925
Ile Cys Glu Leu Val Glu Lys Tyr Asp Ala Val Ile Ala Leu Glu Asp
930 935 940
Leu Asn Ser Gly Phe Lys Asn Ser Arg Val Lys Val Glu Lys Gln Val
945 950 955 960
Tyr Gln Lys Phe Glu Lys Met Leu Ile Asp Lys Leu Asn Tyr Met Val
965 970 975
Asp Lys Lys Ser Asn Pro Cys Ala Thr Gly Gly Ala Leu Lys Gly Tyr
980 985 990
Gln Ile Thr Asn Lys Phe Glu Ser Phe Lys Ser Met Ser Thr Gln Asn
995 1000 1005
Gly Phe Ile Phe Tyr Ile Pro Ala Trp Leu Thr Ser Lys Ile Asp
1010 1015 1020
Pro Ser Thr Gly Phe Val Asn Leu Leu Lys Thr Lys Tyr Thr Ser
1025 1030 1035
Ile Ala Asp Ser Lys Lys Phe Ile Ser Ser Phe Asp Arg Ile Met
1040 1045 1050
Tyr Val Pro Glu Glu Asp Leu Phe Glu Phe Ala Leu Asp Tyr Lys
1055 1060 1065
Asn Phe Ser Arg Thr Asp Ala Asp Tyr Ile Lys Lys Trp Lys Leu
1070 1075 1080
Tyr Ser Tyr Gly Asn Arg Ile Arg Ile Phe Arg Asn Pro Lys Lys
1085 1090 1095
Asn Asn Val Phe Asp Trp Glu Glu Val Cys Leu Thr Ser Ala Tyr
1100 1105 1110
Lys Glu Leu Phe Asn Lys Tyr Gly Ile Asn Tyr Gln Gln Gly Asp
1115 1120 1125
Ile Arg Ala Leu Leu Cys Glu Gln Ser Asp Lys Ala Phe Tyr Ser
1130 1135 1140
Ser Phe Met Ala Leu Met Ser Leu Met Leu Gln Met Ala Asn Ser
1145 1150 1155
Ile Thr Gly Arg Thr Asp Val Asp Phe Leu Ile Ser Pro Val Lys
1160 1165 1170
Asn Ser Asp Gly Ile Phe Tyr Asp Ser Arg Asn Tyr Glu Ala Gln
1175 1180 1185
Glu Asn Ala Ile Leu Pro Lys Asn Ala Asp Ala Asn Gly Ala Tyr
1190 1195 1200
Asn Ile Ala Arg Lys Val Leu Trp Ala Ile Gly Gln Phe Lys Lys
1205 1210 1215
Ala Glu Asp Glu Lys Leu Asp Lys Val Lys Ile Ala Ile Ser Asn
1220 1225 1230
Lys Glu Trp Leu Glu Tyr Ala Gln Thr Ser Val Lys His Ser Gly
1235 1240 1245
Gly Ser Ser Gly Gly Ser Ser Gly Ser Glu Thr Pro Gly Thr Ser
1250 1255 1260
Glu Ser Ala Thr Pro Glu Ser Ser Gly Gly Ser Ser Gly Gly Ser
1265 1270 1275
Thr Leu Asn Ile Glu Asp Glu Tyr Arg Leu His Glu Thr Ser Lys
1280 1285 1290
Glu Pro Asp Val Ser Leu Gly Ser Thr Trp Leu Ser Asp Phe Pro
1295 1300 1305
Gln Ala Trp Ala Glu Thr Gly Gly Met Gly Leu Ala Val Arg Gln
1310 1315 1320
Ala Pro Leu Ile Ile Pro Leu Lys Ala Thr Ser Thr Pro Val Ser
1325 1330 1335
Ile Lys Gln Tyr Pro Met Ser Gln Glu Ala Arg Leu Gly Ile Lys
1340 1345 1350
Pro His Ile Gln Arg Leu Leu Asp Gln Gly Ile Leu Val Pro Cys
1355 1360 1365
Gln Ser Pro Trp Asn Thr Pro Leu Leu Pro Val Lys Lys Pro Gly
1370 1375 1380
Thr Asn Asp Tyr Arg Pro Val Gln Asp Leu Arg Glu Val Asn Lys
1385 1390 1395
Arg Val Glu Asp Ile His Pro Thr Val Pro Asn Pro Tyr Asn Leu
1400 1405 1410
Leu Ser Gly Leu Pro Pro Ser His Gln Trp Tyr Thr Val Leu Asp
1415 1420 1425
Leu Lys Asp Ala Phe Phe Cys Leu Arg Leu His Pro Thr Ser Gln
1430 1435 1440
Pro Leu Phe Ala Phe Glu Trp Arg Asp Pro Glu Met Gly Ile Ser
1445 1450 1455
Gly Gln Leu Thr Trp Thr Arg Leu Pro Gln Gly Phe Lys Asn Ser
1460 1465 1470
Pro Thr Leu Phe Asn Glu Ala Leu His Arg Asp Leu Ala Asp Phe
1475 1480 1485
Arg Ile Gln His Pro Asp Leu Ile Leu Leu Gln Tyr Val Asp Asp
1490 1495 1500
Leu Leu Leu Ala Ala Thr Ser Glu Leu Asp Cys Gln Gln Gly Thr
1505 1510 1515
Arg Ala Leu Leu Gln Thr Leu Gly Asn Leu Gly Tyr Arg Ala Ser
1520 1525 1530
Ala Lys Lys Ala Gln Ile Cys Gln Lys Gln Val Lys Tyr Leu Gly
1535 1540 1545
Tyr Leu Leu Lys Glu Gly Gln Arg Trp Leu Thr Glu Ala Arg Lys
1550 1555 1560
Glu Thr Val Met Gly Gln Pro Thr Pro Lys Thr Pro Arg Gln Leu
1565 1570 1575
Arg Glu Phe Leu Gly Lys Ala Gly Phe Cys Arg Leu Phe Ile Pro
1580 1585 1590
Gly Phe Ala Glu Met Ala Ala Pro Leu Tyr Pro Leu Thr Lys Pro
1595 1600 1605
Gly Thr Leu Phe Asn Trp Gly Pro Asp Gln Gln Lys Ala Tyr Gln
1610 1615 1620
Glu Ile Lys Gln Ala Leu Leu Thr Ala Pro Ala Leu Gly Leu Pro
1625 1630 1635
Asp Leu Thr Lys Pro Phe Glu Leu Phe Val Asp Glu Lys Gln Gly
1640 1645 1650
Tyr Ala Lys Gly Val Leu Thr Gln Lys Leu Gly Pro Trp Arg Arg
1655 1660 1665
Pro Val Ala Tyr Leu Ser Lys Lys Leu Asp Pro Val Ala Ala Gly
1670 1675 1680
Trp Pro Pro Cys Leu Arg Met Val Ala Ala Ile Ala Val Leu Thr
1685 1690 1695
Lys Asp Ala Gly Lys Leu Thr Met Gly Gln Pro Leu Val Ile Leu
1700 1705 1710
Ala Pro His Ala Val Glu Ala Leu Val Lys Gln Pro Pro Asp Arg
1715 1720 1725
Trp Leu Ser Asn Ala Arg Met Thr His Tyr Gln Ala Leu Leu Leu
1730 1735 1740
Asp Thr Asp Arg Val Gln Phe Gly Pro Val Val Ala Leu Asn Pro
1745 1750 1755
Ala Thr Leu Leu Pro Leu Pro Glu Glu Gly Leu Gln His Asn Cys
1760 1765 1770
Leu Asp Ile Leu Ala Glu Ala His Gly Thr Arg Pro Asp Leu Thr
1775 1780 1785
Asp Gln Pro Leu Pro Asp Ala Asp His Thr Trp Tyr Thr Asp Gly
1790 1795 1800
Ser Ser Leu Leu Gln Glu Gly Gln Arg Lys Ala Gly Ala Ala Val
1805 1810 1815
Thr Thr Glu Thr Glu Val Ile Trp Ala Lys Ala Leu Pro Ala Gly
1820 1825 1830
Thr Ser Ala Gln Arg Ala Glu Leu Ile Ala Leu Thr Gln Ala Leu
1835 1840 1845
Lys Met Ala Glu Gly Lys Lys Leu Asn Val Tyr Thr Asp Ser Arg
1850 1855 1860
Tyr Ala Phe Ala Thr Ala His Ile His Gly Glu Ile Tyr Arg Arg
1865 1870 1875
Arg Gly Trp Leu Thr Ser Glu Gly Lys Glu Ile Lys Asn Lys Asp
1880 1885 1890
Glu Ile Leu Ala Leu Leu Lys Ala Leu Phe Leu Pro Lys Arg Leu
1895 1900 1905
Ser Ile Ile His Cys Pro Gly His Gln Lys Gly His Ser Ala Glu
1910 1915 1920
Ala Arg Gly Asn Arg Met Ala Asp Gln Ala Ala Arg Lys Ala Ala
1925 1930 1935
Ile Thr Glu Thr Pro Asp Thr Ser Thr Leu Leu Ile Glu Asn Ser
1940 1945 1950
Ser Pro Ser Gly Gly Ser Lys Arg Thr Ala Asp Gly Ser Glu Phe
1955 1960 1965
Glu Pro Lys Lys Lys Arg Lys Val Gly Ser Gly
1970 1975
<210> 83
<211> 1963
<212> PRT
<213> 人工的
<220>
<223> 合成的REDRAW编辑器
<400> 83
Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys
1 5 10 15
Arg Lys Val Thr Leu Asn Ile Glu Asp Glu Tyr Arg Leu His Glu Thr
20 25 30
Ser Lys Glu Pro Asp Val Ser Leu Gly Ser Thr Trp Leu Ser Asp Phe
35 40 45
Pro Gln Ala Trp Ala Glu Thr Gly Gly Met Gly Leu Ala Val Arg Gln
50 55 60
Ala Pro Leu Ile Ile Pro Leu Lys Ala Thr Ser Thr Pro Val Ser Ile
65 70 75 80
Lys Gln Tyr Pro Met Ser Gln Glu Ala Arg Leu Gly Ile Lys Pro His
85 90 95
Ile Gln Arg Leu Leu Asp Gln Gly Ile Leu Val Pro Cys Gln Ser Pro
100 105 110
Trp Asn Thr Pro Leu Leu Pro Val Lys Lys Pro Gly Thr Asn Asp Tyr
115 120 125
Arg Pro Val Gln Asp Leu Arg Glu Val Asn Lys Arg Val Glu Asp Ile
130 135 140
His Pro Thr Val Pro Asn Pro Tyr Asn Leu Leu Ser Gly Leu Pro Pro
145 150 155 160
Ser His Gln Trp Tyr Thr Val Leu Asp Leu Lys Asp Ala Phe Phe Cys
165 170 175
Leu Arg Leu His Pro Thr Ser Gln Pro Leu Phe Ala Phe Glu Trp Arg
180 185 190
Asp Pro Glu Met Gly Ile Ser Gly Gln Leu Thr Trp Thr Arg Leu Pro
195 200 205
Gln Gly Phe Lys Asn Ser Pro Thr Leu Phe Asn Glu Ala Leu His Arg
210 215 220
Asp Leu Ala Asp Phe Arg Ile Gln His Pro Asp Leu Ile Leu Leu Gln
225 230 235 240
Tyr Val Asp Asp Leu Leu Leu Ala Ala Thr Ser Glu Leu Asp Cys Gln
245 250 255
Gln Gly Thr Arg Ala Leu Leu Gln Thr Leu Gly Asn Leu Gly Tyr Arg
260 265 270
Ala Ser Ala Lys Lys Ala Gln Ile Cys Gln Lys Gln Val Lys Tyr Leu
275 280 285
Gly Tyr Leu Leu Lys Glu Gly Gln Arg Trp Leu Thr Glu Ala Arg Lys
290 295 300
Glu Thr Val Met Gly Gln Pro Thr Pro Lys Thr Pro Arg Gln Leu Arg
305 310 315 320
Glu Phe Leu Gly Lys Ala Gly Phe Cys Arg Leu Phe Ile Pro Gly Phe
325 330 335
Ala Glu Met Ala Ala Pro Leu Tyr Pro Leu Thr Lys Pro Gly Thr Leu
340 345 350
Phe Asn Trp Gly Pro Asp Gln Gln Lys Ala Tyr Gln Glu Ile Lys Gln
355 360 365
Ala Leu Leu Thr Ala Pro Ala Leu Gly Leu Pro Asp Leu Thr Lys Pro
370 375 380
Phe Glu Leu Phe Val Asp Glu Lys Gln Gly Tyr Ala Lys Gly Val Leu
385 390 395 400
Thr Gln Lys Leu Gly Pro Trp Arg Arg Pro Val Ala Tyr Leu Ser Lys
405 410 415
Lys Leu Asp Pro Val Ala Ala Gly Trp Pro Pro Cys Leu Arg Met Val
420 425 430
Ala Ala Ile Ala Val Leu Thr Lys Asp Ala Gly Lys Leu Thr Met Gly
435 440 445
Gln Pro Leu Val Ile Leu Ala Pro His Ala Val Glu Ala Leu Val Lys
450 455 460
Gln Pro Pro Asp Arg Trp Leu Ser Asn Ala Arg Met Thr His Tyr Gln
465 470 475 480
Ala Leu Leu Leu Asp Thr Asp Arg Val Gln Phe Gly Pro Val Val Ala
485 490 495
Leu Asn Pro Ala Thr Leu Leu Pro Leu Pro Glu Glu Gly Leu Gln His
500 505 510
Asn Cys Leu Asp Ile Leu Ala Glu Ala His Gly Thr Arg Pro Asp Leu
515 520 525
Thr Asp Gln Pro Leu Pro Asp Ala Asp His Thr Trp Tyr Thr Asp Gly
530 535 540
Ser Ser Leu Leu Gln Glu Gly Gln Arg Lys Ala Gly Ala Ala Val Thr
545 550 555 560
Thr Glu Thr Glu Val Ile Trp Ala Lys Ala Leu Pro Ala Gly Thr Ser
565 570 575
Ala Gln Arg Ala Glu Leu Ile Ala Leu Thr Gln Ala Leu Lys Met Ala
580 585 590
Glu Gly Lys Lys Leu Asn Val Tyr Thr Asp Ser Arg Tyr Ala Phe Ala
595 600 605
Thr Ala His Ile His Gly Glu Ile Tyr Arg Arg Arg Gly Trp Leu Thr
610 615 620
Ser Glu Gly Lys Glu Ile Lys Asn Lys Asp Glu Ile Leu Ala Leu Leu
625 630 635 640
Lys Ala Leu Phe Leu Pro Lys Arg Leu Ser Ile Ile His Cys Pro Gly
645 650 655
His Gln Lys Gly His Ser Ala Glu Ala Arg Gly Asn Arg Met Ala Asp
660 665 670
Gln Ala Ala Arg Lys Ala Ala Ile Thr Glu Thr Pro Asp Thr Ser Thr
675 680 685
Leu Leu Ile Glu Asn Ser Ser Pro Ser Gly Ser Glu Thr Pro Gly Thr
690 695 700
Ser Glu Ser Ala Thr Pro Glu Ser Ser Lys Leu Glu Lys Phe Thr Asn
705 710 715 720
Cys Tyr Ser Leu Ser Lys Thr Leu Arg Phe Lys Ala Ile Pro Val Gly
725 730 735
Lys Thr Gln Glu Asn Ile Asp Asn Lys Arg Leu Leu Val Glu Asp Glu
740 745 750
Lys Arg Ala Glu Asp Tyr Lys Gly Val Lys Lys Leu Leu Asp Arg Tyr
755 760 765
Tyr Leu Ser Phe Ile Asn Asp Val Leu His Ser Ile Lys Leu Lys Asn
770 775 780
Leu Asn Asn Tyr Ile Ser Leu Phe Arg Lys Lys Thr Arg Thr Glu Lys
785 790 795 800
Glu Asn Lys Glu Leu Glu Asn Leu Glu Ile Asn Leu Arg Lys Glu Ile
805 810 815
Ala Lys Ala Phe Lys Gly Asn Glu Gly Tyr Lys Ser Leu Phe Lys Lys
820 825 830
Asp Ile Ile Glu Thr Ile Leu Pro Glu Phe Leu Asp Asp Lys Asp Glu
835 840 845
Ile Ala Leu Val Asn Ser Phe Asn Gly Phe Thr Thr Ala Phe Thr Gly
850 855 860
Phe Phe Asp Asn Arg Glu Asn Met Phe Ser Glu Glu Ala Lys Ser Thr
865 870 875 880
Ser Ile Ala Phe Arg Cys Ile Asn Glu Asn Leu Thr Arg Tyr Ile Ser
885 890 895
Asn Met Asp Ile Phe Glu Lys Val Asp Ala Ile Phe Asp Lys His Glu
900 905 910
Val Gln Glu Ile Lys Glu Lys Ile Leu Asn Ser Asp Tyr Asp Val Glu
915 920 925
Asp Phe Phe Glu Gly Glu Phe Phe Asn Phe Val Leu Thr Gln Glu Gly
930 935 940
Ile Asp Val Tyr Asn Ala Ile Ile Gly Gly Phe Val Thr Glu Ser Gly
945 950 955 960
Glu Lys Ile Lys Gly Leu Asn Glu Tyr Ile Asn Leu Tyr Asn Gln Lys
965 970 975
Thr Lys Gln Lys Leu Pro Lys Phe Lys Pro Leu Tyr Lys Gln Val Leu
980 985 990
Ser Asp Arg Glu Ser Leu Ser Phe Tyr Gly Glu Gly Tyr Thr Ser Asp
995 1000 1005
Glu Glu Val Leu Glu Val Phe Arg Asn Thr Leu Asn Lys Asn Ser
1010 1015 1020
Glu Ile Phe Ser Ser Ile Lys Lys Leu Glu Lys Leu Phe Lys Asn
1025 1030 1035
Phe Asp Glu Tyr Ser Ser Ala Gly Ile Phe Val Lys Asn Gly Pro
1040 1045 1050
Ala Ile Ser Thr Ile Ser Lys Asp Ile Phe Gly Glu Trp Asn Val
1055 1060 1065
Ile Arg Asp Lys Trp Asn Ala Glu Tyr Asp Asp Ile His Leu Lys
1070 1075 1080
Lys Lys Ala Val Val Thr Glu Lys Tyr Glu Asp Asp Arg Arg Lys
1085 1090 1095
Ser Phe Lys Lys Ile Gly Ser Phe Ser Leu Glu Gln Leu Gln Glu
1100 1105 1110
Tyr Ala Asp Ala Asp Leu Ser Val Val Glu Lys Leu Lys Glu Ile
1115 1120 1125
Ile Ile Gln Lys Val Asp Glu Ile Tyr Lys Val Tyr Gly Ser Ser
1130 1135 1140
Glu Lys Leu Phe Asp Ala Asp Phe Val Leu Glu Lys Ser Leu Lys
1145 1150 1155
Lys Asn Asp Ala Val Val Ala Ile Met Lys Asp Leu Leu Asp Ser
1160 1165 1170
Val Lys Ser Phe Glu Asn Tyr Ile Lys Ala Phe Phe Gly Glu Gly
1175 1180 1185
Lys Glu Thr Asn Arg Asp Glu Ser Phe Tyr Gly Asp Phe Val Leu
1190 1195 1200
Ala Tyr Asp Ile Leu Leu Lys Val Asp His Ile Tyr Asp Ala Ile
1205 1210 1215
Arg Asn Tyr Val Thr Gln Lys Pro Tyr Ser Lys Asp Lys Phe Lys
1220 1225 1230
Leu Tyr Phe Gln Asn Pro Gln Phe Met Gly Gly Trp Asp Lys Asp
1235 1240 1245
Lys Glu Thr Asp Tyr Arg Ala Thr Ile Leu Arg Tyr Gly Ser Lys
1250 1255 1260
Tyr Tyr Leu Ala Ile Met Asp Lys Lys Tyr Ala Lys Cys Leu Gln
1265 1270 1275
Lys Ile Asp Lys Asp Asp Val Asn Gly Asn Tyr Glu Lys Ile Asn
1280 1285 1290
Tyr Lys Leu Leu Pro Gly Pro Asn Lys Met Leu Pro Lys Val Phe
1295 1300 1305
Phe Ser Lys Lys Trp Met Ala Tyr Tyr Asn Pro Ser Glu Asp Ile
1310 1315 1320
Gln Lys Ile Tyr Lys Asn Gly Thr Phe Lys Lys Gly Asp Met Phe
1325 1330 1335
Asn Leu Asn Asp Cys His Lys Leu Ile Asp Phe Phe Lys Asp Ser
1340 1345 1350
Ile Ser Arg Tyr Pro Lys Trp Ser Asn Ala Tyr Asp Phe Asn Phe
1355 1360 1365
Ser Glu Thr Glu Lys Tyr Lys Asp Ile Ala Gly Phe Tyr Arg Glu
1370 1375 1380
Val Glu Glu Gln Gly Tyr Lys Val Ser Phe Glu Ser Ala Ser Lys
1385 1390 1395
Lys Glu Val Asp Lys Leu Val Glu Glu Gly Lys Leu Tyr Met Phe
1400 1405 1410
Gln Ile Tyr Asn Lys Asp Phe Ser Asp Lys Ser His Gly Thr Pro
1415 1420 1425
Asn Leu His Thr Met Tyr Phe Lys Leu Leu Phe Asp Glu Asn Asn
1430 1435 1440
His Gly Gln Ile Arg Leu Ser Gly Gly Ala Glu Leu Phe Met Arg
1445 1450 1455
Arg Ala Ser Leu Lys Lys Glu Glu Leu Val Val Ala Pro Ala Asn
1460 1465 1470
Ser Pro Ile Ala Asn Lys Asn Pro Asp Asn Pro Lys Lys Thr Thr
1475 1480 1485
Thr Leu Ser Tyr Asp Val Tyr Lys Asp Lys Arg Phe Ser Glu Asp
1490 1495 1500
Gln Tyr Glu Leu His Ile Pro Ile Ala Ile Asn Lys Cys Pro Lys
1505 1510 1515
Asn Ile Phe Lys Ile Asn Thr Glu Val Arg Val Leu Leu Lys His
1520 1525 1530
Asp Asp Asn Pro Tyr Val Ile Gly Ile Asp Arg Gly Glu Arg Asn
1535 1540 1545
Leu Leu Tyr Ile Val Val Val Asp Gly Lys Gly Asn Ile Val Glu
1550 1555 1560
Gln Tyr Ser Leu Asn Glu Ile Ile Asn Asn Phe Asn Gly Ile Arg
1565 1570 1575
Ile Lys Thr Asp Tyr His Ser Leu Leu Asp Lys Lys Glu Lys Glu
1580 1585 1590
Arg Phe Glu Ala Arg Gln Asn Trp Thr Ser Ile Glu Asn Ile Lys
1595 1600 1605
Glu Leu Lys Ala Gly Tyr Ile Ser Gln Val Val His Lys Ile Cys
1610 1615 1620
Glu Leu Val Glu Lys Tyr Asp Ala Val Ile Ala Leu Glu Asp Leu
1625 1630 1635
Asn Ser Gly Phe Lys Asn Ser Arg Val Lys Val Glu Lys Gln Val
1640 1645 1650
Tyr Gln Lys Phe Glu Lys Met Leu Ile Asp Lys Leu Asn Tyr Met
1655 1660 1665
Val Asp Lys Lys Ser Asn Pro Cys Ala Thr Gly Gly Ala Leu Lys
1670 1675 1680
Gly Tyr Gln Ile Thr Asn Lys Phe Glu Ser Phe Lys Ser Met Ser
1685 1690 1695
Thr Gln Asn Gly Phe Ile Phe Tyr Ile Pro Ala Trp Leu Thr Ser
1700 1705 1710
Lys Ile Asp Pro Ser Thr Gly Phe Val Asn Leu Leu Lys Thr Lys
1715 1720 1725
Tyr Thr Ser Ile Ala Asp Ser Lys Lys Phe Ile Ser Ser Phe Asp
1730 1735 1740
Arg Ile Met Tyr Val Pro Glu Glu Asp Leu Phe Glu Phe Ala Leu
1745 1750 1755
Asp Tyr Lys Asn Phe Ser Arg Thr Asp Ala Asp Tyr Ile Lys Lys
1760 1765 1770
Trp Lys Leu Tyr Ser Tyr Gly Asn Arg Ile Arg Ile Phe Arg Asn
1775 1780 1785
Pro Lys Lys Asn Asn Val Phe Asp Trp Glu Glu Val Cys Leu Thr
1790 1795 1800
Ser Ala Tyr Lys Glu Leu Phe Asn Lys Tyr Gly Ile Asn Tyr Gln
1805 1810 1815
Gln Gly Asp Ile Arg Ala Leu Leu Cys Glu Gln Ser Asp Lys Ala
1820 1825 1830
Phe Tyr Ser Ser Phe Met Ala Leu Met Ser Leu Met Leu Gln Met
1835 1840 1845
Ala Asn Ser Ile Thr Gly Arg Thr Asp Val Asp Phe Leu Ile Ser
1850 1855 1860
Pro Val Lys Asn Ser Asp Gly Ile Phe Tyr Asp Ser Arg Asn Tyr
1865 1870 1875
Glu Ala Gln Glu Asn Ala Ile Leu Pro Lys Asn Ala Asp Ala Asn
1880 1885 1890
Gly Ala Tyr Asn Ile Ala Arg Lys Val Leu Trp Ala Ile Gly Gln
1895 1900 1905
Phe Lys Lys Ala Glu Asp Glu Lys Leu Asp Lys Val Lys Ile Ala
1910 1915 1920
Ile Ser Asn Lys Glu Trp Leu Glu Tyr Ala Gln Thr Ser Val Lys
1925 1930 1935
His Ser Gly Gly Ser Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu
1940 1945 1950
Pro Lys Lys Lys Arg Lys Val Gly Ser Gly
1955 1960
<210> 84
<211> 1979
<212> PRT
<213> 人工的
<220>
<223> 合成的REDRAW编辑器
<400> 84
Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys
1 5 10 15
Arg Lys Val Thr Leu Asn Ile Glu Asp Glu Tyr Arg Leu His Glu Thr
20 25 30
Ser Lys Glu Pro Asp Val Ser Leu Gly Ser Thr Trp Leu Ser Asp Phe
35 40 45
Pro Gln Ala Trp Ala Glu Thr Gly Gly Met Gly Leu Ala Val Arg Gln
50 55 60
Ala Pro Leu Ile Ile Pro Leu Lys Ala Thr Ser Thr Pro Val Ser Ile
65 70 75 80
Lys Gln Tyr Pro Met Ser Gln Glu Ala Arg Leu Gly Ile Lys Pro His
85 90 95
Ile Gln Arg Leu Leu Asp Gln Gly Ile Leu Val Pro Cys Gln Ser Pro
100 105 110
Trp Asn Thr Pro Leu Leu Pro Val Lys Lys Pro Gly Thr Asn Asp Tyr
115 120 125
Arg Pro Val Gln Asp Leu Arg Glu Val Asn Lys Arg Val Glu Asp Ile
130 135 140
His Pro Thr Val Pro Asn Pro Tyr Asn Leu Leu Ser Gly Leu Pro Pro
145 150 155 160
Ser His Gln Trp Tyr Thr Val Leu Asp Leu Lys Asp Ala Phe Phe Cys
165 170 175
Leu Arg Leu His Pro Thr Ser Gln Pro Leu Phe Ala Phe Glu Trp Arg
180 185 190
Asp Pro Glu Met Gly Ile Ser Gly Gln Leu Thr Trp Thr Arg Leu Pro
195 200 205
Gln Gly Phe Lys Asn Ser Pro Thr Leu Phe Asn Glu Ala Leu His Arg
210 215 220
Asp Leu Ala Asp Phe Arg Ile Gln His Pro Asp Leu Ile Leu Leu Gln
225 230 235 240
Tyr Val Asp Asp Leu Leu Leu Ala Ala Thr Ser Glu Leu Asp Cys Gln
245 250 255
Gln Gly Thr Arg Ala Leu Leu Gln Thr Leu Gly Asn Leu Gly Tyr Arg
260 265 270
Ala Ser Ala Lys Lys Ala Gln Ile Cys Gln Lys Gln Val Lys Tyr Leu
275 280 285
Gly Tyr Leu Leu Lys Glu Gly Gln Arg Trp Leu Thr Glu Ala Arg Lys
290 295 300
Glu Thr Val Met Gly Gln Pro Thr Pro Lys Thr Pro Arg Gln Leu Arg
305 310 315 320
Glu Phe Leu Gly Lys Ala Gly Phe Cys Arg Leu Phe Ile Pro Gly Phe
325 330 335
Ala Glu Met Ala Ala Pro Leu Tyr Pro Leu Thr Lys Pro Gly Thr Leu
340 345 350
Phe Asn Trp Gly Pro Asp Gln Gln Lys Ala Tyr Gln Glu Ile Lys Gln
355 360 365
Ala Leu Leu Thr Ala Pro Ala Leu Gly Leu Pro Asp Leu Thr Lys Pro
370 375 380
Phe Glu Leu Phe Val Asp Glu Lys Gln Gly Tyr Ala Lys Gly Val Leu
385 390 395 400
Thr Gln Lys Leu Gly Pro Trp Arg Arg Pro Val Ala Tyr Leu Ser Lys
405 410 415
Lys Leu Asp Pro Val Ala Ala Gly Trp Pro Pro Cys Leu Arg Met Val
420 425 430
Ala Ala Ile Ala Val Leu Thr Lys Asp Ala Gly Lys Leu Thr Met Gly
435 440 445
Gln Pro Leu Val Ile Leu Ala Pro His Ala Val Glu Ala Leu Val Lys
450 455 460
Gln Pro Pro Asp Arg Trp Leu Ser Asn Ala Arg Met Thr His Tyr Gln
465 470 475 480
Ala Leu Leu Leu Asp Thr Asp Arg Val Gln Phe Gly Pro Val Val Ala
485 490 495
Leu Asn Pro Ala Thr Leu Leu Pro Leu Pro Glu Glu Gly Leu Gln His
500 505 510
Asn Cys Leu Asp Ile Leu Ala Glu Ala His Gly Thr Arg Pro Asp Leu
515 520 525
Thr Asp Gln Pro Leu Pro Asp Ala Asp His Thr Trp Tyr Thr Asp Gly
530 535 540
Ser Ser Leu Leu Gln Glu Gly Gln Arg Lys Ala Gly Ala Ala Val Thr
545 550 555 560
Thr Glu Thr Glu Val Ile Trp Ala Lys Ala Leu Pro Ala Gly Thr Ser
565 570 575
Ala Gln Arg Ala Glu Leu Ile Ala Leu Thr Gln Ala Leu Lys Met Ala
580 585 590
Glu Gly Lys Lys Leu Asn Val Tyr Thr Asp Ser Arg Tyr Ala Phe Ala
595 600 605
Thr Ala His Ile His Gly Glu Ile Tyr Arg Arg Arg Gly Trp Leu Thr
610 615 620
Ser Glu Gly Lys Glu Ile Lys Asn Lys Asp Glu Ile Leu Ala Leu Leu
625 630 635 640
Lys Ala Leu Phe Leu Pro Lys Arg Leu Ser Ile Ile His Cys Pro Gly
645 650 655
His Gln Lys Gly His Ser Ala Glu Ala Arg Gly Asn Arg Met Ala Asp
660 665 670
Gln Ala Ala Arg Lys Ala Ala Ile Thr Glu Thr Pro Asp Thr Ser Thr
675 680 685
Leu Leu Ile Glu Asn Ser Ser Pro Ser Gly Gly Ser Ser Gly Gly Ser
690 695 700
Ser Gly Ser Glu Thr Pro Gly Thr Ser Glu Ser Ala Thr Pro Glu Ser
705 710 715 720
Ser Gly Gly Ser Ser Gly Gly Ser Ser Lys Leu Glu Lys Phe Thr Asn
725 730 735
Cys Tyr Ser Leu Ser Lys Thr Leu Arg Phe Lys Ala Ile Pro Val Gly
740 745 750
Lys Thr Gln Glu Asn Ile Asp Asn Lys Arg Leu Leu Val Glu Asp Glu
755 760 765
Lys Arg Ala Glu Asp Tyr Lys Gly Val Lys Lys Leu Leu Asp Arg Tyr
770 775 780
Tyr Leu Ser Phe Ile Asn Asp Val Leu His Ser Ile Lys Leu Lys Asn
785 790 795 800
Leu Asn Asn Tyr Ile Ser Leu Phe Arg Lys Lys Thr Arg Thr Glu Lys
805 810 815
Glu Asn Lys Glu Leu Glu Asn Leu Glu Ile Asn Leu Arg Lys Glu Ile
820 825 830
Ala Lys Ala Phe Lys Gly Asn Glu Gly Tyr Lys Ser Leu Phe Lys Lys
835 840 845
Asp Ile Ile Glu Thr Ile Leu Pro Glu Phe Leu Asp Asp Lys Asp Glu
850 855 860
Ile Ala Leu Val Asn Ser Phe Asn Gly Phe Thr Thr Ala Phe Thr Gly
865 870 875 880
Phe Phe Asp Asn Arg Glu Asn Met Phe Ser Glu Glu Ala Lys Ser Thr
885 890 895
Ser Ile Ala Phe Arg Cys Ile Asn Glu Asn Leu Thr Arg Tyr Ile Ser
900 905 910
Asn Met Asp Ile Phe Glu Lys Val Asp Ala Ile Phe Asp Lys His Glu
915 920 925
Val Gln Glu Ile Lys Glu Lys Ile Leu Asn Ser Asp Tyr Asp Val Glu
930 935 940
Asp Phe Phe Glu Gly Glu Phe Phe Asn Phe Val Leu Thr Gln Glu Gly
945 950 955 960
Ile Asp Val Tyr Asn Ala Ile Ile Gly Gly Phe Val Thr Glu Ser Gly
965 970 975
Glu Lys Ile Lys Gly Leu Asn Glu Tyr Ile Asn Leu Tyr Asn Gln Lys
980 985 990
Thr Lys Gln Lys Leu Pro Lys Phe Lys Pro Leu Tyr Lys Gln Val Leu
995 1000 1005
Ser Asp Arg Glu Ser Leu Ser Phe Tyr Gly Glu Gly Tyr Thr Ser
1010 1015 1020
Asp Glu Glu Val Leu Glu Val Phe Arg Asn Thr Leu Asn Lys Asn
1025 1030 1035
Ser Glu Ile Phe Ser Ser Ile Lys Lys Leu Glu Lys Leu Phe Lys
1040 1045 1050
Asn Phe Asp Glu Tyr Ser Ser Ala Gly Ile Phe Val Lys Asn Gly
1055 1060 1065
Pro Ala Ile Ser Thr Ile Ser Lys Asp Ile Phe Gly Glu Trp Asn
1070 1075 1080
Val Ile Arg Asp Lys Trp Asn Ala Glu Tyr Asp Asp Ile His Leu
1085 1090 1095
Lys Lys Lys Ala Val Val Thr Glu Lys Tyr Glu Asp Asp Arg Arg
1100 1105 1110
Lys Ser Phe Lys Lys Ile Gly Ser Phe Ser Leu Glu Gln Leu Gln
1115 1120 1125
Glu Tyr Ala Asp Ala Asp Leu Ser Val Val Glu Lys Leu Lys Glu
1130 1135 1140
Ile Ile Ile Gln Lys Val Asp Glu Ile Tyr Lys Val Tyr Gly Ser
1145 1150 1155
Ser Glu Lys Leu Phe Asp Ala Asp Phe Val Leu Glu Lys Ser Leu
1160 1165 1170
Lys Lys Asn Asp Ala Val Val Ala Ile Met Lys Asp Leu Leu Asp
1175 1180 1185
Ser Val Lys Ser Phe Glu Asn Tyr Ile Lys Ala Phe Phe Gly Glu
1190 1195 1200
Gly Lys Glu Thr Asn Arg Asp Glu Ser Phe Tyr Gly Asp Phe Val
1205 1210 1215
Leu Ala Tyr Asp Ile Leu Leu Lys Val Asp His Ile Tyr Asp Ala
1220 1225 1230
Ile Arg Asn Tyr Val Thr Gln Lys Pro Tyr Ser Lys Asp Lys Phe
1235 1240 1245
Lys Leu Tyr Phe Gln Asn Pro Gln Phe Met Gly Gly Trp Asp Lys
1250 1255 1260
Asp Lys Glu Thr Asp Tyr Arg Ala Thr Ile Leu Arg Tyr Gly Ser
1265 1270 1275
Lys Tyr Tyr Leu Ala Ile Met Asp Lys Lys Tyr Ala Lys Cys Leu
1280 1285 1290
Gln Lys Ile Asp Lys Asp Asp Val Asn Gly Asn Tyr Glu Lys Ile
1295 1300 1305
Asn Tyr Lys Leu Leu Pro Gly Pro Asn Lys Met Leu Pro Lys Val
1310 1315 1320
Phe Phe Ser Lys Lys Trp Met Ala Tyr Tyr Asn Pro Ser Glu Asp
1325 1330 1335
Ile Gln Lys Ile Tyr Lys Asn Gly Thr Phe Lys Lys Gly Asp Met
1340 1345 1350
Phe Asn Leu Asn Asp Cys His Lys Leu Ile Asp Phe Phe Lys Asp
1355 1360 1365
Ser Ile Ser Arg Tyr Pro Lys Trp Ser Asn Ala Tyr Asp Phe Asn
1370 1375 1380
Phe Ser Glu Thr Glu Lys Tyr Lys Asp Ile Ala Gly Phe Tyr Arg
1385 1390 1395
Glu Val Glu Glu Gln Gly Tyr Lys Val Ser Phe Glu Ser Ala Ser
1400 1405 1410
Lys Lys Glu Val Asp Lys Leu Val Glu Glu Gly Lys Leu Tyr Met
1415 1420 1425
Phe Gln Ile Tyr Asn Lys Asp Phe Ser Asp Lys Ser His Gly Thr
1430 1435 1440
Pro Asn Leu His Thr Met Tyr Phe Lys Leu Leu Phe Asp Glu Asn
1445 1450 1455
Asn His Gly Gln Ile Arg Leu Ser Gly Gly Ala Glu Leu Phe Met
1460 1465 1470
Arg Arg Ala Ser Leu Lys Lys Glu Glu Leu Val Val Ala Pro Ala
1475 1480 1485
Asn Ser Pro Ile Ala Asn Lys Asn Pro Asp Asn Pro Lys Lys Thr
1490 1495 1500
Thr Thr Leu Ser Tyr Asp Val Tyr Lys Asp Lys Arg Phe Ser Glu
1505 1510 1515
Asp Gln Tyr Glu Leu His Ile Pro Ile Ala Ile Asn Lys Cys Pro
1520 1525 1530
Lys Asn Ile Phe Lys Ile Asn Thr Glu Val Arg Val Leu Leu Lys
1535 1540 1545
His Asp Asp Asn Pro Tyr Val Ile Gly Ile Asp Arg Gly Glu Arg
1550 1555 1560
Asn Leu Leu Tyr Ile Val Val Val Asp Gly Lys Gly Asn Ile Val
1565 1570 1575
Glu Gln Tyr Ser Leu Asn Glu Ile Ile Asn Asn Phe Asn Gly Ile
1580 1585 1590
Arg Ile Lys Thr Asp Tyr His Ser Leu Leu Asp Lys Lys Glu Lys
1595 1600 1605
Glu Arg Phe Glu Ala Arg Gln Asn Trp Thr Ser Ile Glu Asn Ile
1610 1615 1620
Lys Glu Leu Lys Ala Gly Tyr Ile Ser Gln Val Val His Lys Ile
1625 1630 1635
Cys Glu Leu Val Glu Lys Tyr Asp Ala Val Ile Ala Leu Glu Asp
1640 1645 1650
Leu Asn Ser Gly Phe Lys Asn Ser Arg Val Lys Val Glu Lys Gln
1655 1660 1665
Val Tyr Gln Lys Phe Glu Lys Met Leu Ile Asp Lys Leu Asn Tyr
1670 1675 1680
Met Val Asp Lys Lys Ser Asn Pro Cys Ala Thr Gly Gly Ala Leu
1685 1690 1695
Lys Gly Tyr Gln Ile Thr Asn Lys Phe Glu Ser Phe Lys Ser Met
1700 1705 1710
Ser Thr Gln Asn Gly Phe Ile Phe Tyr Ile Pro Ala Trp Leu Thr
1715 1720 1725
Ser Lys Ile Asp Pro Ser Thr Gly Phe Val Asn Leu Leu Lys Thr
1730 1735 1740
Lys Tyr Thr Ser Ile Ala Asp Ser Lys Lys Phe Ile Ser Ser Phe
1745 1750 1755
Asp Arg Ile Met Tyr Val Pro Glu Glu Asp Leu Phe Glu Phe Ala
1760 1765 1770
Leu Asp Tyr Lys Asn Phe Ser Arg Thr Asp Ala Asp Tyr Ile Lys
1775 1780 1785
Lys Trp Lys Leu Tyr Ser Tyr Gly Asn Arg Ile Arg Ile Phe Arg
1790 1795 1800
Asn Pro Lys Lys Asn Asn Val Phe Asp Trp Glu Glu Val Cys Leu
1805 1810 1815
Thr Ser Ala Tyr Lys Glu Leu Phe Asn Lys Tyr Gly Ile Asn Tyr
1820 1825 1830
Gln Gln Gly Asp Ile Arg Ala Leu Leu Cys Glu Gln Ser Asp Lys
1835 1840 1845
Ala Phe Tyr Ser Ser Phe Met Ala Leu Met Ser Leu Met Leu Gln
1850 1855 1860
Met Ala Asn Ser Ile Thr Gly Arg Thr Asp Val Asp Phe Leu Ile
1865 1870 1875
Ser Pro Val Lys Asn Ser Asp Gly Ile Phe Tyr Asp Ser Arg Asn
1880 1885 1890
Tyr Glu Ala Gln Glu Asn Ala Ile Leu Pro Lys Asn Ala Asp Ala
1895 1900 1905
Asn Gly Ala Tyr Asn Ile Ala Arg Lys Val Leu Trp Ala Ile Gly
1910 1915 1920
Gln Phe Lys Lys Ala Glu Asp Glu Lys Leu Asp Lys Val Lys Ile
1925 1930 1935
Ala Ile Ser Asn Lys Glu Trp Leu Glu Tyr Ala Gln Thr Ser Val
1940 1945 1950
Lys His Ser Gly Gly Ser Lys Arg Thr Ala Asp Gly Ser Glu Phe
1955 1960 1965
Glu Pro Lys Lys Lys Arg Lys Val Gly Ser Gly
1970 1975
<210> 85
<211> 1963
<212> PRT
<213> 人工的
<220>
<223> 合成的REDRAW编辑器
<400> 85
Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys
1 5 10 15
Arg Lys Val Ser Lys Leu Glu Lys Phe Thr Asn Cys Tyr Ser Leu Ser
20 25 30
Lys Thr Leu Arg Phe Lys Ala Ile Pro Val Gly Lys Thr Gln Glu Asn
35 40 45
Ile Asp Asn Lys Arg Leu Leu Val Glu Asp Glu Lys Arg Ala Glu Asp
50 55 60
Tyr Lys Gly Val Lys Lys Leu Leu Asp Arg Tyr Tyr Leu Ser Phe Ile
65 70 75 80
Asn Asp Val Leu His Ser Ile Lys Leu Lys Asn Leu Asn Asn Tyr Ile
85 90 95
Ser Leu Phe Arg Lys Lys Thr Arg Thr Glu Lys Glu Asn Lys Glu Leu
100 105 110
Glu Asn Leu Glu Ile Asn Leu Arg Lys Glu Ile Ala Lys Ala Phe Lys
115 120 125
Gly Asn Glu Gly Tyr Lys Ser Leu Phe Lys Lys Asp Ile Ile Glu Thr
130 135 140
Ile Leu Pro Glu Phe Leu Asp Asp Lys Asp Glu Ile Ala Leu Val Asn
145 150 155 160
Ser Phe Asn Gly Phe Thr Thr Ala Phe Thr Gly Phe Phe Asp Asn Arg
165 170 175
Glu Asn Met Phe Ser Glu Glu Ala Lys Ser Thr Ser Ile Ala Phe Arg
180 185 190
Cys Ile Asn Glu Asn Leu Thr Arg Tyr Ile Ser Asn Met Asp Ile Phe
195 200 205
Glu Lys Val Asp Ala Ile Phe Asp Lys His Glu Val Gln Glu Ile Lys
210 215 220
Glu Lys Ile Leu Asn Ser Asp Tyr Asp Val Glu Asp Phe Phe Glu Gly
225 230 235 240
Glu Phe Phe Asn Phe Val Leu Thr Gln Glu Gly Ile Asp Val Tyr Asn
245 250 255
Ala Ile Ile Gly Gly Phe Val Thr Glu Ser Gly Glu Lys Ile Lys Gly
260 265 270
Leu Asn Glu Tyr Ile Asn Leu Tyr Asn Gln Lys Thr Lys Gln Lys Leu
275 280 285
Pro Lys Phe Lys Pro Leu Tyr Lys Gln Val Leu Ser Asp Arg Glu Ser
290 295 300
Leu Ser Phe Tyr Gly Glu Gly Tyr Thr Ser Asp Glu Glu Val Leu Glu
305 310 315 320
Val Phe Arg Asn Thr Leu Asn Lys Asn Ser Glu Ile Phe Ser Ser Ile
325 330 335
Lys Lys Leu Glu Lys Leu Phe Lys Asn Phe Asp Glu Tyr Ser Ser Ala
340 345 350
Gly Ile Phe Val Lys Asn Gly Pro Ala Ile Ser Thr Ile Ser Lys Asp
355 360 365
Ile Phe Gly Glu Trp Asn Val Ile Arg Asp Lys Trp Asn Ala Glu Tyr
370 375 380
Asp Asp Ile His Leu Lys Lys Lys Ala Val Val Thr Glu Lys Tyr Glu
385 390 395 400
Asp Asp Arg Arg Lys Ser Phe Lys Lys Ile Gly Ser Phe Ser Leu Glu
405 410 415
Gln Leu Gln Glu Tyr Ala Asp Ala Asp Leu Ser Val Val Glu Lys Leu
420 425 430
Lys Glu Ile Ile Ile Gln Lys Val Asp Glu Ile Tyr Lys Val Tyr Gly
435 440 445
Ser Ser Glu Lys Leu Phe Asp Ala Asp Phe Val Leu Glu Lys Ser Leu
450 455 460
Lys Lys Asn Asp Ala Val Val Ala Ile Met Lys Asp Leu Leu Asp Ser
465 470 475 480
Val Lys Ser Phe Glu Asn Tyr Ile Lys Ala Phe Phe Gly Glu Gly Lys
485 490 495
Glu Thr Asn Arg Asp Glu Ser Phe Tyr Gly Asp Phe Val Leu Ala Tyr
500 505 510
Asp Ile Leu Leu Lys Val Asp His Ile Tyr Asp Ala Ile Arg Asn Tyr
515 520 525
Val Thr Gln Lys Pro Tyr Ser Lys Asp Lys Phe Lys Leu Tyr Phe Gln
530 535 540
Asn Pro Gln Phe Met Gly Gly Trp Asp Lys Asp Lys Glu Thr Asp Tyr
545 550 555 560
Arg Ala Thr Ile Leu Arg Tyr Gly Ser Lys Tyr Tyr Leu Ala Ile Met
565 570 575
Asp Lys Lys Tyr Ala Lys Cys Leu Gln Lys Ile Asp Lys Asp Asp Val
580 585 590
Asn Gly Asn Tyr Glu Lys Ile Asn Tyr Lys Leu Leu Pro Gly Pro Asn
595 600 605
Lys Met Leu Pro Lys Val Phe Phe Ser Lys Lys Trp Met Ala Tyr Tyr
610 615 620
Asn Pro Ser Glu Asp Ile Gln Lys Ile Tyr Lys Asn Gly Thr Phe Lys
625 630 635 640
Lys Gly Asp Met Phe Asn Leu Asn Asp Cys His Lys Leu Ile Asp Phe
645 650 655
Phe Lys Asp Ser Ile Ser Arg Tyr Pro Lys Trp Ser Asn Ala Tyr Asp
660 665 670
Phe Asn Phe Ser Glu Thr Glu Lys Tyr Lys Asp Ile Ala Gly Phe Tyr
675 680 685
Arg Glu Val Glu Glu Gln Gly Tyr Lys Val Ser Phe Glu Ser Ala Ser
690 695 700
Lys Lys Glu Val Asp Lys Leu Val Glu Glu Gly Lys Leu Tyr Met Phe
705 710 715 720
Gln Ile Tyr Asn Lys Asp Phe Ser Asp Lys Ser His Gly Thr Pro Asn
725 730 735
Leu His Thr Met Tyr Phe Lys Leu Leu Phe Asp Glu Asn Asn His Gly
740 745 750
Gln Ile Arg Leu Ser Gly Gly Ala Glu Leu Phe Met Arg Arg Ala Ser
755 760 765
Leu Lys Lys Glu Glu Leu Val Val Ala Pro Ala Asn Ser Pro Ile Ala
770 775 780
Asn Lys Asn Pro Asp Asn Pro Lys Lys Thr Thr Thr Leu Ser Tyr Asp
785 790 795 800
Val Tyr Lys Asp Lys Arg Phe Ser Glu Asp Gln Tyr Glu Leu His Ile
805 810 815
Pro Ile Ala Ile Asn Lys Cys Pro Lys Asn Ile Phe Lys Ile Asn Thr
820 825 830
Glu Val Arg Val Leu Leu Lys His Asp Asp Asn Pro Tyr Val Ile Gly
835 840 845
Ile Asp Arg Gly Glu Arg Asn Leu Leu Tyr Ile Val Val Val Asp Gly
850 855 860
Lys Gly Asn Ile Val Glu Gln Tyr Ser Leu Asn Glu Ile Ile Asn Asn
865 870 875 880
Phe Asn Gly Ile Arg Ile Lys Thr Asp Tyr His Ser Leu Leu Asp Lys
885 890 895
Lys Glu Lys Glu Arg Phe Glu Ala Arg Gln Asn Trp Thr Ser Ile Glu
900 905 910
Asn Ile Lys Glu Leu Lys Ala Gly Tyr Ile Ser Gln Val Val His Lys
915 920 925
Ile Cys Glu Leu Val Glu Lys Tyr Asp Ala Val Ile Ala Leu Glu Asp
930 935 940
Leu Asn Ser Gly Phe Lys Asn Ser Arg Val Lys Val Glu Lys Gln Val
945 950 955 960
Tyr Gln Lys Phe Glu Lys Met Leu Ile Asp Lys Leu Asn Tyr Met Val
965 970 975
Asp Lys Lys Ser Asn Pro Cys Ala Thr Gly Gly Ala Leu Lys Gly Tyr
980 985 990
Gln Ile Thr Asn Lys Phe Glu Ser Phe Lys Ser Met Ser Thr Gln Asn
995 1000 1005
Gly Phe Ile Phe Tyr Ile Pro Ala Trp Leu Thr Ser Lys Ile Asp
1010 1015 1020
Pro Ser Thr Gly Phe Val Asn Leu Leu Lys Thr Lys Tyr Thr Ser
1025 1030 1035
Ile Ala Asp Ser Lys Lys Phe Ile Ser Ser Phe Asp Arg Ile Met
1040 1045 1050
Tyr Val Pro Glu Glu Asp Leu Phe Glu Phe Ala Leu Asp Tyr Lys
1055 1060 1065
Asn Phe Ser Arg Thr Asp Ala Asp Tyr Ile Lys Lys Trp Lys Leu
1070 1075 1080
Tyr Ser Tyr Gly Asn Arg Ile Arg Ile Phe Arg Asn Pro Lys Lys
1085 1090 1095
Asn Asn Val Phe Asp Trp Glu Glu Val Cys Leu Thr Ser Ala Tyr
1100 1105 1110
Lys Glu Leu Phe Asn Lys Tyr Gly Ile Asn Tyr Gln Gln Gly Asp
1115 1120 1125
Ile Arg Ala Leu Leu Cys Glu Gln Ser Asp Lys Ala Phe Tyr Ser
1130 1135 1140
Ser Phe Met Ala Leu Met Ser Leu Met Leu Gln Met Ala Asn Ser
1145 1150 1155
Ile Thr Gly Arg Thr Asp Val Asp Phe Leu Ile Ser Pro Val Lys
1160 1165 1170
Asn Ser Asp Gly Ile Phe Tyr Asp Ser Arg Asn Tyr Glu Ala Gln
1175 1180 1185
Glu Asn Ala Ile Leu Pro Lys Asn Ala Asp Ala Asn Gly Ala Tyr
1190 1195 1200
Asn Ile Ala Arg Lys Val Leu Trp Ala Ile Gly Gln Phe Lys Lys
1205 1210 1215
Ala Glu Asp Glu Lys Leu Asp Lys Val Lys Ile Ala Ile Ser Asn
1220 1225 1230
Lys Glu Trp Leu Glu Tyr Ala Gln Thr Ser Val Lys His Ser Gly
1235 1240 1245
Ser Glu Thr Pro Gly Thr Ser Glu Ser Ala Thr Pro Glu Ser Thr
1250 1255 1260
Leu Asn Ile Glu Asp Glu Tyr Arg Leu His Glu Thr Ser Lys Glu
1265 1270 1275
Pro Asp Val Ser Leu Gly Ser Thr Trp Leu Ser Asp Phe Pro Gln
1280 1285 1290
Ala Trp Ala Glu Thr Gly Gly Met Gly Leu Ala Val Arg Gln Ala
1295 1300 1305
Pro Leu Ile Ile Pro Leu Lys Ala Thr Ser Thr Pro Val Ser Ile
1310 1315 1320
Lys Gln Tyr Pro Met Ser Gln Glu Ala Arg Leu Gly Ile Lys Pro
1325 1330 1335
His Ile Gln Arg Leu Leu Asp Gln Gly Ile Leu Val Pro Cys Gln
1340 1345 1350
Ser Pro Trp Asn Thr Pro Leu Leu Pro Val Lys Lys Pro Gly Thr
1355 1360 1365
Asn Asp Tyr Arg Pro Val Gln Asp Leu Arg Glu Val Asn Lys Arg
1370 1375 1380
Val Glu Asp Ile His Pro Thr Val Pro Asn Pro Tyr Asn Leu Leu
1385 1390 1395
Ser Gly Leu Pro Pro Ser His Gln Trp Tyr Thr Val Leu Asp Leu
1400 1405 1410
Lys Asp Ala Phe Phe Cys Leu Arg Leu His Pro Thr Ser Gln Pro
1415 1420 1425
Leu Phe Ala Phe Glu Trp Arg Asp Pro Glu Met Gly Ile Ser Gly
1430 1435 1440
Gln Leu Thr Trp Thr Arg Leu Pro Gln Gly Phe Lys Asn Ser Pro
1445 1450 1455
Thr Leu Phe Asn Glu Ala Leu His Arg Asp Leu Ala Asp Phe Arg
1460 1465 1470
Ile Gln His Pro Asp Leu Ile Leu Leu Gln Tyr Val Asp Asp Leu
1475 1480 1485
Leu Leu Ala Ala Thr Ser Glu Leu Asp Cys Gln Gln Gly Thr Arg
1490 1495 1500
Ala Leu Leu Gln Thr Leu Gly Asn Leu Gly Tyr Arg Ala Ser Ala
1505 1510 1515
Lys Lys Ala Gln Ile Cys Gln Lys Gln Val Lys Tyr Leu Gly Tyr
1520 1525 1530
Leu Leu Lys Glu Gly Gln Arg Trp Leu Thr Glu Ala Arg Lys Glu
1535 1540 1545
Thr Val Met Gly Gln Pro Thr Pro Lys Thr Pro Arg Gln Leu Arg
1550 1555 1560
Glu Phe Leu Gly Lys Ala Gly Phe Cys Arg Leu Phe Ile Pro Gly
1565 1570 1575
Phe Ala Glu Met Ala Ala Pro Leu Tyr Pro Leu Thr Lys Pro Gly
1580 1585 1590
Thr Leu Phe Asn Trp Gly Pro Asp Gln Gln Lys Ala Tyr Gln Glu
1595 1600 1605
Ile Lys Gln Ala Leu Leu Thr Ala Pro Ala Leu Gly Leu Pro Asp
1610 1615 1620
Leu Thr Lys Pro Phe Glu Leu Phe Val Asp Glu Lys Gln Gly Tyr
1625 1630 1635
Ala Lys Gly Val Leu Thr Gln Lys Leu Gly Pro Trp Arg Arg Pro
1640 1645 1650
Val Ala Tyr Leu Ser Lys Lys Leu Asp Pro Val Ala Ala Gly Trp
1655 1660 1665
Pro Pro Cys Leu Arg Met Val Ala Ala Ile Ala Val Leu Thr Lys
1670 1675 1680
Asp Ala Gly Lys Leu Thr Met Gly Gln Pro Leu Val Ile Leu Ala
1685 1690 1695
Pro His Ala Val Glu Ala Leu Val Lys Gln Pro Pro Asp Arg Trp
1700 1705 1710
Leu Ser Asn Ala Arg Met Thr His Tyr Gln Ala Leu Leu Leu Asp
1715 1720 1725
Thr Asp Arg Val Gln Phe Gly Pro Val Val Ala Leu Asn Pro Ala
1730 1735 1740
Thr Leu Leu Pro Leu Pro Glu Glu Gly Leu Gln His Asn Cys Leu
1745 1750 1755
Asp Ile Leu Ala Glu Ala His Gly Thr Arg Pro Asp Leu Thr Asp
1760 1765 1770
Gln Pro Leu Pro Asp Ala Asp His Thr Trp Tyr Thr Asp Gly Ser
1775 1780 1785
Ser Leu Leu Gln Glu Gly Gln Arg Lys Ala Gly Ala Ala Val Thr
1790 1795 1800
Thr Glu Thr Glu Val Ile Trp Ala Lys Ala Leu Pro Ala Gly Thr
1805 1810 1815
Ser Ala Gln Arg Ala Glu Leu Ile Ala Leu Thr Gln Ala Leu Lys
1820 1825 1830
Met Ala Glu Gly Lys Lys Leu Asn Val Tyr Thr Asp Ser Arg Tyr
1835 1840 1845
Ala Phe Ala Thr Ala His Ile His Gly Glu Ile Tyr Arg Arg Arg
1850 1855 1860
Gly Trp Leu Thr Ser Glu Gly Lys Glu Ile Lys Asn Lys Asp Glu
1865 1870 1875
Ile Leu Ala Leu Leu Lys Ala Leu Phe Leu Pro Lys Arg Leu Ser
1880 1885 1890
Ile Ile His Cys Pro Gly His Gln Lys Gly His Ser Ala Glu Ala
1895 1900 1905
Arg Gly Asn Arg Met Ala Asp Gln Ala Ala Arg Lys Ala Ala Ile
1910 1915 1920
Thr Glu Thr Pro Asp Thr Ser Thr Leu Leu Ile Glu Asn Ser Ser
1925 1930 1935
Pro Ser Gly Gly Ser Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu
1940 1945 1950
Pro Lys Lys Lys Arg Lys Val Gly Ser Gly
1955 1960
<210> 86
<211> 1979
<212> PRT
<213> 人工的
<220>
<223> 合成的REDRAW编辑器
<400> 86
Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys
1 5 10 15
Arg Lys Val Ser Lys Leu Glu Lys Phe Thr Asn Cys Tyr Ser Leu Ser
20 25 30
Lys Thr Leu Arg Phe Lys Ala Ile Pro Val Gly Lys Thr Gln Glu Asn
35 40 45
Ile Asp Asn Lys Arg Leu Leu Val Glu Asp Glu Lys Arg Ala Glu Asp
50 55 60
Tyr Lys Gly Val Lys Lys Leu Leu Asp Arg Tyr Tyr Leu Ser Phe Ile
65 70 75 80
Asn Asp Val Leu His Ser Ile Lys Leu Lys Asn Leu Asn Asn Tyr Ile
85 90 95
Ser Leu Phe Arg Lys Lys Thr Arg Thr Glu Lys Glu Asn Lys Glu Leu
100 105 110
Glu Asn Leu Glu Ile Asn Leu Arg Lys Glu Ile Ala Lys Ala Phe Lys
115 120 125
Gly Asn Glu Gly Tyr Lys Ser Leu Phe Lys Lys Asp Ile Ile Glu Thr
130 135 140
Ile Leu Pro Glu Phe Leu Asp Asp Lys Asp Glu Ile Ala Leu Val Asn
145 150 155 160
Ser Phe Asn Gly Phe Thr Thr Ala Phe Thr Gly Phe Phe Asp Asn Arg
165 170 175
Glu Asn Met Phe Ser Glu Glu Ala Lys Ser Thr Ser Ile Ala Phe Arg
180 185 190
Cys Ile Asn Glu Asn Leu Thr Arg Tyr Ile Ser Asn Met Asp Ile Phe
195 200 205
Glu Lys Val Asp Ala Ile Phe Asp Lys His Glu Val Gln Glu Ile Lys
210 215 220
Glu Lys Ile Leu Asn Ser Asp Tyr Asp Val Glu Asp Phe Phe Glu Gly
225 230 235 240
Glu Phe Phe Asn Phe Val Leu Thr Gln Glu Gly Ile Asp Val Tyr Asn
245 250 255
Ala Ile Ile Gly Gly Phe Val Thr Glu Ser Gly Glu Lys Ile Lys Gly
260 265 270
Leu Asn Glu Tyr Ile Asn Leu Tyr Asn Gln Lys Thr Lys Gln Lys Leu
275 280 285
Pro Lys Phe Lys Pro Leu Tyr Lys Gln Val Leu Ser Asp Arg Glu Ser
290 295 300
Leu Ser Phe Tyr Gly Glu Gly Tyr Thr Ser Asp Glu Glu Val Leu Glu
305 310 315 320
Val Phe Arg Asn Thr Leu Asn Lys Asn Ser Glu Ile Phe Ser Ser Ile
325 330 335
Lys Lys Leu Glu Lys Leu Phe Lys Asn Phe Asp Glu Tyr Ser Ser Ala
340 345 350
Gly Ile Phe Val Lys Asn Gly Pro Ala Ile Ser Thr Ile Ser Lys Asp
355 360 365
Ile Phe Gly Glu Trp Asn Val Ile Arg Asp Lys Trp Asn Ala Glu Tyr
370 375 380
Asp Asp Ile His Leu Lys Lys Lys Ala Val Val Thr Glu Lys Tyr Glu
385 390 395 400
Asp Asp Arg Arg Lys Ser Phe Lys Lys Ile Gly Ser Phe Ser Leu Glu
405 410 415
Gln Leu Gln Glu Tyr Ala Asp Ala Asp Leu Ser Val Val Glu Lys Leu
420 425 430
Lys Glu Ile Ile Ile Gln Lys Val Asp Glu Ile Tyr Lys Val Tyr Gly
435 440 445
Ser Ser Glu Lys Leu Phe Asp Ala Asp Phe Val Leu Glu Lys Ser Leu
450 455 460
Lys Lys Asn Asp Ala Val Val Ala Ile Met Lys Asp Leu Leu Asp Ser
465 470 475 480
Val Lys Ser Phe Glu Asn Tyr Ile Lys Ala Phe Phe Gly Glu Gly Lys
485 490 495
Glu Thr Asn Arg Asp Glu Ser Phe Tyr Gly Asp Phe Val Leu Ala Tyr
500 505 510
Asp Ile Leu Leu Lys Val Asp His Ile Tyr Asp Ala Ile Arg Asn Tyr
515 520 525
Val Thr Gln Lys Pro Tyr Ser Lys Asp Lys Phe Lys Leu Tyr Phe Gln
530 535 540
Asn Pro Gln Phe Met Gly Gly Trp Asp Lys Asp Lys Glu Thr Asp Tyr
545 550 555 560
Arg Ala Thr Ile Leu Arg Tyr Gly Ser Lys Tyr Tyr Leu Ala Ile Met
565 570 575
Asp Lys Lys Tyr Ala Lys Cys Leu Gln Lys Ile Asp Lys Asp Asp Val
580 585 590
Asn Gly Asn Tyr Glu Lys Ile Asn Tyr Lys Leu Leu Pro Gly Pro Asn
595 600 605
Lys Met Leu Pro Lys Val Phe Phe Ser Lys Lys Trp Met Ala Tyr Tyr
610 615 620
Asn Pro Ser Glu Asp Ile Gln Lys Ile Tyr Lys Asn Gly Thr Phe Lys
625 630 635 640
Lys Gly Asp Met Phe Asn Leu Asn Asp Cys His Lys Leu Ile Asp Phe
645 650 655
Phe Lys Asp Ser Ile Ser Arg Tyr Pro Lys Trp Ser Asn Ala Tyr Asp
660 665 670
Phe Asn Phe Ser Glu Thr Glu Lys Tyr Lys Asp Ile Ala Gly Phe Tyr
675 680 685
Arg Glu Val Glu Glu Gln Gly Tyr Lys Val Ser Phe Glu Ser Ala Ser
690 695 700
Lys Lys Glu Val Asp Lys Leu Val Glu Glu Gly Lys Leu Tyr Met Phe
705 710 715 720
Gln Ile Tyr Asn Lys Asp Phe Ser Asp Lys Ser His Gly Thr Pro Asn
725 730 735
Leu His Thr Met Tyr Phe Lys Leu Leu Phe Asp Glu Asn Asn His Gly
740 745 750
Gln Ile Arg Leu Ser Gly Gly Ala Glu Leu Phe Met Arg Arg Ala Ser
755 760 765
Leu Lys Lys Glu Glu Leu Val Val Ala Pro Ala Asn Ser Pro Ile Ala
770 775 780
Asn Lys Asn Pro Asp Asn Pro Lys Lys Thr Thr Thr Leu Ser Tyr Asp
785 790 795 800
Val Tyr Lys Asp Lys Arg Phe Ser Glu Asp Gln Tyr Glu Leu His Ile
805 810 815
Pro Ile Ala Ile Asn Lys Cys Pro Lys Asn Ile Phe Lys Ile Asn Thr
820 825 830
Glu Val Arg Val Leu Leu Lys His Asp Asp Asn Pro Tyr Val Ile Gly
835 840 845
Ile Asp Arg Gly Glu Arg Asn Leu Leu Tyr Ile Val Val Val Asp Gly
850 855 860
Lys Gly Asn Ile Val Glu Gln Tyr Ser Leu Asn Glu Ile Ile Asn Asn
865 870 875 880
Phe Asn Gly Ile Arg Ile Lys Thr Asp Tyr His Ser Leu Leu Asp Lys
885 890 895
Lys Glu Lys Glu Arg Phe Glu Ala Arg Gln Asn Trp Thr Ser Ile Glu
900 905 910
Asn Ile Lys Glu Leu Lys Ala Gly Tyr Ile Ser Gln Val Val His Lys
915 920 925
Ile Cys Glu Leu Val Glu Lys Tyr Asp Ala Val Ile Ala Leu Glu Asp
930 935 940
Leu Asn Ser Gly Phe Lys Asn Ser Arg Val Lys Val Glu Lys Gln Val
945 950 955 960
Tyr Gln Lys Phe Glu Lys Met Leu Ile Asp Lys Leu Asn Tyr Met Val
965 970 975
Asp Lys Lys Ser Asn Pro Cys Ala Thr Gly Gly Ala Leu Lys Gly Tyr
980 985 990
Gln Ile Thr Asn Lys Phe Glu Ser Phe Lys Ser Met Ser Thr Gln Asn
995 1000 1005
Gly Phe Ile Phe Tyr Ile Pro Ala Trp Leu Thr Ser Lys Ile Asp
1010 1015 1020
Pro Ser Thr Gly Phe Val Asn Leu Leu Lys Thr Lys Tyr Thr Ser
1025 1030 1035
Ile Ala Asp Ser Lys Lys Phe Ile Ser Ser Phe Asp Arg Ile Met
1040 1045 1050
Tyr Val Pro Glu Glu Asp Leu Phe Glu Phe Ala Leu Asp Tyr Lys
1055 1060 1065
Asn Phe Ser Arg Thr Asp Ala Asp Tyr Ile Lys Lys Trp Lys Leu
1070 1075 1080
Tyr Ser Tyr Gly Asn Arg Ile Arg Ile Phe Arg Asn Pro Lys Lys
1085 1090 1095
Asn Asn Val Phe Asp Trp Glu Glu Val Cys Leu Thr Ser Ala Tyr
1100 1105 1110
Lys Glu Leu Phe Asn Lys Tyr Gly Ile Asn Tyr Gln Gln Gly Asp
1115 1120 1125
Ile Arg Ala Leu Leu Cys Glu Gln Ser Asp Lys Ala Phe Tyr Ser
1130 1135 1140
Ser Phe Met Ala Leu Met Ser Leu Met Leu Gln Met Ala Asn Ser
1145 1150 1155
Ile Thr Gly Arg Thr Asp Val Asp Phe Leu Ile Ser Pro Val Lys
1160 1165 1170
Asn Ser Asp Gly Ile Phe Tyr Asp Ser Arg Asn Tyr Glu Ala Gln
1175 1180 1185
Glu Asn Ala Ile Leu Pro Lys Asn Ala Asp Ala Asn Gly Ala Tyr
1190 1195 1200
Asn Ile Ala Arg Lys Val Leu Trp Ala Ile Gly Gln Phe Lys Lys
1205 1210 1215
Ala Glu Asp Glu Lys Leu Asp Lys Val Lys Ile Ala Ile Ser Asn
1220 1225 1230
Lys Glu Trp Leu Glu Tyr Ala Gln Thr Ser Val Lys His Ser Gly
1235 1240 1245
Gly Ser Ser Gly Gly Ser Ser Gly Ser Glu Thr Pro Gly Thr Ser
1250 1255 1260
Glu Ser Ala Thr Pro Glu Ser Ser Gly Gly Ser Ser Gly Gly Ser
1265 1270 1275
Thr Leu Asn Ile Glu Asp Glu Tyr Arg Leu His Glu Thr Ser Lys
1280 1285 1290
Glu Pro Asp Val Ser Leu Gly Ser Thr Trp Leu Ser Asp Phe Pro
1295 1300 1305
Gln Ala Trp Ala Glu Thr Gly Gly Met Gly Leu Ala Val Arg Gln
1310 1315 1320
Ala Pro Leu Ile Ile Pro Leu Lys Ala Thr Ser Thr Pro Val Ser
1325 1330 1335
Ile Lys Gln Tyr Pro Met Ser Gln Glu Ala Arg Leu Gly Ile Lys
1340 1345 1350
Pro His Ile Gln Arg Leu Leu Asp Gln Gly Ile Leu Val Pro Cys
1355 1360 1365
Gln Ser Pro Trp Asn Thr Pro Leu Leu Pro Val Lys Lys Pro Gly
1370 1375 1380
Thr Asn Asp Tyr Arg Pro Val Gln Asp Leu Arg Glu Val Asn Lys
1385 1390 1395
Arg Val Glu Asp Ile His Pro Thr Val Pro Asn Pro Tyr Asn Leu
1400 1405 1410
Leu Ser Gly Leu Pro Pro Ser His Gln Trp Tyr Thr Val Leu Asp
1415 1420 1425
Leu Lys Asp Ala Phe Phe Cys Leu Arg Leu His Pro Thr Ser Gln
1430 1435 1440
Pro Leu Phe Ala Phe Glu Trp Arg Asp Pro Glu Met Gly Ile Ser
1445 1450 1455
Gly Gln Leu Thr Trp Thr Arg Leu Pro Gln Gly Phe Lys Asn Ser
1460 1465 1470
Pro Thr Leu Phe Asn Glu Ala Leu His Arg Asp Leu Ala Asp Phe
1475 1480 1485
Arg Ile Gln His Pro Asp Leu Ile Leu Leu Gln Tyr Val Asp Asp
1490 1495 1500
Leu Leu Leu Ala Ala Thr Ser Glu Leu Asp Cys Gln Gln Gly Thr
1505 1510 1515
Arg Ala Leu Leu Gln Thr Leu Gly Asn Leu Gly Tyr Arg Ala Ser
1520 1525 1530
Ala Lys Lys Ala Gln Ile Cys Gln Lys Gln Val Lys Tyr Leu Gly
1535 1540 1545
Tyr Leu Leu Lys Glu Gly Gln Arg Trp Leu Thr Glu Ala Arg Lys
1550 1555 1560
Glu Thr Val Met Gly Gln Pro Thr Pro Lys Thr Pro Arg Gln Leu
1565 1570 1575
Arg Glu Phe Leu Gly Lys Ala Gly Phe Cys Arg Leu Phe Ile Pro
1580 1585 1590
Gly Phe Ala Glu Met Ala Ala Pro Leu Tyr Pro Leu Thr Lys Pro
1595 1600 1605
Gly Thr Leu Phe Asn Trp Gly Pro Asp Gln Gln Lys Ala Tyr Gln
1610 1615 1620
Glu Ile Lys Gln Ala Leu Leu Thr Ala Pro Ala Leu Gly Leu Pro
1625 1630 1635
Asp Leu Thr Lys Pro Phe Glu Leu Phe Val Asp Glu Lys Gln Gly
1640 1645 1650
Tyr Ala Lys Gly Val Leu Thr Gln Lys Leu Gly Pro Trp Arg Arg
1655 1660 1665
Pro Val Ala Tyr Leu Ser Lys Lys Leu Asp Pro Val Ala Ala Gly
1670 1675 1680
Trp Pro Pro Cys Leu Arg Met Val Ala Ala Ile Ala Val Leu Thr
1685 1690 1695
Lys Asp Ala Gly Lys Leu Thr Met Gly Gln Pro Leu Val Ile Leu
1700 1705 1710
Ala Pro His Ala Val Glu Ala Leu Val Lys Gln Pro Pro Asp Arg
1715 1720 1725
Trp Leu Ser Asn Ala Arg Met Thr His Tyr Gln Ala Leu Leu Leu
1730 1735 1740
Asp Thr Asp Arg Val Gln Phe Gly Pro Val Val Ala Leu Asn Pro
1745 1750 1755
Ala Thr Leu Leu Pro Leu Pro Glu Glu Gly Leu Gln His Asn Cys
1760 1765 1770
Leu Asp Ile Leu Ala Glu Ala His Gly Thr Arg Pro Asp Leu Thr
1775 1780 1785
Asp Gln Pro Leu Pro Asp Ala Asp His Thr Trp Tyr Thr Asp Gly
1790 1795 1800
Ser Ser Leu Leu Gln Glu Gly Gln Arg Lys Ala Gly Ala Ala Val
1805 1810 1815
Thr Thr Glu Thr Glu Val Ile Trp Ala Lys Ala Leu Pro Ala Gly
1820 1825 1830
Thr Ser Ala Gln Arg Ala Glu Leu Ile Ala Leu Thr Gln Ala Leu
1835 1840 1845
Lys Met Ala Glu Gly Lys Lys Leu Asn Val Tyr Thr Asp Ser Arg
1850 1855 1860
Tyr Ala Phe Ala Thr Ala His Ile His Gly Glu Ile Tyr Arg Arg
1865 1870 1875
Arg Gly Trp Leu Thr Ser Glu Gly Lys Glu Ile Lys Asn Lys Asp
1880 1885 1890
Glu Ile Leu Ala Leu Leu Lys Ala Leu Phe Leu Pro Lys Arg Leu
1895 1900 1905
Ser Ile Ile His Cys Pro Gly His Gln Lys Gly His Ser Ala Glu
1910 1915 1920
Ala Arg Gly Asn Arg Met Ala Asp Gln Ala Ala Arg Lys Ala Ala
1925 1930 1935
Ile Thr Glu Thr Pro Asp Thr Ser Thr Leu Leu Ile Glu Asn Ser
1940 1945 1950
Ser Pro Ser Gly Gly Ser Lys Arg Thr Ala Asp Gly Ser Glu Phe
1955 1960 1965
Glu Pro Lys Lys Lys Arg Lys Val Gly Ser Gly
1970 1975
<210> 87
<211> 166
<212> DNA
<213> 人工的
<220>
<223> 合成的tagRNA
<400> 87
gtttcaaaga ttaaataatt tctactaagt gtagattacg gctccgcagt ggatggcggt 60
aatttctact aagtgtagat gcggcgcgtt gtttcatcaa ggcgtacggt caccgtaacc 120
agcaaatcaa tatcactgtg tggcttcagg ccgccatcca ctgcgg 166
<210> 88
<211> 8019
<212> DNA
<213> 人工的
<220>
<223> 合成的质粒
<400> 88
gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc 60
catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca 120
acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga 180
ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc 240
aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct 300
ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat 360
tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc 420
ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt 480
ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa 540
tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctggttta gtgaaccgtc 600
agatccgcta gagatccgcg gccgctaata cgactcacta tagggagagc cgccaccatg 660
aaacggacag ccgacggaag cgagttcgag tcaccaaaga agaagcggaa agtctccaag 720
ctggagaagt ttacaaactg ttacagcctc tccaaaaccc tcaggtttaa agcgatcccg 780
gtgggcaaga cccaggagaa catcgacaac aagaggctcc tggtggaaga cgagaagcgc 840
gccgaagact acaagggcgt gaagaagctg ctcgataggt actacctcag ctttattaac 900
gacgtgctgc acagcatcaa actcaagaat ctcaacaact acatctccct cttccgcaaa 960
aagacccgca ccgagaagga gaacaaggag ctggagaacc tggagatcaa cctccgcaag 1020
gaaatcgcca aagcgttcaa gggcaatgaa gggtacaaga gcctcttcaa gaaagacatc 1080
atcgaaacta tcctcccaga gtttctcgat gacaaggacg agatcgcgct ggtgaactcc 1140
tttaacgggt tcacaaccgc gtttaccggc ttctttgata acagggaaaa tatgttctcc 1200
gaggaggcca agtccaccag catcgccttc aggtgtatca acgagaacct cacccgctac 1260
atttccaata tggacatttt cgagaaggtg gatgcgatct tcgataagca cgaggtgcag 1320
gagatcaaag agaagattct caattccgat tatgacgtcg aggatttctt cgaaggggag 1380
ttctttaatt ttgtgctcac acaagagggc attgacgtgt acaacgcgat tatcgggggc 1440
ttcgtcacag agtccgggga gaagattaag gggctgaatg agtacatcaa tctgtacaat 1500
cagaagacca agcagaaact gccgaaattc aagccgctct acaagcaagt cctgtccgat 1560
agggaaagcc tctccttcta cggcgagggc tataccagcg acgaggaggt gctggaagtc 1620
ttccgcaaca cactgaataa gaatagcgag attttctcct ccatcaagaa gctcgagaag 1680
ctctttaaga actttgacga gtacagctcc gccgggattt tcgtgaagaa cgggccggcg 1740
atcagcacca tctccaagga catctttggc gagtggaacg tcatcaggga caagtggaac 1800
gccgagtacg acgacatcca cctgaagaag aaggcggtgg tgaccgagaa gtatgaggac 1860
gatcgcagga agtccttcaa aaaaatcggc tccttcagcc tcgaacagct ccaggagtat 1920
gccgatgcgg atctgtccgt cgtcgagaag ctgaaggaaa tcatcattca gaaggtcgac 1980
gagatctata aagtgtacgg gtccagcgag aagctgttcg acgccgactt tgtgctcgag 2040
aagtccctca aaaagaatga cgccgtggtg gccattatga aagacctgct cgactccgtg 2100
aagtccttcg aaaattacat taaagcgttc tttggggagg ggaaggaaac taacagggat 2160
gagtccttct atggcgactt tgtcctcgcg tacgacatcc tgctgaaggt cgaccacatt 2220
tacgacgcga tccgcaacta cgtgacacag aagccgtact ccaaagacaa gttcaagctg 2280
tacttccaga acccgcaatt tatggggggc tgggacaagg ataaagagac agactaccgc 2340
gcgacaattc tccgctatgg ctccaaatac tatctggcca tcatggacaa gaagtacgcg 2400
aagtgcctgc agaagatcga caaagacgac gtcaatggca actatgaaaa gatcaactac 2460
aagctgctgc cgggcccgaa caagatgctc ccgaaggtgt tcttcagcaa gaagtggatg 2520
gcctactaca atccaagcga ggatattcag aaaatctata aaaacgggac cttcaagaag 2580
ggggacatgt ttaacctcaa cgactgccac aagctcattg atttcttcaa ggatagcatt 2640
tcccgctacc cgaaatggtc caatgcgtac gattttaact tctccgagac agaaaagtac 2700
aaagacatcg cgggctttta cagggaggtg gaggagcaag ggtataaagt ttcttttgaa 2760
tccgcgagca agaaggaagt cgacaagctc gtcgaggagg gcaagctcta catgttccaa 2820
atttataaca aggacttttc cgacaagagc catgggaccc caaacctcca caccatgtac 2880
ttcaaactgc tctttgacga gaacaaccac gggcaaatca ggctgagcgg cggcgccgaa 2940
ttattcatgc gcagggcctc cctcaagaag gaagagctgg tcgtccatcc agccaattcc 3000
ccgatcgcga acaagaaccc ggacaatccg aaaaagacca ccaccctgtc ctacgacgtc 3060
tacaaggaca aacgcttcag cgaagaccag tacgaattac acatcccaat tgcgattaat 3120
aagtgcccaa agaatatctt caaaattaat acagaggtca gggtgctgct caaacacgac 3180
gacaatccgt atgtcatcgg cattgacagg ggcgagcgca atctgctcta tatcgtggtc 3240
gtggatggga agggcaatat tgtggagcag tactccctga acgagattat caacaacttc 3300
aatgggatta ggattaagac cgactatcac agcctgctcg acaagaaaga aaaagagagg 3360
tttgaggccc gccaaaactg gacctccatt gagaatatca aagaattaaa ggccggctat 3420
atttcccaag tcgtccacaa gatctgcgag ctggtggaga aatatgacgc cgtgattgcg 3480
ctcgaagact taaattctgg gttcaagaac tcccgcgtga aggtggaaaa acaggtgtat 3540
cagaaattcg agaaaatgct gatcgacaaa ctcaattata tggtggataa gaagtccaac 3600
ccgtgtgcca cagggggcgc gctgaagggc tatcagatca ccaacaagtt cgagagcttc 3660
aagagcatga gcacccagaa cgggtttatt ttctacatcc cggcgtggct cacctccaag 3720
attgacccga gcaccggctt cgtgaacctc ctgaagacaa agtatacctc cattgccgac 3780
agcaagaagt ttatctcctc cttcgaccgc attatgtatg tgccggagga ggacctcttc 3840
gagttcgccc tcgactacaa aaacttcagc cgcacagatg cggattacat caagaagtgg 3900
aagctgtact cctacgggaa caggatccgc atcttcagga atccaaaaaa aaataacgtc 3960
tttgactggg aggaagtgtg cctgacatcc gcctacaagg aactgttcaa taaatacggc 4020
atcaattacc agcagggcga cattcgcgcc ctcctctgtg agcagtccga caaagcgttt 4080
tactccagct tcatggccct catgtccctg atgctccaaa tgaggaatag catcacaggg 4140
cgcaccgacg tcgacttcct catcagcccg gtgaagaact ccgacgggat cttttacgac 4200
tcccgcaact atgaggcgca agagaatgcg atcctcccga agaacgccga tgcgaacggg 4260
gcctataata tcgccaggaa agtgctctgg gccatcgggc agttcaaaaa ggcggaggat 4320
gagaagctcg acaaggtgaa aattgccatt tccaacaagg agtggctgga gtacgcgcag 4380
acctccgtga agcactctgg cggctcaaaa agaaccgccg acggcagcga attcgagccc 4440
aagaagaaga ggaaagtcgg aagcggagct actaacttca gcctgctgaa gcaggctgga 4500
gacgtggagg agaaccctgg acctatggtg agcaagggcg aggagctgtt caccggggtg 4560
gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc acaagttcag cgtgtccggc 4620
gagggcgagg gcgatgccac ctacggcaag ctgaccctga agttcatctg caccaccggc 4680
aagctgcccg tgccctggcc caccctcgtg accaccctga cctatggagt gcagtgcttc 4740
agccgctacc ccgaccacat gaagcagcac gacttcttca agtccgccat gcccgaaggc 4800
tacgtccagg agcgcaccat cttcttcaag gacgacggca actacaagac ccgcgccgag 4860
gtgaagttcg agggcgacac cctggtgaac cgcatcgagc tgaagggcat cgacttcaag 4920
gaggacggca acatcctggg gcacaagctg gagtacaact acaacagcca caacgtctat 4980
atcatggccg acaagcagaa gaacggcatc aaggtgaact tcaagatccg ccacaacatc 5040
gaggacggca gcgtgcagct cgccgaccac taccagcaga acacccccat cggcgacggc 5100
cccgtgctgc tgcccgacaa ccactacctg agcacccagt ccgccctgag caaagacccc 5160
aacgagaagc gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg gatcactctc 5220
ggcatggacg agctgtacaa gtctggtggt tctcccaaga agaagaggaa agtctaaccg 5280
gtcatcatca ccatcaccat tgagtttaaa cccgctgatc agcctcgact gtgccttcta 5340
gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca 5400
ctcccactgt cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc 5460
attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagacaata 5520
gcaggcatgc tggggatgcg gtgggctcta tggcttctga ggcggaaaga accagctggg 5580
gctcgatacc gtcgacctct agctagagct tggcgtaatc atggtcatag ctgtttcctg 5640
tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta 5700
aagcctaggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg 5760
ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga 5820
gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg 5880
tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag 5940
aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc 6000
gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca 6060
aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt 6120
ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc 6180
tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc 6240
tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc 6300
ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact 6360
tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg 6420
ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta 6480
tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca 6540
aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa 6600
aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacact cagtggaacg 6660
aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc 6720
ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg 6780
acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat 6840
ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg 6900
gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa 6960
taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca 7020
tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc 7080
gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt 7140
cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa 7200
aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat 7260
cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct 7320
tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga 7380
gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag 7440
tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga 7500
gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca 7560
ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg 7620
cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc 7680
agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag 7740
gggttccgcg cacatttccc cgaaaagtgc cacctgacgt cgacggatcg ggagatcgat 7800
ctcccgatcc cctagggtcg actctcagta caatctgctc tgatgccgca tagttaagcc 7860
agtatctgct ccctgcttgt gtgttggagg tcgctgagta gtgcgcgagc aaaatttaag 7920
ctacaacaag gcaaggcttg accgacaatt gcatgaagaa tctgcttagg gttaggcgtt 7980
ttgcgctgct tcgcgatgta cgggccagat atacgcgtt 8019
<210> 89
<211> 10116
<212> DNA
<213> 人工的
<220>
<223> 合成的REDRAW编辑器
<400> 89
gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc 60
catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca 120
acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga 180
ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc 240
aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct 300
ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat 360
tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc 420
ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt 480
ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa 540
tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctggttta gtgaaccgtc 600
agatccgcta gagatccgcg gccgctaata cgactcacta tagggagagc cgccaccatg 660
aaacggacag ccgacggaag cgagttcgag tcaccaaaga agaagcggaa agtcacactt 720
aatattgagg atgaacatag attgcacgag acctctaagg aacctgatgt ttctcttgga 780
tcaacttggt tgtcagattt cccacaagca tgggcagaga ccggaggtat gggtcttgct 840
gttaggcagg caccacttat tattcctttg aaggcaacct ctactcctgt gtcaattaag 900
caatatccaa tgtctcagga agctaggctt ggaattaagc ctcacattca aagacttttg 960
gatcagggta ttttggtgcc atgtcaatca ccttggaaca caccactttt gcctgttaag 1020
aagcctggaa ctaatgatta cagaccagtg caagatttga gggaggttaa caagagagtg 1080
gaagatattc acccaactgt tccaaaccct tataatcttt tgtctggatt gccaccttca 1140
catcaatggt acactgtgct tgatttgaag gatgcatttt tctgccttag gttgcatcca 1200
acatctcagc ctctttttgc tttcgagtgg agagatcctg aaatgggaat ttctggtcaa 1260
cttacatgga ccaggttgcc tcagggtttc aagaactcac caaccttgtt taatgaggca 1320
cttcacaggg atttggctga ttttaggatt caacatcctg atcttatcct tttgcagtat 1380
gttgatgatc ttttgcttgc tgcaacttct gaattggatt gtcaacaggg aactagggca 1440
ttgcttcaaa cacttggaaa tttgggttac agagcttcag caaagaaggc tcagatttgc 1500
caaaagcagg ttaagtatct tggatacttg cttaaggaag gacaaaggtg gttgaccgag 1560
gctagaaagg aaactgtgat gggtcaacca acacctaaga cccctaggca gcttagagag 1620
ttcttgggaa aggcaggttt ttgtaggctt ttcattccag gatttgctga aatggctgca 1680
ccactttatc ctttgaccaa gcctggaact ttgtttaact ggggtccaga tcaacagaag 1740
gcataccaag aaattaagca ggctttgctt actgctccag cacttggttt gcctgatctt 1800
acaaagccat ttgagttgtt cgttgatgaa aagcaaggat atgcaaaggg tgtgcttacc 1860
cagaagttgg gaccttggag aaggcctgtt gcttaccttt ctaagaaact tgatccagtg 1920
gctgcaggtt ggccaccttg tcttagaatg gttgctgcaa ttgcagtgct tacaaaggat 1980
gctggaaagt tgactatggg acaacctctt gttattttgg caccacacgc tgttgaggca 2040
cttgtgaagc agccacctga taggtggttg tcaaacgcaa gaatgaccca ttatcaagct 2100
cttcttttgg atactgatag ggtgcagttc ggtcctgttg tggctttgaa tccagcaaca 2160
cttttgccac ttcctgagga aggattgcaa cacaactgcc ttgatatttt ggctgaggca 2220
catggtacaa gacctgatct taccgatcag ccattgcctg atgctgatca cacttggtac 2280
acagatggat cttcactttt gcaagaagga cagaggaagg ctggtgctgc agttactaca 2340
gagactgaag tgatttgggc taaggcactt ccagctggaa catctgctca aagagcagag 2400
cttattgctt tgacccaggc acttaagatg gctgaaggaa agaagttgaa cgtttacact 2460
gattctaggt atgctttcgc aacagctcat attcacggag aaatctatag aaggagagga 2520
tggttgacat cagagggaaa ggaaattaag aacaaggatg aaattcttgc acttttgaag 2580
gctctttttc ttcctaagag attgtctatt attcattgcc caggacacca aaagggtcat 2640
tcagcagaag ctaggggaaa tagaatggct gatcaggctg caagaaaggc tgcaattact 2700
gagacacctg atacctctac tcttttgatc gaaaactctt caccaaattc aaggcttatt 2760
aactctggca gcgaaactcc gggcacttcc gagtcagcta ctcctgagtc ttccaagctg 2820
gagaagttta caaactgtta cagcctctcc aaaaccctca ggtttaaagc gatcccggtg 2880
ggcaagaccc aggagaacat cgacaacaag aggctcctgg tggaagacga gaagcgcgcc 2940
gaagactaca agggcgtgaa gaagctgctc gataggtact acctcagctt tattaacgac 3000
gtgctgcaca gcatcaaact caagaatctc aacaactaca tctccctctt ccgcaaaaag 3060
acccgcaccg agaaggagaa caaggagctg gagaacctgg agatcaacct ccgcaaggaa 3120
atcgccaaag cgttcaaggg caatgaaggg tacaagagcc tcttcaagaa agacatcatc 3180
gaaactatcc tcccagagtt tctcgatgac aaggacgaga tcgcgctggt gaactccttt 3240
aacgggttca caaccgcgtt taccggcttc tttgataaca gggaaaatat gttctccgag 3300
gaggccaagt ccaccagcat cgccttcagg tgtatcaacg agaacctcac ccgctacatt 3360
tccaatatgg acattttcga gaaggtggat gcgatcttcg ataagcacga ggtgcaggag 3420
atcaaagaga agattctcaa ttccgattat gacgtcgagg atttcttcga aggggagttc 3480
tttaattttg tgctcacaca agagggcatt gacgtgtaca acgcgattat cgggggcttc 3540
gtcacagagt ccggggagaa gattaagggg ctgaatgagt acatcaatct gtacaatcag 3600
aagaccaagc agaaactgcc gaaattcaag ccgctctaca agcaagtcct gtccgatagg 3660
gaaagcctct ccttctacgg cgagggctat accagcgacg aggaggtgct ggaagtcttc 3720
cgcaacacac tgaataagaa tagcgagatt ttctcctcca tcaagaagct cgagaagctc 3780
tttaagaact ttgacgagta cagctccgcc gggattttcg tgaagaacgg gccggcgatc 3840
agcaccatct ccaaggacat ctttggcgag tggaacgtca tcagggacaa gtggaacgcc 3900
gagtacgacg acatccacct gaagaagaag gcggtggtga ccgagaagta tgaggacgat 3960
cgcaggaagt ccttcaaaaa aatcggctcc ttcagcctcg aacagctcca ggagtatgcc 4020
gatgcggatc tgtccgtcgt cgagaagctg aaggaaatca tcattcagaa ggtcgacgag 4080
atctataaag tgtacgggtc cagcgagaag ctgttcgacg ccgactttgt gctcgagaag 4140
tccctcaaaa agaatgacgc cgtggtggcc attatgaaag acctgctcga ctccgtgaag 4200
tccttcgaaa attacattaa agcgttcttt ggggagggga aggaaactaa cagggatgag 4260
tccttctatg gcgactttgt cctcgcgtac gacatcctgc tgaaggtcga ccacatttac 4320
gacgcgatcc gcaactacgt gacacagaag ccgtactcca aagacaagtt caagctgtac 4380
ttccagaacc cgcaatttat ggggggctgg gacaaggata aagagacaga ctaccgcgcg 4440
acaattctcc gctatggctc caaatactat ctggccatca tggacaagaa gtacgcgaag 4500
tgcctgcaga agatcgacaa agacgacgtc aatggcaact atgaaaagat caactacaag 4560
ctgctgccgg gcccgaacaa gatgctcccg aaggtgttct tcagcaagaa gtggatggcc 4620
tactacaatc caagcgagga tattcagaaa atctataaaa acgggacctt caagaagggg 4680
gacatgttta acctcaacga ctgccacaag ctcattgatt tcttcaagga tagcatttcc 4740
cgctacccga aatggtccaa tgcgtacgat tttaacttct ccgagacaga aaagtacaaa 4800
gacatcgcgg gcttttacag ggaggtggag gagcaagggt ataaagtttc ttttgaatcc 4860
gcgagcaaga aggaagtcga caagctcgtc gaggagggca agctctacat gttccaaatt 4920
tataacaagg acttttccga caagagccat gggaccccaa acctccacac catgtacttc 4980
aaactgctct ttgacgagaa caaccacggg caaatcaggc tgagcggcgg cgccgaatta 5040
ttcatgcgca gggcctccct caagaaggaa gagctggtcg tccatccagc caattccccg 5100
atcgcgaaca agaacccgga caatccgaaa aagaccacca ccctgtccta cgacgtctac 5160
aaggacaaac gcttcagcga agaccagtac gaattacaca tcccaattgc gattaataag 5220
tgcccaaaga atatcttcaa aattaataca gaggtcaggg tgctgctcaa acacgacgac 5280
aatccgtatg tcatcggcat tgacaggggc gagcgcaatc tgctctatat cgtggtcgtg 5340
gatgggaagg gcaatattgt ggagcagtac tccctgaacg agattatcaa caacttcaat 5400
gggattagga ttaagaccga ctatcacagc ctgctcgaca agaaagaaaa agagaggttt 5460
gaggcccgcc aaaactggac ctccattgag aatatcaaag aattaaaggc cggctatatt 5520
tcccaagtcg tccacaagat ctgcgagctg gtggagaaat atgacgccgt gattgcgctc 5580
gaagacttaa attctgggtt caagaactcc cgcgtgaagg tggaaaaaca ggtgtatcag 5640
aaattcgaga aaatgctgat cgacaaactc aattatatgg tggataagaa gtccaacccg 5700
tgtgccacag ggggcgcgct gaagggctat cagatcacca acaagttcga gagcttcaag 5760
agcatgagca cccagaacgg gtttattttc tacatcccgg cgtggctcac ctccaagatt 5820
gacccgagca ccggcttcgt gaacctcctg aagacaaagt atacctccat tgccgacagc 5880
aagaagttta tctcctcctt cgaccgcatt atgtatgtgc cggaggagga cctcttcgag 5940
ttcgccctcg actacaaaaa cttcagccgc acagatgcgg attacatcaa gaagtggaag 6000
ctgtactcct acgggaacag gatccgcatc ttcaggaatc caaaaaaaaa taacgtcttt 6060
gactgggagg aagtgtgcct gacatccgcc tacaaggaac tgttcaataa atacggcatc 6120
aattaccagc agggcgacat tcgcgccctc ctctgtgagc agtccgacaa agcgttttac 6180
tccagcttca tggccctcat gtccctgatg ctccaaatgg cgaatagcat cacagggcgc 6240
accgacgtcg acttcctcat cagcccggtg aagaactccg acgggatctt ttacgactcc 6300
cgcaactatg aggcgcaaga gaatgcgatc ctcccgaaga acgccgatgc gaacggggcc 6360
tataatatcg ccaggaaagt gctctgggcc atcgggcagt tcaaaaaggc ggaggatgag 6420
aagctcgaca aggtgaaaat tgccatttcc aacaaggagt ggctggagta cgcgcagacc 6480
tccgtgaagc actctggcgg ctcaaaaaga accgccgacg gcagcgaatt cgagcccaag 6540
aagaagagga aagtcggaag cggagctact aacttcagcc tgctgaagca ggctggagac 6600
gtggaggaga accctggacc tatggtgagc aagggcgagg agctgttcac cggggtggtg 6660
cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt gtccggcgag 6720
ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag 6780
ctgcccgtgc cctggcccac cctcgtgacc accctgacct atggagtgca gtgcttcagc 6840
cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac 6900
gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg 6960
aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag 7020
gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa cgtctatatc 7080
atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca caacatcgag 7140
gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc 7200
gtgctgctgc ccgacaacca ctacctgagc acccagtccg ccctgagcaa agaccccaac 7260
gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc 7320
atggacgagc tgtacaagtc tggtggttct cccaagaaga agaggaaagt ctaaccggtc 7380
atcatcacca tcaccattga gtttaaaccc gctgatcagc ctcgactgtg ccttctagtt 7440
gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc 7500
ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt 7560
ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca 7620
ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc agctggggct 7680
cgataccgtc gacctctagc tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt 7740
gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag 7800
cctagggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt 7860
tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag 7920
gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg 7980
ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat 8040
caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta 8100
aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa 8160
atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc 8220
cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt 8280
ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca 8340
gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg 8400
accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat 8460
cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta 8520
cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct 8580
gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac 8640
aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa 8700
aaggatctca agaagatcct ttgatctttt ctacggggtc tgacactcag tggaacgaaa 8760
actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt 8820
taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca 8880
gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca 8940
tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc 9000
ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa 9060
accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc 9120
agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca 9180
acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat 9240
tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag 9300
cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac 9360
tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt 9420
ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt 9480
gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc 9540
tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat 9600
ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca 9660
gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga 9720
cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg 9780
gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg 9840
ttccgcgcac atttccccga aaagtgccac ctgacgtcga cggatcggga gatcgatctc 9900
ccgatcccct agggtcgact ctcagtacaa tctgctctga tgccgcatag ttaagccagt 9960
atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg cgcgagcaaa atttaagcta 10020
caacaaggca aggcttgacc gacaattgca tgaagaatct gcttagggtt aggcgttttg 10080
cgctgcttcg cgatgtacgg gccagatata cgcgtt 10116
<210> 90
<211> 6717
<212> DNA
<213> 人工的
<220>
<223> 合成的tagRNA
<400> 90
atgaaacgga cagccgacgg aagcgagttc gagtcaccaa agaagaagcg gaaagtcaca 60
cttaatattg aggatgaaca tagattgcac gagacctcta aggaacctga tgtttctctt 120
ggatcaactt ggttgtcaga tttcccacaa gcatgggcag agaccggagg tatgggtctt 180
gctgttaggc aggcaccact tattattcct ttgaaggcaa cctctactcc tgtgtcaatt 240
aagcaatatc caatgtctca ggaagctagg cttggaatta agcctcacat tcaaagactt 300
ttggatcagg gtattttggt gccatgtcaa tcaccttgga acacaccact tttgcctgtt 360
aagaagcctg gaactaatga ttacagacca gtgcaagatt tgagggaggt taacaagaga 420
gtggaagata ttcacccaac tgttccaaac ccttataatc ttttgtctgg attgccacct 480
tcacatcaat ggtacactgt gcttgatttg aaggatgcat ttttctgcct taggttgcat 540
ccaacatctc agcctctttt tgctttcgag tggagagatc ctgaaatggg aatttctggt 600
caacttacat ggaccaggtt gcctcagggt ttcaagaact caccaacctt gtttaatgag 660
gcacttcaca gggatttggc tgattttagg attcaacatc ctgatcttat ccttttgcag 720
tatgttgatg atcttttgct tgctgcaact tctgaattgg attgtcaaca gggaactagg 780
gcattgcttc aaacacttgg aaatttgggt tacagagctt cagcaaagaa ggctcagatt 840
tgccaaaagc aggttaagta tcttggatac ttgcttaagg aaggacaaag gtggttgacc 900
gaggctagaa aggaaactgt gatgggtcaa ccaacaccta agacccctag gcagcttaga 960
gagttcttgg gaaaggcagg tttttgtagg cttttcattc caggatttgc tgaaatggct 1020
gcaccacttt atcctttgac caagcctgga actttgttta actggggtcc agatcaacag 1080
aaggcatacc aagaaattaa gcaggctttg cttactgctc cagcacttgg tttgcctgat 1140
cttacaaagc catttgagtt gttcgttgat gaaaagcaag gatatgcaaa gggtgtgctt 1200
acccagaagt tgggaccttg gagaaggcct gttgcttacc tttctaagaa acttgatcca 1260
gtggctgcag gttggccacc ttgtcttaga atggttgctg caattgcagt gcttacaaag 1320
gatgctggaa agttgactat gggacaacct cttgttattt tggcaccaca cgctgttgag 1380
gcacttgtga agcagccacc tgataggtgg ttgtcaaacg caagaatgac ccattatcaa 1440
gctcttcttt tggatactga tagggtgcag ttcggtcctg ttgtggcttt gaatccagca 1500
acacttttgc cacttcctga ggaaggattg caacacaact gccttgatat tttggctgag 1560
gcacatggta caagacctga tcttaccgat cagccattgc ctgatgctga tcacacttgg 1620
tacacagatg gatcttcact tttgcaagaa ggacagagga aggctggtgc tgcagttact 1680
acagagactg aagtgatttg ggctaaggca cttccagctg gaacatctgc tcaaagagca 1740
gagcttattg ctttgaccca ggcacttaag atggctgaag gaaagaagtt gaacgtttac 1800
actgattcta ggtatgcttt cgcaacagct catattcacg gagaaatcta tagaaggaga 1860
ggatggttga catcagaggg aaaggaaatt aagaacaagg atgaaattct tgcacttttg 1920
aaggctcttt ttcttcctaa gagattgtct attattcatt gcccaggaca ccaaaagggt 1980
cattcagcag aagctagggg aaatagaatg gctgatcagg ctgcaagaaa ggctgcaatt 2040
actgagacac ctgatacctc tactcttttg atcgaaaact cttcaccaaa ttcaaggctt 2100
attaactctg gcagcgaaac tccgggcact tccgagtcag ctactcctga gtcttccaag 2160
ctggagaagt ttacaaactg ttacagcctc tccaaaaccc tcaggtttaa agcgatcccg 2220
gtgggcaaga cccaggagaa catcgacaac aagaggctcc tggtggaaga cgagaagcgc 2280
gccgaagact acaagggcgt gaagaagctg ctcgataggt actacctcag ctttattaac 2340
gacgtgctgc acagcatcaa actcaagaat ctcaacaact acatctccct cttccgcaaa 2400
aagacccgca ccgagaagga gaacaaggag ctggagaacc tggagatcaa cctccgcaag 2460
gaaatcgcca aagcgttcaa gggcaatgaa gggtacaaga gcctcttcaa gaaagacatc 2520
atcgaaacta tcctcccaga gtttctcgat gacaaggacg agatcgcgct ggtgaactcc 2580
tttaacgggt tcacaaccgc gtttaccggc ttctttgata acagggaaaa tatgttctcc 2640
gaggaggcca agtccaccag catcgccttc aggtgtatca acgagaacct cacccgctac 2700
atttccaata tggacatttt cgagaaggtg gatgcgatct tcgataagca cgaggtgcag 2760
gagatcaaag agaagattct caattccgat tatgacgtcg aggatttctt cgaaggggag 2820
ttctttaatt ttgtgctcac acaagagggc attgacgtgt acaacgcgat tatcgggggc 2880
ttcgtcacag agtccgggga gaagattaag gggctgaatg agtacatcaa tctgtacaat 2940
cagaagacca agcagaaact gccgaaattc aagccgctct acaagcaagt cctgtccgat 3000
agggaaagcc tctccttcta cggcgagggc tataccagcg acgaggaggt gctggaagtc 3060
ttccgcaaca cactgaataa gaatagcgag attttctcct ccatcaagaa gctcgagaag 3120
ctctttaaga actttgacga gtacagctcc gccgggattt tcgtgaagaa cgggccggcg 3180
atcagcacca tctccaagga catctttggc gagtggaacg tcatcaggga caagtggaac 3240
gccgagtacg acgacatcca cctgaagaag aaggcggtgg tgaccgagaa gtatgaggac 3300
gatcgcagga agtccttcaa aaaaatcggc tccttcagcc tcgaacagct ccaggagtat 3360
gccgatgcgg atctgtccgt cgtcgagaag ctgaaggaaa tcatcattca gaaggtcgac 3420
gagatctata aagtgtacgg gtccagcgag aagctgttcg acgccgactt tgtgctcgag 3480
aagtccctca aaaagaatga cgccgtggtg gccattatga aagacctgct cgactccgtg 3540
aagtccttcg aaaattacat taaagcgttc tttggggagg ggaaggaaac taacagggat 3600
gagtccttct atggcgactt tgtcctcgcg tacgacatcc tgctgaaggt cgaccacatt 3660
tacgacgcga tccgcaacta cgtgacacag aagccgtact ccaaagacaa gttcaagctg 3720
tacttccaga acccgcaatt tatggggggc tgggacaagg ataaagagac agactaccgc 3780
gcgacaattc tccgctatgg ctccaaatac tatctggcca tcatggacaa gaagtacgcg 3840
aagtgcctgc agaagatcga caaagacgac gtcaatggca actatgaaaa gatcaactac 3900
aagctgctgc cgggcccgaa caagatgctc ccgaaggtgt tcttcagcaa gaagtggatg 3960
gcctactaca atccaagcga ggatattcag aaaatctata aaaacgggac cttcaagaag 4020
ggggacatgt ttaacctcaa cgactgccac aagctcattg atttcttcaa ggatagcatt 4080
tcccgctacc cgaaatggtc caatgcgtac gattttaact tctccgagac agaaaagtac 4140
aaagacatcg cgggctttta cagggaggtg gaggagcaag ggtataaagt ttcttttgaa 4200
tccgcgagca agaaggaagt cgacaagctc gtcgaggagg gcaagctcta catgttccaa 4260
atttataaca aggacttttc cgacaagagc catgggaccc caaacctcca caccatgtac 4320
ttcaaactgc tctttgacga gaacaaccac gggcaaatca ggctgagcgg cggcgccgaa 4380
ttattcatgc gcagggcctc cctcaagaag gaagagctgg tcgtccatcc agccaattcc 4440
ccgatcgcga acaagaaccc ggacaatccg aaaaagacca ccaccctgtc ctacgacgtc 4500
tacaaggaca aacgcttcag cgaagaccag tacgaattac acatcccaat tgcgattaat 4560
aagtgcccaa agaatatctt caaaattaat acagaggtca gggtgctgct caaacacgac 4620
gacaatccgt atgtcatcgg cattgacagg ggcgagcgca atctgctcta tatcgtggtc 4680
gtggatggga agggcaatat tgtggagcag tactccctga acgagattat caacaacttc 4740
aatgggatta ggattaagac cgactatcac agcctgctcg acaagaaaga aaaagagagg 4800
tttgaggccc gccaaaactg gacctccatt gagaatatca aagaattaaa ggccggctat 4860
atttcccaag tcgtccacaa gatctgcgag ctggtggaga aatatgacgc cgtgattgcg 4920
ctcgaagact taaattctgg gttcaagaac tcccgcgtga aggtggaaaa acaggtgtat 4980
cagaaattcg agaaaatgct gatcgacaaa ctcaattata tggtggataa gaagtccaac 5040
ccgtgtgcca cagggggcgc gctgaagggc tatcagatca ccaacaagtt cgagagcttc 5100
aagagcatga gcacccagaa cgggtttatt ttctacatcc cggcgtggct cacctccaag 5160
attgacccga gcaccggctt cgtgaacctc ctgaagacaa agtatacctc cattgccgac 5220
agcaagaagt ttatctcctc cttcgaccgc attatgtatg tgccggagga ggacctcttc 5280
gagttcgccc tcgactacaa aaacttcagc cgcacagatg cggattacat caagaagtgg 5340
aagctgtact cctacgggaa caggatccgc atcttcagga atccaaaaaa aaataacgtc 5400
tttgactggg aggaagtgtg cctgacatcc gcctacaagg aactgttcaa taaatacggc 5460
atcaattacc agcagggcga cattcgcgcc ctcctctgtg agcagtccga caaagcgttt 5520
tactccagct tcatggccct catgtccctg atgctccaaa tggcgaatag catcacaggg 5580
cgcaccgacg tcgacttcct catcagcccg gtgaagaact ccgacgggat cttttacgac 5640
tcccgcaact atgaggcgca agagaatgcg atcctcccga agaacgccga tgcgaacggg 5700
gcctataata tcgccaggaa agtgctctgg gccatcgggc agttcaaaaa ggcggaggat 5760
gagaagctcg acaaggtgaa aattgccatt tccaacaagg agtggctgga gtacgcgcag 5820
acctccgtga agcactctgg cggctcaaaa agaaccgccg acggcagcga attcgagccc 5880
aagaagaaga ggaaagtcgg aagcggagct actaacttca gcctgctgaa gcaggctgga 5940
gacgtggagg agaaccctgg acctatggtg agcaagggcg aggagctgtt caccggggtg 6000
gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc acaagttcag cgtgtccggc 6060
gagggcgagg gcgatgccac ctacggcaag ctgaccctga agttcatctg caccaccggc 6120
aagctgcccg tgccctggcc caccctcgtg accaccctga cctatggagt gcagtgcttc 6180
agccgctacc ccgaccacat gaagcagcac gacttcttca agtccgccat gcccgaaggc 6240
tacgtccagg agcgcaccat cttcttcaag gacgacggca actacaagac ccgcgccgag 6300
gtgaagttcg agggcgacac cctggtgaac cgcatcgagc tgaagggcat cgacttcaag 6360
gaggacggca acatcctggg gcacaagctg gagtacaact acaacagcca caacgtctat 6420
atcatggccg acaagcagaa gaacggcatc aaggtgaact tcaagatccg ccacaacatc 6480
gaggacggca gcgtgcagct cgccgaccac taccagcaga acacccccat cggcgacggc 6540
cccgtgctgc tgcccgacaa ccactacctg agcacccagt ccgccctgag caaagacccc 6600
aacgagaagc gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg gatcactctc 6660
ggcatggacg agctgtacaa gtctggtggt tctcccaaga agaagaggaa agtctaa 6717
<210> 91
<211> 2238
<212> PRT
<213> 人工的
<220>
<223> 合成的编辑器序列
<400> 91
Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys
1 5 10 15
Arg Lys Val Thr Leu Asn Ile Glu Asp Glu His Arg Leu His Glu Thr
20 25 30
Ser Lys Glu Pro Asp Val Ser Leu Gly Ser Thr Trp Leu Ser Asp Phe
35 40 45
Pro Gln Ala Trp Ala Glu Thr Gly Gly Met Gly Leu Ala Val Arg Gln
50 55 60
Ala Pro Leu Ile Ile Pro Leu Lys Ala Thr Ser Thr Pro Val Ser Ile
65 70 75 80
Lys Gln Tyr Pro Met Ser Gln Glu Ala Arg Leu Gly Ile Lys Pro His
85 90 95
Ile Gln Arg Leu Leu Asp Gln Gly Ile Leu Val Pro Cys Gln Ser Pro
100 105 110
Trp Asn Thr Pro Leu Leu Pro Val Lys Lys Pro Gly Thr Asn Asp Tyr
115 120 125
Arg Pro Val Gln Asp Leu Arg Glu Val Asn Lys Arg Val Glu Asp Ile
130 135 140
His Pro Thr Val Pro Asn Pro Tyr Asn Leu Leu Ser Gly Leu Pro Pro
145 150 155 160
Ser His Gln Trp Tyr Thr Val Leu Asp Leu Lys Asp Ala Phe Phe Cys
165 170 175
Leu Arg Leu His Pro Thr Ser Gln Pro Leu Phe Ala Phe Glu Trp Arg
180 185 190
Asp Pro Glu Met Gly Ile Ser Gly Gln Leu Thr Trp Thr Arg Leu Pro
195 200 205
Gln Gly Phe Lys Asn Ser Pro Thr Leu Phe Asn Glu Ala Leu His Arg
210 215 220
Asp Leu Ala Asp Phe Arg Ile Gln His Pro Asp Leu Ile Leu Leu Gln
225 230 235 240
Tyr Val Asp Asp Leu Leu Leu Ala Ala Thr Ser Glu Leu Asp Cys Gln
245 250 255
Gln Gly Thr Arg Ala Leu Leu Gln Thr Leu Gly Asn Leu Gly Tyr Arg
260 265 270
Ala Ser Ala Lys Lys Ala Gln Ile Cys Gln Lys Gln Val Lys Tyr Leu
275 280 285
Gly Tyr Leu Leu Lys Glu Gly Gln Arg Trp Leu Thr Glu Ala Arg Lys
290 295 300
Glu Thr Val Met Gly Gln Pro Thr Pro Lys Thr Pro Arg Gln Leu Arg
305 310 315 320
Glu Phe Leu Gly Lys Ala Gly Phe Cys Arg Leu Phe Ile Pro Gly Phe
325 330 335
Ala Glu Met Ala Ala Pro Leu Tyr Pro Leu Thr Lys Pro Gly Thr Leu
340 345 350
Phe Asn Trp Gly Pro Asp Gln Gln Lys Ala Tyr Gln Glu Ile Lys Gln
355 360 365
Ala Leu Leu Thr Ala Pro Ala Leu Gly Leu Pro Asp Leu Thr Lys Pro
370 375 380
Phe Glu Leu Phe Val Asp Glu Lys Gln Gly Tyr Ala Lys Gly Val Leu
385 390 395 400
Thr Gln Lys Leu Gly Pro Trp Arg Arg Pro Val Ala Tyr Leu Ser Lys
405 410 415
Lys Leu Asp Pro Val Ala Ala Gly Trp Pro Pro Cys Leu Arg Met Val
420 425 430
Ala Ala Ile Ala Val Leu Thr Lys Asp Ala Gly Lys Leu Thr Met Gly
435 440 445
Gln Pro Leu Val Ile Leu Ala Pro His Ala Val Glu Ala Leu Val Lys
450 455 460
Gln Pro Pro Asp Arg Trp Leu Ser Asn Ala Arg Met Thr His Tyr Gln
465 470 475 480
Ala Leu Leu Leu Asp Thr Asp Arg Val Gln Phe Gly Pro Val Val Ala
485 490 495
Leu Asn Pro Ala Thr Leu Leu Pro Leu Pro Glu Glu Gly Leu Gln His
500 505 510
Asn Cys Leu Asp Ile Leu Ala Glu Ala His Gly Thr Arg Pro Asp Leu
515 520 525
Thr Asp Gln Pro Leu Pro Asp Ala Asp His Thr Trp Tyr Thr Asp Gly
530 535 540
Ser Ser Leu Leu Gln Glu Gly Gln Arg Lys Ala Gly Ala Ala Val Thr
545 550 555 560
Thr Glu Thr Glu Val Ile Trp Ala Lys Ala Leu Pro Ala Gly Thr Ser
565 570 575
Ala Gln Arg Ala Glu Leu Ile Ala Leu Thr Gln Ala Leu Lys Met Ala
580 585 590
Glu Gly Lys Lys Leu Asn Val Tyr Thr Asp Ser Arg Tyr Ala Phe Ala
595 600 605
Thr Ala His Ile His Gly Glu Ile Tyr Arg Arg Arg Gly Trp Leu Thr
610 615 620
Ser Glu Gly Lys Glu Ile Lys Asn Lys Asp Glu Ile Leu Ala Leu Leu
625 630 635 640
Lys Ala Leu Phe Leu Pro Lys Arg Leu Ser Ile Ile His Cys Pro Gly
645 650 655
His Gln Lys Gly His Ser Ala Glu Ala Arg Gly Asn Arg Met Ala Asp
660 665 670
Gln Ala Ala Arg Lys Ala Ala Ile Thr Glu Thr Pro Asp Thr Ser Thr
675 680 685
Leu Leu Ile Glu Asn Ser Ser Pro Asn Ser Arg Leu Ile Asn Ser Gly
690 695 700
Ser Glu Thr Pro Gly Thr Ser Glu Ser Ala Thr Pro Glu Ser Ser Lys
705 710 715 720
Leu Glu Lys Phe Thr Asn Cys Tyr Ser Leu Ser Lys Thr Leu Arg Phe
725 730 735
Lys Ala Ile Pro Val Gly Lys Thr Gln Glu Asn Ile Asp Asn Lys Arg
740 745 750
Leu Leu Val Glu Asp Glu Lys Arg Ala Glu Asp Tyr Lys Gly Val Lys
755 760 765
Lys Leu Leu Asp Arg Tyr Tyr Leu Ser Phe Ile Asn Asp Val Leu His
770 775 780
Ser Ile Lys Leu Lys Asn Leu Asn Asn Tyr Ile Ser Leu Phe Arg Lys
785 790 795 800
Lys Thr Arg Thr Glu Lys Glu Asn Lys Glu Leu Glu Asn Leu Glu Ile
805 810 815
Asn Leu Arg Lys Glu Ile Ala Lys Ala Phe Lys Gly Asn Glu Gly Tyr
820 825 830
Lys Ser Leu Phe Lys Lys Asp Ile Ile Glu Thr Ile Leu Pro Glu Phe
835 840 845
Leu Asp Asp Lys Asp Glu Ile Ala Leu Val Asn Ser Phe Asn Gly Phe
850 855 860
Thr Thr Ala Phe Thr Gly Phe Phe Asp Asn Arg Glu Asn Met Phe Ser
865 870 875 880
Glu Glu Ala Lys Ser Thr Ser Ile Ala Phe Arg Cys Ile Asn Glu Asn
885 890 895
Leu Thr Arg Tyr Ile Ser Asn Met Asp Ile Phe Glu Lys Val Asp Ala
900 905 910
Ile Phe Asp Lys His Glu Val Gln Glu Ile Lys Glu Lys Ile Leu Asn
915 920 925
Ser Asp Tyr Asp Val Glu Asp Phe Phe Glu Gly Glu Phe Phe Asn Phe
930 935 940
Val Leu Thr Gln Glu Gly Ile Asp Val Tyr Asn Ala Ile Ile Gly Gly
945 950 955 960
Phe Val Thr Glu Ser Gly Glu Lys Ile Lys Gly Leu Asn Glu Tyr Ile
965 970 975
Asn Leu Tyr Asn Gln Lys Thr Lys Gln Lys Leu Pro Lys Phe Lys Pro
980 985 990
Leu Tyr Lys Gln Val Leu Ser Asp Arg Glu Ser Leu Ser Phe Tyr Gly
995 1000 1005
Glu Gly Tyr Thr Ser Asp Glu Glu Val Leu Glu Val Phe Arg Asn
1010 1015 1020
Thr Leu Asn Lys Asn Ser Glu Ile Phe Ser Ser Ile Lys Lys Leu
1025 1030 1035
Glu Lys Leu Phe Lys Asn Phe Asp Glu Tyr Ser Ser Ala Gly Ile
1040 1045 1050
Phe Val Lys Asn Gly Pro Ala Ile Ser Thr Ile Ser Lys Asp Ile
1055 1060 1065
Phe Gly Glu Trp Asn Val Ile Arg Asp Lys Trp Asn Ala Glu Tyr
1070 1075 1080
Asp Asp Ile His Leu Lys Lys Lys Ala Val Val Thr Glu Lys Tyr
1085 1090 1095
Glu Asp Asp Arg Arg Lys Ser Phe Lys Lys Ile Gly Ser Phe Ser
1100 1105 1110
Leu Glu Gln Leu Gln Glu Tyr Ala Asp Ala Asp Leu Ser Val Val
1115 1120 1125
Glu Lys Leu Lys Glu Ile Ile Ile Gln Lys Val Asp Glu Ile Tyr
1130 1135 1140
Lys Val Tyr Gly Ser Ser Glu Lys Leu Phe Asp Ala Asp Phe Val
1145 1150 1155
Leu Glu Lys Ser Leu Lys Lys Asn Asp Ala Val Val Ala Ile Met
1160 1165 1170
Lys Asp Leu Leu Asp Ser Val Lys Ser Phe Glu Asn Tyr Ile Lys
1175 1180 1185
Ala Phe Phe Gly Glu Gly Lys Glu Thr Asn Arg Asp Glu Ser Phe
1190 1195 1200
Tyr Gly Asp Phe Val Leu Ala Tyr Asp Ile Leu Leu Lys Val Asp
1205 1210 1215
His Ile Tyr Asp Ala Ile Arg Asn Tyr Val Thr Gln Lys Pro Tyr
1220 1225 1230
Ser Lys Asp Lys Phe Lys Leu Tyr Phe Gln Asn Pro Gln Phe Met
1235 1240 1245
Gly Gly Trp Asp Lys Asp Lys Glu Thr Asp Tyr Arg Ala Thr Ile
1250 1255 1260
Leu Arg Tyr Gly Ser Lys Tyr Tyr Leu Ala Ile Met Asp Lys Lys
1265 1270 1275
Tyr Ala Lys Cys Leu Gln Lys Ile Asp Lys Asp Asp Val Asn Gly
1280 1285 1290
Asn Tyr Glu Lys Ile Asn Tyr Lys Leu Leu Pro Gly Pro Asn Lys
1295 1300 1305
Met Leu Pro Lys Val Phe Phe Ser Lys Lys Trp Met Ala Tyr Tyr
1310 1315 1320
Asn Pro Ser Glu Asp Ile Gln Lys Ile Tyr Lys Asn Gly Thr Phe
1325 1330 1335
Lys Lys Gly Asp Met Phe Asn Leu Asn Asp Cys His Lys Leu Ile
1340 1345 1350
Asp Phe Phe Lys Asp Ser Ile Ser Arg Tyr Pro Lys Trp Ser Asn
1355 1360 1365
Ala Tyr Asp Phe Asn Phe Ser Glu Thr Glu Lys Tyr Lys Asp Ile
1370 1375 1380
Ala Gly Phe Tyr Arg Glu Val Glu Glu Gln Gly Tyr Lys Val Ser
1385 1390 1395
Phe Glu Ser Ala Ser Lys Lys Glu Val Asp Lys Leu Val Glu Glu
1400 1405 1410
Gly Lys Leu Tyr Met Phe Gln Ile Tyr Asn Lys Asp Phe Ser Asp
1415 1420 1425
Lys Ser His Gly Thr Pro Asn Leu His Thr Met Tyr Phe Lys Leu
1430 1435 1440
Leu Phe Asp Glu Asn Asn His Gly Gln Ile Arg Leu Ser Gly Gly
1445 1450 1455
Ala Glu Leu Phe Met Arg Arg Ala Ser Leu Lys Lys Glu Glu Leu
1460 1465 1470
Val Val His Pro Ala Asn Ser Pro Ile Ala Asn Lys Asn Pro Asp
1475 1480 1485
Asn Pro Lys Lys Thr Thr Thr Leu Ser Tyr Asp Val Tyr Lys Asp
1490 1495 1500
Lys Arg Phe Ser Glu Asp Gln Tyr Glu Leu His Ile Pro Ile Ala
1505 1510 1515
Ile Asn Lys Cys Pro Lys Asn Ile Phe Lys Ile Asn Thr Glu Val
1520 1525 1530
Arg Val Leu Leu Lys His Asp Asp Asn Pro Tyr Val Ile Gly Ile
1535 1540 1545
Asp Arg Gly Glu Arg Asn Leu Leu Tyr Ile Val Val Val Asp Gly
1550 1555 1560
Lys Gly Asn Ile Val Glu Gln Tyr Ser Leu Asn Glu Ile Ile Asn
1565 1570 1575
Asn Phe Asn Gly Ile Arg Ile Lys Thr Asp Tyr His Ser Leu Leu
1580 1585 1590
Asp Lys Lys Glu Lys Glu Arg Phe Glu Ala Arg Gln Asn Trp Thr
1595 1600 1605
Ser Ile Glu Asn Ile Lys Glu Leu Lys Ala Gly Tyr Ile Ser Gln
1610 1615 1620
Val Val His Lys Ile Cys Glu Leu Val Glu Lys Tyr Asp Ala Val
1625 1630 1635
Ile Ala Leu Glu Asp Leu Asn Ser Gly Phe Lys Asn Ser Arg Val
1640 1645 1650
Lys Val Glu Lys Gln Val Tyr Gln Lys Phe Glu Lys Met Leu Ile
1655 1660 1665
Asp Lys Leu Asn Tyr Met Val Asp Lys Lys Ser Asn Pro Cys Ala
1670 1675 1680
Thr Gly Gly Ala Leu Lys Gly Tyr Gln Ile Thr Asn Lys Phe Glu
1685 1690 1695
Ser Phe Lys Ser Met Ser Thr Gln Asn Gly Phe Ile Phe Tyr Ile
1700 1705 1710
Pro Ala Trp Leu Thr Ser Lys Ile Asp Pro Ser Thr Gly Phe Val
1715 1720 1725
Asn Leu Leu Lys Thr Lys Tyr Thr Ser Ile Ala Asp Ser Lys Lys
1730 1735 1740
Phe Ile Ser Ser Phe Asp Arg Ile Met Tyr Val Pro Glu Glu Asp
1745 1750 1755
Leu Phe Glu Phe Ala Leu Asp Tyr Lys Asn Phe Ser Arg Thr Asp
1760 1765 1770
Ala Asp Tyr Ile Lys Lys Trp Lys Leu Tyr Ser Tyr Gly Asn Arg
1775 1780 1785
Ile Arg Ile Phe Arg Asn Pro Lys Lys Asn Asn Val Phe Asp Trp
1790 1795 1800
Glu Glu Val Cys Leu Thr Ser Ala Tyr Lys Glu Leu Phe Asn Lys
1805 1810 1815
Tyr Gly Ile Asn Tyr Gln Gln Gly Asp Ile Arg Ala Leu Leu Cys
1820 1825 1830
Glu Gln Ser Asp Lys Ala Phe Tyr Ser Ser Phe Met Ala Leu Met
1835 1840 1845
Ser Leu Met Leu Gln Met Ala Asn Ser Ile Thr Gly Arg Thr Asp
1850 1855 1860
Val Asp Phe Leu Ile Ser Pro Val Lys Asn Ser Asp Gly Ile Phe
1865 1870 1875
Tyr Asp Ser Arg Asn Tyr Glu Ala Gln Glu Asn Ala Ile Leu Pro
1880 1885 1890
Lys Asn Ala Asp Ala Asn Gly Ala Tyr Asn Ile Ala Arg Lys Val
1895 1900 1905
Leu Trp Ala Ile Gly Gln Phe Lys Lys Ala Glu Asp Glu Lys Leu
1910 1915 1920
Asp Lys Val Lys Ile Ala Ile Ser Asn Lys Glu Trp Leu Glu Tyr
1925 1930 1935
Ala Gln Thr Ser Val Lys His Ser Gly Gly Ser Lys Arg Thr Ala
1940 1945 1950
Asp Gly Ser Glu Phe Glu Pro Lys Lys Lys Arg Lys Val Gly Ser
1955 1960 1965
Gly Ala Thr Asn Phe Ser Leu Leu Lys Gln Ala Gly Asp Val Glu
1970 1975 1980
Glu Asn Pro Gly Pro Met Val Ser Lys Gly Glu Glu Leu Phe Thr
1985 1990 1995
Gly Val Val Pro Ile Leu Val Glu Leu Asp Gly Asp Val Asn Gly
2000 2005 2010
His Lys Phe Ser Val Ser Gly Glu Gly Glu Gly Asp Ala Thr Tyr
2015 2020 2025
Gly Lys Leu Thr Leu Lys Phe Ile Cys Thr Thr Gly Lys Leu Pro
2030 2035 2040
Val Pro Trp Pro Thr Leu Val Thr Thr Leu Thr Tyr Gly Val Gln
2045 2050 2055
Cys Phe Ser Arg Tyr Pro Asp His Met Lys Gln His Asp Phe Phe
2060 2065 2070
Lys Ser Ala Met Pro Glu Gly Tyr Val Gln Glu Arg Thr Ile Phe
2075 2080 2085
Phe Lys Asp Asp Gly Asn Tyr Lys Thr Arg Ala Glu Val Lys Phe
2090 2095 2100
Glu Gly Asp Thr Leu Val Asn Arg Ile Glu Leu Lys Gly Ile Asp
2105 2110 2115
Phe Lys Glu Asp Gly Asn Ile Leu Gly His Lys Leu Glu Tyr Asn
2120 2125 2130
Tyr Asn Ser His Asn Val Tyr Ile Met Ala Asp Lys Gln Lys Asn
2135 2140 2145
Gly Ile Lys Val Asn Phe Lys Ile Arg His Asn Ile Glu Asp Gly
2150 2155 2160
Ser Val Gln Leu Ala Asp His Tyr Gln Gln Asn Thr Pro Ile Gly
2165 2170 2175
Asp Gly Pro Val Leu Leu Pro Asp Asn His Tyr Leu Ser Thr Gln
2180 2185 2190
Ser Ala Leu Ser Lys Asp Pro Asn Glu Lys Arg Asp His Met Val
2195 2200 2205
Leu Leu Glu Phe Val Thr Ala Ala Gly Ile Thr Leu Gly Met Asp
2210 2215 2220
Glu Leu Tyr Lys Ser Gly Gly Ser Pro Lys Lys Lys Arg Lys Val
2225 2230 2235
<210> 92
<211> 231
<212> DNA
<213> 人工的
<220>
<223> 合成的tagRNA
<400> 92
tatttctata agtgtagatt actcgtgtat atatactccg caccgaggtt ggtacgaaca 60
ccgggagtct ttaacacgac cgccacggat caggatcacg gagtgctcct gcaggttgtg 120
accttcacca ccgatgtagg aagtcacttc gaaaccgtta gtcagacgaa cacggcatac 180
tttacgcagc gcggagttcg gtttacgagg agtggtagta tatacacgag t 231
<210> 93
<211> 218
<212> DNA
<213> 人工的
<220>
<223> 合成的tagRNA
<400> 93
taatttctac taagtgtaga ttacggctcc gcagtggatg gcggtaagtc tccatagaat 60
ggaggacagc gcggagaatc tcgctctctc caggggaagc cgaagtttcc aaaaggtcgt 120
tgatcaaagc gcggcgcgtt gtttcatcaa ggcgtacggt caccgtaacc agcaaatcaa 180
tatcactgtg tggcttcagg ccgccatcca ctgcggat 218
<210> 94
<211> 213
<212> DNA
<213> 人工的
<220>
<223> 合成的tagRNA
<400> 94
taatttcaac taagtgtaga ttacggctcc gcagtggatg gcggtaagtc tccatagaat 60
ggagggcgga gaatctcgct ctctccaggg gaagccgaag tttccaaaag gtcgttgatc 120
aaagcgcggc gcgttgtttc atcaaggcgt acggtcaccg taaccagcaa atcaatatca 180
ctgtgtggct tcaggccgcc atccactgcg gat 213
<210> 95
<211> 21
<212> DNA
<213> 人工的
<220>
<223> 合成的诱饵发夹
<400> 95
taagtctcca tagaatggag g 21
<210> 96
<211> 1267
<212> PRT
<213> 人工的
<220>
<223> 合成的LbCas12a多肽
<400> 96
Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys
1 5 10 15
Arg Lys Val Ser Lys Leu Glu Lys Phe Thr Asn Cys Tyr Ser Leu Ser
20 25 30
Lys Thr Leu Arg Phe Lys Ala Ile Pro Val Gly Lys Thr Gln Glu Asn
35 40 45
Ile Asp Asn Lys Arg Leu Leu Val Glu Asp Glu Lys Arg Ala Glu Asp
50 55 60
Tyr Lys Gly Val Lys Lys Leu Leu Asp Arg Tyr Tyr Leu Ser Phe Ile
65 70 75 80
Asn Asp Val Leu His Ser Ile Lys Leu Lys Asn Leu Asn Asn Tyr Ile
85 90 95
Ser Leu Phe Arg Lys Lys Thr Arg Thr Glu Lys Glu Asn Lys Glu Leu
100 105 110
Glu Asn Leu Glu Ile Asn Leu Arg Lys Glu Ile Ala Lys Ala Phe Lys
115 120 125
Gly Asn Glu Gly Tyr Lys Ser Leu Phe Lys Lys Asp Ile Ile Glu Thr
130 135 140
Ile Leu Pro Glu Phe Leu Asp Asp Lys Asp Glu Ile Ala Leu Val Asn
145 150 155 160
Ser Phe Asn Gly Phe Thr Thr Ala Phe Thr Gly Phe Phe Asp Asn Arg
165 170 175
Glu Asn Met Phe Ser Glu Glu Ala Lys Ser Thr Ser Ile Ala Phe Arg
180 185 190
Cys Ile Asn Glu Asn Leu Thr Arg Tyr Ile Ser Asn Met Asp Ile Phe
195 200 205
Glu Lys Val Asp Ala Ile Phe Asp Lys His Glu Val Gln Glu Ile Lys
210 215 220
Glu Lys Ile Leu Asn Ser Asp Tyr Asp Val Glu Asp Phe Phe Glu Gly
225 230 235 240
Glu Phe Phe Asn Phe Val Leu Thr Gln Glu Gly Ile Asp Val Tyr Asn
245 250 255
Ala Ile Ile Gly Gly Phe Val Thr Glu Ser Gly Glu Lys Ile Lys Gly
260 265 270
Leu Asn Glu Tyr Ile Asn Leu Tyr Asn Gln Lys Thr Lys Gln Lys Leu
275 280 285
Pro Lys Phe Lys Pro Leu Tyr Lys Gln Val Leu Ser Asp Arg Glu Ser
290 295 300
Leu Ser Phe Tyr Gly Glu Gly Tyr Thr Ser Asp Glu Glu Val Leu Glu
305 310 315 320
Val Phe Arg Asn Thr Leu Asn Lys Asn Ser Glu Ile Phe Ser Ser Ile
325 330 335
Lys Lys Leu Glu Lys Leu Phe Lys Asn Phe Asp Glu Tyr Ser Ser Ala
340 345 350
Gly Ile Phe Val Lys Asn Gly Pro Ala Ile Ser Thr Ile Ser Lys Asp
355 360 365
Ile Phe Gly Glu Trp Asn Val Ile Arg Asp Lys Trp Asn Ala Glu Tyr
370 375 380
Asp Asp Ile His Leu Lys Lys Lys Ala Val Val Thr Glu Lys Tyr Glu
385 390 395 400
Asp Asp Arg Arg Lys Ser Phe Lys Lys Ile Gly Ser Phe Ser Leu Glu
405 410 415
Gln Leu Gln Glu Tyr Ala Asp Ala Asp Leu Ser Val Val Glu Lys Leu
420 425 430
Lys Glu Ile Ile Ile Gln Lys Val Asp Glu Ile Tyr Lys Val Tyr Gly
435 440 445
Ser Ser Glu Lys Leu Phe Asp Ala Asp Phe Val Leu Glu Lys Ser Leu
450 455 460
Lys Lys Asn Asp Ala Val Val Ala Ile Met Lys Asp Leu Leu Asp Ser
465 470 475 480
Val Lys Ser Phe Glu Asn Tyr Ile Lys Ala Phe Phe Gly Glu Gly Lys
485 490 495
Glu Thr Asn Arg Asp Glu Ser Phe Tyr Gly Asp Phe Val Leu Ala Tyr
500 505 510
Asp Ile Leu Leu Lys Val Asp His Ile Tyr Asp Ala Ile Arg Asn Tyr
515 520 525
Val Thr Gln Lys Pro Tyr Ser Lys Asp Lys Phe Lys Leu Tyr Phe Gln
530 535 540
Asn Pro Gln Phe Met Gly Gly Trp Asp Lys Asp Lys Glu Thr Asp Tyr
545 550 555 560
Arg Ala Thr Ile Leu Arg Tyr Gly Ser Lys Tyr Tyr Leu Ala Ile Met
565 570 575
Asp Lys Lys Tyr Ala Lys Cys Leu Gln Lys Ile Asp Lys Asp Asp Val
580 585 590
Asn Gly Asn Tyr Glu Lys Ile Asn Tyr Lys Leu Leu Pro Gly Pro Asn
595 600 605
Lys Met Leu Pro Lys Val Phe Phe Ser Lys Lys Trp Met Ala Tyr Tyr
610 615 620
Asn Pro Ser Glu Asp Ile Gln Lys Ile Tyr Lys Asn Gly Thr Phe Lys
625 630 635 640
Lys Gly Asp Met Phe Asn Leu Asn Asp Cys His Lys Leu Ile Asp Phe
645 650 655
Phe Lys Asp Ser Ile Ser Arg Tyr Pro Lys Trp Ser Asn Ala Tyr Asp
660 665 670
Phe Asn Phe Ser Glu Thr Glu Lys Tyr Lys Asp Ile Ala Gly Phe Tyr
675 680 685
Arg Glu Val Glu Glu Gln Gly Tyr Lys Val Ser Phe Glu Ser Ala Ser
690 695 700
Lys Lys Glu Val Asp Lys Leu Val Glu Glu Gly Lys Leu Tyr Met Phe
705 710 715 720
Gln Ile Tyr Asn Lys Asp Phe Ser Asp Lys Ser His Gly Thr Pro Asn
725 730 735
Leu His Thr Met Tyr Phe Lys Leu Leu Phe Asp Glu Asn Asn His Gly
740 745 750
Gln Ile Arg Leu Ser Gly Gly Ala Glu Leu Phe Met Arg Arg Ala Ser
755 760 765
Leu Lys Lys Glu Glu Leu Val Val Ala Pro Ala Asn Ser Pro Ile Ala
770 775 780
Asn Lys Asn Pro Asp Asn Pro Lys Lys Thr Thr Thr Leu Ser Tyr Asp
785 790 795 800
Val Tyr Lys Asp Lys Arg Phe Ser Glu Asp Gln Tyr Glu Leu His Ile
805 810 815
Pro Ile Ala Ile Asn Lys Cys Pro Lys Asn Ile Phe Lys Ile Asn Thr
820 825 830
Glu Val Arg Val Leu Leu Lys His Asp Asp Asn Pro Tyr Val Ile Gly
835 840 845
Ile Asp Arg Gly Glu Arg Asn Leu Leu Tyr Ile Val Val Val Asp Gly
850 855 860
Lys Gly Asn Ile Val Glu Gln Tyr Ser Leu Asn Glu Ile Ile Asn Asn
865 870 875 880
Phe Asn Gly Ile Arg Ile Lys Thr Asp Tyr His Ser Leu Leu Asp Lys
885 890 895
Lys Glu Lys Glu Arg Phe Glu Ala Arg Gln Asn Trp Thr Ser Ile Glu
900 905 910
Asn Ile Lys Glu Leu Lys Ala Gly Tyr Ile Ser Gln Val Val His Lys
915 920 925
Ile Cys Glu Leu Val Glu Lys Tyr Asp Ala Val Ile Ala Leu Glu Asp
930 935 940
Leu Asn Ser Gly Phe Lys Asn Ser Arg Val Lys Val Glu Lys Gln Val
945 950 955 960
Tyr Gln Lys Phe Glu Lys Met Leu Ile Asp Lys Leu Asn Tyr Met Val
965 970 975
Asp Lys Lys Ser Asn Pro Cys Ala Thr Gly Gly Ala Leu Lys Gly Tyr
980 985 990
Gln Ile Thr Asn Lys Phe Glu Ser Phe Lys Ser Met Ser Thr Gln Asn
995 1000 1005
Gly Phe Ile Phe Tyr Ile Pro Ala Trp Leu Thr Ser Lys Ile Asp
1010 1015 1020
Pro Ser Thr Gly Phe Val Asn Leu Leu Lys Thr Lys Tyr Thr Ser
1025 1030 1035
Ile Ala Asp Ser Lys Lys Phe Ile Ser Ser Phe Asp Arg Ile Met
1040 1045 1050
Tyr Val Pro Glu Glu Asp Leu Phe Glu Phe Ala Leu Asp Tyr Lys
1055 1060 1065
Asn Phe Ser Arg Thr Asp Ala Asp Tyr Ile Lys Lys Trp Lys Leu
1070 1075 1080
Tyr Ser Tyr Gly Asn Arg Ile Arg Ile Phe Arg Asn Pro Lys Lys
1085 1090 1095
Asn Asn Val Phe Asp Trp Glu Glu Val Cys Leu Thr Ser Ala Tyr
1100 1105 1110
Lys Glu Leu Phe Asn Lys Tyr Gly Ile Asn Tyr Gln Gln Gly Asp
1115 1120 1125
Ile Arg Ala Leu Leu Cys Glu Gln Ser Asp Lys Ala Phe Tyr Ser
1130 1135 1140
Ser Phe Met Ala Leu Met Ser Leu Met Leu Gln Met Arg Asn Ser
1145 1150 1155
Ile Thr Gly Arg Thr Asp Val Asp Phe Leu Ile Ser Pro Val Lys
1160 1165 1170
Asn Ser Asp Gly Ile Phe Tyr Asp Ser Arg Asn Tyr Glu Ala Gln
1175 1180 1185
Glu Asn Ala Ile Leu Pro Lys Asn Ala Asp Ala Asn Gly Ala Tyr
1190 1195 1200
Asn Ile Ala Arg Lys Val Leu Trp Ala Ile Gly Gln Phe Lys Lys
1205 1210 1215
Ala Glu Asp Glu Lys Leu Asp Lys Val Lys Ile Ala Ile Ser Asn
1220 1225 1230
Lys Glu Trp Leu Glu Tyr Ala Gln Thr Ser Val Lys His Ser Gly
1235 1240 1245
Gly Ser Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Pro Lys Lys
1250 1255 1260
Lys Arg Lys Val
1265
<210> 97
<211> 717
<212> PRT
<213> 人工的
<220>
<223> 合成的核酸外切酶
<400> 97
Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys
1 5 10 15
Arg Lys Val Thr Leu Asn Ile Glu Asp Glu His Arg Leu His Glu Thr
20 25 30
Ser Lys Glu Pro Asp Val Ser Leu Gly Ser Thr Trp Leu Ser Asp Phe
35 40 45
Pro Gln Ala Trp Ala Glu Thr Gly Gly Met Gly Leu Ala Val Arg Gln
50 55 60
Ala Pro Leu Ile Ile Pro Leu Lys Ala Thr Ser Thr Pro Val Ser Ile
65 70 75 80
Lys Gln Tyr Pro Met Ser Gln Glu Ala Arg Leu Gly Ile Lys Pro His
85 90 95
Ile Gln Arg Leu Leu Asp Gln Gly Ile Leu Val Pro Cys Gln Ser Pro
100 105 110
Trp Asn Thr Pro Leu Leu Pro Val Lys Lys Pro Gly Thr Asn Asp Tyr
115 120 125
Arg Pro Val Gln Asp Leu Arg Glu Val Asn Lys Arg Val Glu Asp Ile
130 135 140
His Pro Thr Val Pro Asn Pro Tyr Asn Leu Leu Ser Gly Leu Pro Pro
145 150 155 160
Ser His Gln Trp Tyr Thr Val Leu Asp Leu Lys Asp Ala Phe Phe Cys
165 170 175
Leu Arg Leu His Pro Thr Ser Gln Pro Leu Phe Ala Phe Glu Trp Arg
180 185 190
Asp Pro Glu Met Gly Ile Ser Gly Gln Leu Thr Trp Thr Arg Leu Pro
195 200 205
Gln Gly Phe Lys Asn Ser Pro Thr Leu Phe Asn Glu Ala Leu His Arg
210 215 220
Asp Leu Ala Asp Phe Arg Ile Gln His Pro Asp Leu Ile Leu Leu Gln
225 230 235 240
Tyr Val Asp Asp Leu Leu Leu Ala Ala Thr Ser Glu Leu Asp Cys Gln
245 250 255
Gln Gly Thr Arg Ala Leu Leu Gln Thr Leu Gly Asn Leu Gly Tyr Arg
260 265 270
Ala Ser Ala Lys Lys Ala Gln Ile Cys Gln Lys Gln Val Lys Tyr Leu
275 280 285
Gly Tyr Leu Leu Lys Glu Gly Gln Arg Trp Leu Thr Glu Ala Arg Lys
290 295 300
Glu Thr Val Met Gly Gln Pro Thr Pro Lys Thr Pro Arg Gln Leu Arg
305 310 315 320
Glu Phe Leu Gly Lys Ala Gly Phe Cys Arg Leu Phe Ile Pro Gly Phe
325 330 335
Ala Glu Met Ala Ala Pro Leu Tyr Pro Leu Thr Lys Pro Gly Thr Leu
340 345 350
Phe Asn Trp Gly Pro Asp Gln Gln Lys Ala Tyr Gln Glu Ile Lys Gln
355 360 365
Ala Leu Leu Thr Ala Pro Ala Leu Gly Leu Pro Asp Leu Thr Lys Pro
370 375 380
Phe Glu Leu Phe Val Asp Glu Lys Gln Gly Tyr Ala Lys Gly Val Leu
385 390 395 400
Thr Gln Lys Leu Gly Pro Trp Arg Arg Pro Val Ala Tyr Leu Ser Lys
405 410 415
Lys Leu Asp Pro Val Ala Ala Gly Trp Pro Pro Cys Leu Arg Met Val
420 425 430
Ala Ala Ile Ala Val Leu Thr Lys Asp Ala Gly Lys Leu Thr Met Gly
435 440 445
Gln Pro Leu Val Ile Leu Ala Pro His Ala Val Glu Ala Leu Val Lys
450 455 460
Gln Pro Pro Asp Arg Trp Leu Ser Asn Ala Arg Met Thr His Tyr Gln
465 470 475 480
Ala Leu Leu Leu Asp Thr Asp Arg Val Gln Phe Gly Pro Val Val Ala
485 490 495
Leu Asn Pro Ala Thr Leu Leu Pro Leu Pro Glu Glu Gly Leu Gln His
500 505 510
Asn Cys Leu Asp Ile Leu Ala Glu Ala His Gly Thr Arg Pro Asp Leu
515 520 525
Thr Asp Gln Pro Leu Pro Asp Ala Asp His Thr Trp Tyr Thr Asp Gly
530 535 540
Ser Ser Leu Leu Gln Glu Gly Gln Arg Lys Ala Gly Ala Ala Val Thr
545 550 555 560
Thr Glu Thr Glu Val Ile Trp Ala Lys Ala Leu Pro Ala Gly Thr Ser
565 570 575
Ala Gln Arg Ala Glu Leu Ile Ala Leu Thr Gln Ala Leu Lys Met Ala
580 585 590
Glu Gly Lys Lys Leu Asn Val Tyr Thr Asp Ser Arg Tyr Ala Phe Ala
595 600 605
Thr Ala His Ile His Gly Glu Ile Tyr Arg Arg Arg Gly Trp Leu Thr
610 615 620
Ser Glu Gly Lys Glu Ile Lys Asn Lys Asp Glu Ile Leu Ala Leu Leu
625 630 635 640
Lys Ala Leu Phe Leu Pro Lys Arg Leu Ser Ile Ile His Cys Pro Gly
645 650 655
His Gln Lys Gly His Ser Ala Glu Ala Arg Gly Asn Arg Met Ala Asp
660 665 670
Gln Ala Ala Arg Lys Ala Ala Ile Thr Glu Thr Pro Asp Thr Ser Thr
675 680 685
Leu Leu Ile Glu Asn Ser Ser Pro Ser Gly Gly Ser Lys Arg Thr Ala
690 695 700
Asp Gly Ser Glu Phe Glu Pro Lys Lys Lys Arg Lys Val
705 710 715
<210> 98
<211> 343
<212> PRT
<213> 人工的
<220>
<223> 合成的核酸外切酶
<400> 98
Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys
1 5 10 15
Arg Lys Val Glu His Pro His Asn Glu Asn Ala Gly Ser Asp Pro His
20 25 30
Arg Asp Cys Ser Asp Glu Thr Gly Glu Val Ala Asp Pro Val Ile Val
35 40 45
Glu Asp Ile Glu Pro Gly Ile Tyr Tyr Gly Ile Ser Asn Glu Asn Tyr
50 55 60
His Ala Gly Pro Gly Ile Ser Lys Ser Gln Leu Asp Asp Ile Ala Asp
65 70 75 80
Thr Pro Ala Leu Tyr Leu Trp Arg Lys Asn Ala Pro Val Asp Thr Thr
85 90 95
Lys Thr Lys Thr Leu Asp Leu Gly Thr Ala Phe His Cys Arg Val Leu
100 105 110
Glu Pro Glu Glu Phe Ser Asn Arg Phe Ile Val Ala Pro Glu Phe Asn
115 120 125
Arg Arg Thr Asn Ala Gly Lys Glu Glu Glu Lys Ala Phe Leu Met Glu
130 135 140
Cys Ala Ser Thr Gly Lys Thr Val Ile Thr Ala Glu Glu Gly Arg Lys
145 150 155 160
Ile Glu Leu Met Tyr Gln Ser Val Met Ala Leu Pro Leu Gly Gln Trp
165 170 175
Leu Val Glu Ser Ala Gly His Ala Glu Ser Ser Ile Tyr Trp Glu Asp
180 185 190
Pro Glu Thr Gly Ile Leu Cys Arg Cys Arg Pro Asp Lys Ile Ile Pro
195 200 205
Glu Phe His Trp Ile Met Asp Val Lys Thr Thr Ala Asp Ile Gln Arg
210 215 220
Phe Lys Thr Ala Tyr Tyr Asp Tyr Arg Tyr His Val Gln Asp Ala Phe
225 230 235 240
Tyr Ser Asp Gly Tyr Glu Ala Gln Phe Gly Val Gln Pro Thr Phe Val
245 250 255
Phe Leu Val Ala Ser Thr Thr Ile Glu Cys Gly Arg Tyr Pro Val Glu
260 265 270
Ile Phe Met Met Gly Glu Glu Ala Lys Leu Ala Gly Gln Gln Glu Tyr
275 280 285
His Arg Asn Leu Arg Thr Leu Ser Asp Cys Leu Asn Thr Asp Glu Trp
290 295 300
Pro Ala Ile Lys Thr Leu Ser Leu Pro Arg Trp Ala Lys Glu Tyr Ala
305 310 315 320
Asn Asp Ser Gly Gly Ser Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu
325 330 335
Pro Lys Lys Lys Arg Lys Val
340
<210> 99
<211> 617
<212> PRT
<213> 人工的
<220>
<223> 合成的核酸外切酶
<400> 99
Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys
1 5 10 15
Arg Lys Val Met Lys Gln Gln Ile Gln Leu Arg Arg Arg Glu Val Asp
20 25 30
Glu Thr Ala Asp Leu Pro Ala Glu Leu Pro Pro Leu Leu Arg Arg Leu
35 40 45
Tyr Ala Ser Arg Gly Val Arg Ser Ala Gln Glu Leu Glu Arg Ser Val
50 55 60
Lys Gly Met Leu Pro Trp Gln Gln Leu Ser Gly Val Glu Lys Ala Val
65 70 75 80
Glu Ile Leu Tyr Asn Ala Phe Arg Glu Gly Thr Arg Ile Ile Val Val
85 90 95
Gly Asp Phe Asp Ala Asp Gly Ala Thr Ser Thr Ala Leu Ser Val Leu
100 105 110
Ala Met Arg Ser Leu Gly Cys Ser Asn Ile Asp Tyr Leu Val Pro Asn
115 120 125
Arg Phe Glu Asp Gly Tyr Gly Leu Ser Pro Glu Val Val Asp Gln Ala
130 135 140
His Ala Arg Gly Ala Gln Leu Ile Val Thr Val Asp Asn Gly Ile Ser
145 150 155 160
Ser His Ala Gly Val Glu His Ala Arg Ser Leu Gly Ile Pro Val Ile
165 170 175
Val Thr Asp His His Leu Pro Gly Asp Thr Leu Pro Ala Ala Glu Ala
180 185 190
Ile Ile Asn Pro Asn Leu Arg Asp Cys Asn Phe Pro Ser Lys Ser Leu
195 200 205
Ala Gly Val Gly Val Ala Phe Tyr Leu Met Leu Ala Leu Arg Thr Phe
210 215 220
Leu Arg Asp Gln Gly Trp Phe Asp Glu Arg Asn Ile Ala Ile Pro Asn
225 230 235 240
Leu Ala Glu Leu Leu Asp Leu Val Ala Leu Gly Thr Val Ala Asp Val
245 250 255
Val Pro Leu Asp Ala Asn Asn Arg Ile Leu Thr Trp Gln Gly Met Ser
260 265 270
Arg Ile Arg Ala Gly Lys Cys Arg Pro Gly Ile Lys Ala Leu Leu Glu
275 280 285
Val Ala Asn Arg Asp Ala Gln Lys Leu Ala Ala Ser Asp Leu Gly Phe
290 295 300
Ala Leu Gly Pro Arg Leu Asn Ala Ala Gly Arg Leu Asp Asp Met Ser
305 310 315 320
Val Gly Val Ala Leu Leu Leu Cys Asp Asn Ile Gly Glu Ala Arg Val
325 330 335
Leu Ala Asn Glu Leu Asp Ala Leu Asn Gln Thr Arg Lys Glu Ile Glu
340 345 350
Gln Gly Met Gln Ile Glu Ala Leu Thr Leu Cys Glu Lys Leu Glu Arg
355 360 365
Ser Arg Asp Thr Leu Pro Gly Gly Leu Ala Met Tyr His Pro Glu Trp
370 375 380
His Gln Gly Val Val Gly Ile Leu Ala Ser Arg Ile Lys Glu Arg Phe
385 390 395 400
His Arg Pro Val Ile Ala Phe Ala Pro Ala Gly Asp Gly Thr Leu Lys
405 410 415
Gly Ser Gly Arg Ser Ile Gln Gly Leu His Met Arg Asp Ala Leu Glu
420 425 430
Arg Leu Asp Thr Leu Tyr Pro Gly Met Met Leu Lys Phe Gly Gly His
435 440 445
Ala Met Ala Ala Gly Leu Ser Leu Glu Glu Asp Lys Phe Lys Leu Phe
450 455 460
Gln Gln Arg Phe Gly Glu Leu Val Thr Glu Trp Leu Asp Pro Ser Leu
465 470 475 480
Leu Gln Gly Glu Val Val Ser Asp Gly Pro Leu Ser Pro Ala Glu Met
485 490 495
Thr Met Glu Val Ala Gln Leu Leu Arg Asp Ala Gly Pro Trp Gly Gln
500 505 510
Met Phe Pro Glu Pro Leu Phe Asp Gly His Phe Arg Leu Leu Gln Gln
515 520 525
Arg Leu Val Gly Glu Arg His Leu Lys Val Met Val Glu Pro Val Gly
530 535 540
Gly Gly Pro Leu Leu Asp Gly Ile Ala Phe Asn Val Asp Thr Ala Leu
545 550 555 560
Trp Pro Asp Asn Gly Val Arg Glu Val Gln Leu Ala Tyr Lys Leu Asp
565 570 575
Ile Asn Glu Phe Arg Gly Asn Arg Ser Leu Gln Ile Ile Ile Asp Asn
580 585 590
Ile Trp Pro Ile Ser Gly Gly Ser Lys Arg Thr Ala Asp Gly Ser Glu
595 600 605
Phe Glu Pro Lys Lys Lys Arg Lys Val
610 615
<210> 100
<211> 330
<212> PRT
<213> 人工的
<220>
<223> 合成的核酸外切酶
<400> 100
Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys
1 5 10 15
Arg Lys Val Ser Lys Ser Trp Gly Lys Phe Ile Glu Glu Glu Glu Ala
20 25 30
Glu Met Ala Ser Arg Arg Asn Leu Met Ile Val Asp Gly Thr Asn Leu
35 40 45
Gly Phe Arg Phe Lys His Asn Asn Ser Lys Lys Pro Phe Ala Ser Ser
50 55 60
Tyr Val Ser Thr Ile Gln Ser Leu Ala Lys Ser Tyr Ser Ala Arg Thr
65 70 75 80
Thr Ile Val Leu Gly Asp Lys Gly Lys Ser Val Phe Arg Leu Glu His
85 90 95
Leu Pro Glu Tyr Lys Gly Asn Arg Asp Glu Lys Tyr Ala Gln Arg Thr
100 105 110
Glu Glu Glu Lys Ala Leu Asp Glu Gln Phe Phe Glu Tyr Leu Lys Asp
115 120 125
Ala Phe Glu Leu Cys Lys Thr Thr Phe Pro Thr Phe Thr Ile Arg Gly
130 135 140
Val Glu Ala Asp Asp Met Ala Ala Tyr Ile Val Lys Leu Ile Gly His
145 150 155 160
Leu Tyr Asp His Val Trp Leu Ile Ser Thr Asp Gly Asp Trp Asp Thr
165 170 175
Leu Leu Thr Asp Lys Val Ser Arg Phe Ser Phe Thr Thr Arg Arg Glu
180 185 190
Tyr His Leu Arg Asp Met Tyr Glu His His Asn Val Asp Asp Val Glu
195 200 205
Gln Phe Ile Ser Leu Lys Ala Ile Met Gly Asp Leu Gly Asp Asn Ile
210 215 220
Arg Gly Val Glu Gly Ile Gly Ala Lys Arg Gly Tyr Asn Ile Ile Arg
225 230 235 240
Glu Phe Gly Asn Val Leu Asp Ile Ile Asp Gln Leu Pro Leu Pro Gly
245 250 255
Lys Gln Lys Tyr Ile Gln Asn Leu Asn Ala Ser Glu Glu Leu Leu Phe
260 265 270
Arg Asn Leu Ile Leu Val Asp Leu Pro Thr Tyr Cys Val Asp Ala Ile
275 280 285
Ala Ala Val Gly Gln Asp Val Leu Asp Lys Phe Thr Lys Asp Ile Leu
290 295 300
Glu Ile Ala Glu Gln Ser Gly Gly Ser Lys Arg Thr Ala Asp Gly Ser
305 310 315 320
Glu Phe Glu Pro Lys Lys Lys Arg Lys Val
325 330
<210> 101
<211> 340
<212> PRT
<213> 人工的
<220>
<223> 合成的核酸外切酶
<400> 101
Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys
1 5 10 15
Arg Lys Val Met Ala Leu Leu Asp Leu Lys Gln Phe Tyr Glu Leu Arg
20 25 30
Glu Gly Cys Asp Asp Lys Gly Ile Leu Val Met Asp Gly Asp Trp Leu
35 40 45
Val Phe Gln Ala Met Ser Ala Ala Glu Phe Asp Ala Ser Trp Glu Glu
50 55 60
Glu Ile Trp His Arg Cys Cys Asp His Ala Lys Ala Arg Gln Ile Leu
65 70 75 80
Glu Asp Ser Ile Lys Ser Tyr Glu Thr Arg Lys Lys Ala Trp Ala Gly
85 90 95
Ala Pro Ile Val Leu Ala Phe Thr Asp Ser Val Asn Trp Arg Lys Glu
100 105 110
Leu Val Asp Pro Asn Tyr Lys Ala Asn Arg Lys Ala Val Lys Lys Pro
115 120 125
Val Gly Tyr Phe Glu Phe Leu Asp Ala Leu Phe Glu Arg Glu Glu Phe
130 135 140
Tyr Cys Ile Arg Glu Pro Met Leu Glu Gly Asp Asp Val Met Gly Val
145 150 155 160
Ile Ala Ser Asn Pro Ser Ala Phe Gly Ala Arg Lys Ala Val Ile Ile
165 170 175
Ser Cys Asp Lys Asp Phe Lys Thr Ile Pro Asn Cys Asp Phe Leu Trp
180 185 190
Cys Thr Thr Gly Asn Ile Leu Thr Gln Thr Glu Glu Ser Ala Asp Trp
195 200 205
Trp His Leu Phe Gln Thr Ile Lys Gly Asp Ile Thr Asp Gly Tyr Ser
210 215 220
Gly Ile Ala Gly Trp Gly Asp Thr Ala Glu Asp Phe Leu Asn Asn Pro
225 230 235 240
Phe Ile Thr Glu Pro Lys Thr Ser Val Leu Lys Ser Gly Lys Asn Lys
245 250 255
Gly Gln Glu Val Thr Lys Trp Val Lys Arg Asp Pro Glu Pro His Glu
260 265 270
Thr Leu Trp Asp Cys Ile Lys Ser Ile Gly Ala Lys Ala Gly Met Thr
275 280 285
Glu Glu Asp Ile Ile Lys Gln Gly Gln Met Ala Arg Ile Leu Arg Phe
290 295 300
Asn Glu Tyr Asn Phe Ile Asp Lys Glu Ile Tyr Leu Trp Arg Pro Ser
305 310 315 320
Gly Gly Ser Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Pro Lys Lys
325 330 335
Lys Arg Lys Val
340
<210> 102
<211> 283
<212> DNA
<213> 智人
<400> 102
ctgtgaggat tgagtgagtt gcacgtgtca agtgcttaga gcaggcgtgc tgcacacagc 60
aggcctttgg tcaggttggc tgctgggctg gccctggggc cgtttccctc actcctgctc 120
ggtgaatttg gctcagcagg cacctgcctc agctgctcac ttgagcctct gggtctagaa 180
ccctctgggg accgtttgag gagtgttcag tctccgtgaa cgttccctta gcactctgcc 240
acttattggg tcagctgtta acatcagtac gttaatgttt cct 283
<210> 103
<211> 23
<212> DNA
<213> 人工的
<220>
<223> 合成的间隔子序列
<400> 103
cctcactcct gctcggtgaa ttt 23
<210> 104
<211> 298
<212> DNA
<213> 智人
<400> 104
gagtcaagga acacggataa agacgctggg agattgacat gcatttcgac caatagcatt 60
gcagagaggc gtatcatttc gcggatgttc caatcagtac gcagagagtc gccgtctcca 120
aggtgaaagc ggaagtaggg ccttcgcgca cctcatggaa tcccttctgc agcacctgga 180
tcgcttttcc gagcttctgg cggtctcaag cactacctac gtcagcacct gggaccccgc 240
caccgtgcgc cgggccttgc agtgggcgcg ctacctgcgc cacatccatc ggcgcttt 298
<210> 105
<211> 23
<212> DNA
<213> 人工的
<220>
<223> 合成的间隔子序列
<400> 105
gcggatgttc caatcagtac gca 23
<210> 106
<211> 1367
<212> PRT
<213> 人工序列
<220>
<223> 合成的nCas9
<400> 106
Asp Lys Lys Tyr Ser Ile Gly Leu Ala Ile Gly Thr Asn Ser Val Gly
1 5 10 15
Trp Ala Val Ile Thr Asp Glu Tyr Lys Val Pro Ser Lys Lys Phe Lys
20 25 30
Val Leu Gly Asn Thr Asp Arg His Ser Ile Lys Lys Asn Leu Ile Gly
35 40 45
Ala Leu Leu Phe Asp Ser Gly Glu Thr Ala Glu Ala Thr Arg Leu Lys
50 55 60
Arg Thr Ala Arg Arg Arg Tyr Thr Arg Arg Lys Asn Arg Ile Cys Tyr
65 70 75 80
Leu Gln Glu Ile Phe Ser Asn Glu Met Ala Lys Val Asp Asp Ser Phe
85 90 95
Phe His Arg Leu Glu Glu Ser Phe Leu Val Glu Glu Asp Lys Lys His
100 105 110
Glu Arg His Pro Ile Phe Gly Asn Ile Val Asp Glu Val Ala Tyr His
115 120 125
Glu Lys Tyr Pro Thr Ile Tyr His Leu Arg Lys Lys Leu Val Asp Ser
130 135 140
Thr Asp Lys Ala Asp Leu Arg Leu Ile Tyr Leu Ala Leu Ala His Met
145 150 155 160
Ile Lys Phe Arg Gly His Phe Leu Ile Glu Gly Asp Leu Asn Pro Asp
165 170 175
Asn Ser Asp Val Asp Lys Leu Phe Ile Gln Leu Val Gln Thr Tyr Asn
180 185 190
Gln Leu Phe Glu Glu Asn Pro Ile Asn Ala Ser Gly Val Asp Ala Lys
195 200 205
Ala Ile Leu Ser Ala Arg Leu Ser Lys Ser Arg Arg Leu Glu Asn Leu
210 215 220
Ile Ala Gln Leu Pro Gly Glu Lys Lys Asn Gly Leu Phe Gly Asn Leu
225 230 235 240
Ile Ala Leu Ser Leu Gly Leu Thr Pro Asn Phe Lys Ser Asn Phe Asp
245 250 255
Leu Ala Glu Asp Ala Lys Leu Gln Leu Ser Lys Asp Thr Tyr Asp Asp
260 265 270
Asp Leu Asp Asn Leu Leu Ala Gln Ile Gly Asp Gln Tyr Ala Asp Leu
275 280 285
Phe Leu Ala Ala Lys Asn Leu Ser Asp Ala Ile Leu Leu Ser Asp Ile
290 295 300
Leu Arg Val Asn Thr Glu Ile Thr Lys Ala Pro Leu Ser Ala Ser Met
305 310 315 320
Ile Lys Arg Tyr Asp Glu His His Gln Asp Leu Thr Leu Leu Lys Ala
325 330 335
Leu Val Arg Gln Gln Leu Pro Glu Lys Tyr Lys Glu Ile Phe Phe Asp
340 345 350
Gln Ser Lys Asn Gly Tyr Ala Gly Tyr Ile Asp Gly Gly Ala Ser Gln
355 360 365
Glu Glu Phe Tyr Lys Phe Ile Lys Pro Ile Leu Glu Lys Met Asp Gly
370 375 380
Thr Glu Glu Leu Leu Val Lys Leu Asn Arg Glu Asp Leu Leu Arg Lys
385 390 395 400
Gln Arg Thr Phe Asp Asn Gly Ser Ile Pro His Gln Ile His Leu Gly
405 410 415
Glu Leu His Ala Ile Leu Arg Arg Gln Glu Asp Phe Tyr Pro Phe Leu
420 425 430
Lys Asp Asn Arg Glu Lys Ile Glu Lys Ile Leu Thr Phe Arg Ile Pro
435 440 445
Tyr Tyr Val Gly Pro Leu Ala Arg Gly Asn Ser Arg Phe Ala Trp Met
450 455 460
Thr Arg Lys Ser Glu Glu Thr Ile Thr Pro Trp Asn Phe Glu Glu Val
465 470 475 480
Val Asp Lys Gly Ala Ser Ala Gln Ser Phe Ile Glu Arg Met Thr Asn
485 490 495
Phe Asp Lys Asn Leu Pro Asn Glu Lys Val Leu Pro Lys His Ser Leu
500 505 510
Leu Tyr Glu Tyr Phe Thr Val Tyr Asn Glu Leu Thr Lys Val Lys Tyr
515 520 525
Val Thr Glu Gly Met Arg Lys Pro Ala Phe Leu Ser Gly Glu Gln Lys
530 535 540
Lys Ala Ile Val Asp Leu Leu Phe Lys Thr Asn Arg Lys Val Thr Val
545 550 555 560
Lys Gln Leu Lys Glu Asp Tyr Phe Lys Lys Ile Glu Cys Phe Asp Ser
565 570 575
Val Glu Ile Ser Gly Val Glu Asp Arg Phe Asn Ala Ser Leu Gly Thr
580 585 590
Tyr His Asp Leu Leu Lys Ile Ile Lys Asp Lys Asp Phe Leu Asp Asn
595 600 605
Glu Glu Asn Glu Asp Ile Leu Glu Asp Ile Val Leu Thr Leu Thr Leu
610 615 620
Phe Glu Asp Arg Glu Met Ile Glu Glu Arg Leu Lys Thr Tyr Ala His
625 630 635 640
Leu Phe Asp Asp Lys Val Met Lys Gln Leu Lys Arg Arg Arg Tyr Thr
645 650 655
Gly Trp Gly Arg Leu Ser Arg Lys Leu Ile Asn Gly Ile Arg Asp Lys
660 665 670
Gln Ser Gly Lys Thr Ile Leu Asp Phe Leu Lys Ser Asp Gly Phe Ala
675 680 685
Asn Arg Asn Phe Met Gln Leu Ile His Asp Asp Ser Leu Thr Phe Lys
690 695 700
Glu Asp Ile Gln Lys Ala Gln Val Ser Gly Gln Gly Asp Ser Leu His
705 710 715 720
Glu His Ile Ala Asn Leu Ala Gly Ser Pro Ala Ile Lys Lys Gly Ile
725 730 735
Leu Gln Thr Val Lys Val Val Asp Glu Leu Val Lys Val Met Gly Arg
740 745 750
His Lys Pro Glu Asn Ile Val Ile Glu Met Ala Arg Glu Asn Gln Thr
755 760 765
Thr Gln Lys Gly Gln Lys Asn Ser Arg Glu Arg Met Lys Arg Ile Glu
770 775 780
Glu Gly Ile Lys Glu Leu Gly Ser Gln Ile Leu Lys Glu His Pro Val
785 790 795 800
Glu Asn Thr Gln Leu Gln Asn Glu Lys Leu Tyr Leu Tyr Tyr Leu Gln
805 810 815
Asn Gly Arg Asp Met Tyr Val Asp Gln Glu Leu Asp Ile Asn Arg Leu
820 825 830
Ser Asp Tyr Asp Val Asp His Ile Val Pro Gln Ser Phe Leu Lys Asp
835 840 845
Asp Ser Ile Asp Asn Lys Val Leu Thr Arg Ser Asp Lys Asn Arg Gly
850 855 860
Lys Ser Asp Asn Val Pro Ser Glu Glu Val Val Lys Lys Met Lys Asn
865 870 875 880
Tyr Trp Arg Gln Leu Leu Asn Ala Lys Leu Ile Thr Gln Arg Lys Phe
885 890 895
Asp Asn Leu Thr Lys Ala Glu Arg Gly Gly Leu Ser Glu Leu Asp Lys
900 905 910
Ala Gly Phe Ile Lys Arg Gln Leu Val Glu Thr Arg Gln Ile Thr Lys
915 920 925
His Val Ala Gln Ile Leu Asp Ser Arg Met Asn Thr Lys Tyr Asp Glu
930 935 940
Asn Asp Lys Leu Ile Arg Glu Val Lys Val Ile Thr Leu Lys Ser Lys
945 950 955 960
Leu Val Ser Asp Phe Arg Lys Asp Phe Gln Phe Tyr Lys Val Arg Glu
965 970 975
Ile Asn Asn Tyr His His Ala His Asp Ala Tyr Leu Asn Ala Val Val
980 985 990
Gly Thr Ala Leu Ile Lys Lys Tyr Pro Lys Leu Glu Ser Glu Phe Val
995 1000 1005
Tyr Gly Asp Tyr Lys Val Tyr Asp Val Arg Lys Met Ile Ala Lys
1010 1015 1020
Ser Glu Gln Glu Ile Gly Lys Ala Thr Ala Lys Tyr Phe Phe Tyr
1025 1030 1035
Ser Asn Ile Met Asn Phe Phe Lys Thr Glu Ile Thr Leu Ala Asn
1040 1045 1050
Gly Glu Ile Arg Lys Arg Pro Leu Ile Glu Thr Asn Gly Glu Thr
1055 1060 1065
Gly Glu Ile Val Trp Asp Lys Gly Arg Asp Phe Ala Thr Val Arg
1070 1075 1080
Lys Val Leu Ser Met Pro Gln Val Asn Ile Val Lys Lys Thr Glu
1085 1090 1095
Val Gln Thr Gly Gly Phe Ser Lys Glu Ser Ile Leu Pro Lys Arg
1100 1105 1110
Asn Ser Asp Lys Leu Ile Ala Arg Lys Lys Asp Trp Asp Pro Lys
1115 1120 1125
Lys Tyr Gly Gly Phe Asp Ser Pro Thr Val Ala Tyr Ser Val Leu
1130 1135 1140
Val Val Ala Lys Val Glu Lys Gly Lys Ser Lys Lys Leu Lys Ser
1145 1150 1155
Val Lys Glu Leu Leu Gly Ile Thr Ile Met Glu Arg Ser Ser Phe
1160 1165 1170
Glu Lys Asn Pro Ile Asp Phe Leu Glu Ala Lys Gly Tyr Lys Glu
1175 1180 1185
Val Lys Lys Asp Leu Ile Ile Lys Leu Pro Lys Tyr Ser Leu Phe
1190 1195 1200
Glu Leu Glu Asn Gly Arg Lys Arg Met Leu Ala Ser Ala Gly Glu
1205 1210 1215
Leu Gln Lys Gly Asn Glu Leu Ala Leu Pro Ser Lys Tyr Val Asn
1220 1225 1230
Phe Leu Tyr Leu Ala Ser His Tyr Glu Lys Leu Lys Gly Ser Pro
1235 1240 1245
Glu Asp Asn Glu Gln Lys Gln Leu Phe Val Glu Gln His Lys His
1250 1255 1260
Tyr Leu Asp Glu Ile Ile Glu Gln Ile Ser Glu Phe Ser Lys Arg
1265 1270 1275
Val Ile Leu Ala Asp Ala Asn Leu Asp Lys Val Leu Ser Ala Tyr
1280 1285 1290
Asn Lys His Arg Asp Lys Pro Ile Arg Glu Gln Ala Glu Asn Ile
1295 1300 1305
Ile His Leu Phe Thr Leu Thr Asn Leu Gly Ala Pro Ala Ala Phe
1310 1315 1320
Lys Tyr Phe Asp Thr Thr Ile Asp Arg Lys Arg Tyr Thr Ser Thr
1325 1330 1335
Lys Glu Val Leu Asp Ala Thr Leu Ile His Gln Ser Ile Thr Gly
1340 1345 1350
Leu Tyr Glu Thr Arg Ile Asp Leu Ser Gln Leu Gly Gly Asp
1355 1360 1365
<210> 107
<211> 1367
<212> PRT
<213> 人工序列
<220>
<223> 合成的enCas9
<400> 107
Asp Lys Lys Tyr Ser Ile Gly Leu Ala Ile Gly Thr Asn Ser Val Gly
1 5 10 15
Trp Ala Val Ile Thr Asp Glu Tyr Lys Val Pro Ser Lys Lys Phe Lys
20 25 30
Val Leu Gly Asn Thr Asp Arg His Ser Ile Lys Lys Asn Leu Ile Gly
35 40 45
Ala Leu Leu Phe Asp Ser Gly Glu Thr Ala Glu Ala Thr Arg Leu Lys
50 55 60
Arg Thr Ala Arg Arg Arg Tyr Thr Arg Arg Lys Asn Arg Ile Cys Tyr
65 70 75 80
Leu Gln Glu Ile Phe Ser Asn Glu Met Ala Lys Val Asp Asp Ser Phe
85 90 95
Phe His Arg Leu Glu Glu Ser Phe Leu Val Glu Glu Asp Lys Lys His
100 105 110
Glu Arg His Pro Ile Phe Gly Asn Ile Val Asp Glu Val Ala Tyr His
115 120 125
Glu Lys Tyr Pro Thr Ile Tyr His Leu Arg Lys Lys Leu Val Asp Ser
130 135 140
Thr Asp Lys Ala Asp Leu Arg Leu Ile Tyr Leu Ala Leu Ala His Met
145 150 155 160
Ile Lys Phe Arg Gly His Phe Leu Ile Glu Gly Asp Leu Asn Pro Asp
165 170 175
Asn Ser Asp Val Asp Lys Leu Phe Ile Gln Leu Val Gln Thr Tyr Asn
180 185 190
Gln Leu Phe Glu Glu Asn Pro Ile Asn Ala Ser Gly Val Asp Ala Lys
195 200 205
Ala Ile Leu Ser Ala Arg Leu Ser Lys Ser Arg Arg Leu Glu Asn Leu
210 215 220
Ile Ala Gln Leu Pro Gly Glu Lys Lys Asn Gly Leu Phe Gly Asn Leu
225 230 235 240
Ile Ala Leu Ser Leu Gly Leu Thr Pro Asn Phe Lys Ser Asn Phe Asp
245 250 255
Leu Ala Glu Asp Ala Lys Leu Gln Leu Ser Lys Asp Thr Tyr Asp Asp
260 265 270
Asp Leu Asp Asn Leu Leu Ala Gln Ile Gly Asp Gln Tyr Ala Asp Leu
275 280 285
Phe Leu Ala Ala Lys Asn Leu Ser Asp Ala Ile Leu Leu Ser Asp Ile
290 295 300
Leu Arg Val Asn Thr Glu Ile Thr Lys Ala Pro Leu Ser Ala Ser Met
305 310 315 320
Ile Lys Arg Tyr Asp Glu His His Gln Asp Leu Thr Leu Leu Lys Ala
325 330 335
Leu Val Arg Gln Gln Leu Pro Glu Lys Tyr Lys Glu Ile Phe Phe Asp
340 345 350
Gln Ser Lys Asn Gly Tyr Ala Gly Tyr Ile Asp Gly Gly Ala Ser Gln
355 360 365
Glu Glu Phe Tyr Lys Phe Ile Lys Pro Ile Leu Glu Lys Met Asp Gly
370 375 380
Thr Glu Glu Leu Leu Val Lys Leu Asn Arg Glu Asp Leu Leu Arg Lys
385 390 395 400
Gln Arg Thr Phe Asp Asn Gly Ser Ile Pro His Gln Ile His Leu Gly
405 410 415
Glu Leu His Ala Ile Leu Arg Arg Gln Glu Asp Phe Tyr Pro Phe Leu
420 425 430
Lys Asp Asn Arg Glu Lys Ile Glu Lys Ile Leu Thr Phe Arg Ile Pro
435 440 445
Tyr Tyr Val Gly Pro Leu Ala Arg Gly Asn Ser Arg Phe Ala Trp Met
450 455 460
Thr Arg Lys Ser Glu Glu Thr Ile Thr Pro Trp Asn Phe Glu Glu Val
465 470 475 480
Val Asp Lys Gly Ala Ser Ala Gln Ser Phe Ile Glu Arg Met Thr Asn
485 490 495
Phe Asp Lys Asn Leu Pro Asn Glu Lys Val Leu Pro Lys His Ser Leu
500 505 510
Leu Tyr Glu Tyr Phe Thr Val Tyr Asn Glu Leu Thr Lys Val Lys Tyr
515 520 525
Val Thr Glu Gly Met Arg Lys Pro Ala Phe Leu Ser Gly Glu Gln Lys
530 535 540
Lys Ala Ile Val Asp Leu Leu Phe Lys Thr Asn Arg Lys Val Thr Val
545 550 555 560
Lys Gln Leu Lys Glu Asp Tyr Phe Lys Lys Ile Glu Cys Phe Asp Ser
565 570 575
Val Glu Ile Ser Gly Val Glu Asp Arg Phe Asn Ala Ser Leu Gly Thr
580 585 590
Tyr His Asp Leu Leu Lys Ile Ile Lys Asp Lys Asp Phe Leu Asp Asn
595 600 605
Glu Glu Asn Glu Asp Ile Leu Glu Asp Ile Val Leu Thr Leu Thr Leu
610 615 620
Phe Glu Asp Arg Glu Met Ile Glu Glu Arg Leu Lys Thr Tyr Ala His
625 630 635 640
Leu Phe Asp Asp Lys Val Met Lys Gln Leu Lys Arg Arg Arg Tyr Thr
645 650 655
Gly Trp Gly Arg Leu Ser Arg Lys Leu Ile Asn Gly Ile Arg Asp Lys
660 665 670
Gln Ser Gly Lys Thr Ile Leu Asp Phe Leu Lys Ser Asp Gly Phe Ala
675 680 685
Asn Arg Asn Phe Met Gln Leu Ile His Asp Asp Ser Leu Thr Phe Lys
690 695 700
Glu Asp Ile Gln Lys Ala Gln Val Ser Gly Gln Gly Asp Ser Leu His
705 710 715 720
Glu His Ile Ala Asn Leu Ala Gly Ser Pro Ala Ile Lys Lys Gly Ile
725 730 735
Leu Gln Thr Val Lys Val Val Asp Glu Leu Val Lys Val Met Gly Arg
740 745 750
His Lys Pro Glu Asn Ile Val Ile Glu Met Ala Arg Glu Asn Gln Thr
755 760 765
Thr Gln Lys Gly Gln Lys Asn Ser Arg Glu Arg Met Lys Arg Ile Glu
770 775 780
Glu Gly Ile Lys Glu Leu Gly Ser Gln Ile Leu Lys Glu His Pro Val
785 790 795 800
Glu Asn Thr Gln Leu Gln Asn Glu Lys Leu Tyr Leu Tyr Tyr Leu Gln
805 810 815
Asn Gly Arg Asp Met Tyr Val Asp Gln Glu Leu Asp Ile Asn Arg Leu
820 825 830
Ser Asp Tyr Asp Val Asp His Ile Val Pro Gln Ser Phe Leu Ala Asp
835 840 845
Asp Ser Ile Asp Asn Lys Val Leu Thr Arg Ser Asp Lys Asn Arg Gly
850 855 860
Lys Ser Asp Asn Val Pro Ser Glu Glu Val Val Lys Lys Met Lys Asn
865 870 875 880
Tyr Trp Arg Gln Leu Leu Asn Ala Lys Leu Ile Thr Gln Arg Lys Phe
885 890 895
Asp Asn Leu Thr Lys Ala Glu Arg Gly Gly Leu Ser Glu Leu Asp Lys
900 905 910
Ala Gly Phe Ile Lys Arg Gln Leu Val Glu Thr Arg Gln Ile Thr Lys
915 920 925
His Val Ala Gln Ile Leu Asp Ser Arg Met Asn Thr Lys Tyr Asp Glu
930 935 940
Asn Asp Lys Leu Ile Arg Glu Val Lys Val Ile Thr Leu Lys Ser Lys
945 950 955 960
Leu Val Ser Asp Phe Arg Lys Asp Phe Gln Phe Tyr Lys Val Arg Glu
965 970 975
Ile Asn Asn Tyr His His Ala His Asp Ala Tyr Leu Asn Ala Val Val
980 985 990
Gly Thr Ala Leu Ile Lys Lys Tyr Pro Ala Leu Glu Ser Glu Phe Val
995 1000 1005
Tyr Gly Asp Tyr Lys Val Tyr Asp Val Arg Lys Met Ile Ala Lys
1010 1015 1020
Ser Glu Gln Glu Ile Gly Lys Ala Thr Ala Lys Tyr Phe Phe Tyr
1025 1030 1035
Ser Asn Ile Met Asn Phe Phe Lys Thr Glu Ile Thr Leu Ala Asn
1040 1045 1050
Gly Glu Ile Arg Lys Ala Pro Leu Ile Glu Thr Asn Gly Glu Thr
1055 1060 1065
Gly Glu Ile Val Trp Asp Lys Gly Arg Asp Phe Ala Thr Val Arg
1070 1075 1080
Lys Val Leu Ser Met Pro Gln Val Asn Ile Val Lys Lys Thr Glu
1085 1090 1095
Val Gln Thr Gly Gly Phe Ser Lys Glu Ser Ile Leu Pro Lys Arg
1100 1105 1110
Asn Ser Asp Lys Leu Ile Ala Arg Lys Lys Asp Trp Asp Pro Lys
1115 1120 1125
Lys Tyr Gly Gly Phe Asp Ser Pro Thr Val Ala Tyr Ser Val Leu
1130 1135 1140
Val Val Ala Lys Val Glu Lys Gly Lys Ser Lys Lys Leu Lys Ser
1145 1150 1155
Val Lys Glu Leu Leu Gly Ile Thr Ile Met Glu Arg Ser Ser Phe
1160 1165 1170
Glu Lys Asn Pro Ile Asp Phe Leu Glu Ala Lys Gly Tyr Lys Glu
1175 1180 1185
Val Lys Lys Asp Leu Ile Ile Lys Leu Pro Lys Tyr Ser Leu Phe
1190 1195 1200
Glu Leu Glu Asn Gly Arg Lys Arg Met Leu Ala Ser Ala Gly Glu
1205 1210 1215
Leu Gln Lys Gly Asn Glu Leu Ala Leu Pro Ser Lys Tyr Val Asn
1220 1225 1230
Phe Leu Tyr Leu Ala Ser His Tyr Glu Lys Leu Lys Gly Ser Pro
1235 1240 1245
Glu Asp Asn Glu Gln Lys Gln Leu Phe Val Glu Gln His Lys His
1250 1255 1260
Tyr Leu Asp Glu Ile Ile Glu Gln Ile Ser Glu Phe Ser Lys Arg
1265 1270 1275
Val Ile Leu Ala Asp Ala Asn Leu Asp Lys Val Leu Ser Ala Tyr
1280 1285 1290
Asn Lys His Arg Asp Lys Pro Ile Arg Glu Gln Ala Glu Asn Ile
1295 1300 1305
Ile His Leu Phe Thr Leu Thr Asn Leu Gly Ala Pro Ala Ala Phe
1310 1315 1320
Lys Tyr Phe Asp Thr Thr Ile Asp Arg Lys Arg Tyr Thr Ser Thr
1325 1330 1335
Lys Glu Val Leu Asp Ala Thr Leu Ile His Gln Ser Ile Thr Gly
1340 1345 1350
Leu Tyr Glu Thr Arg Ile Asp Leu Ser Gln Leu Gly Gly Asp
1355 1360 1365
<210> 108
<211> 4101
<212> DNA
<213> 人工序列
<220>
<223> 合成的Cas9
<400> 108
gacaagaagt acagcatcgg gctggcgatc gggaccaact ccgtcggctg ggctgtgatt 60
accgacgagt acaaggtgcc atccaagaag ttcaaggtcc tcggcaacac tgaccggcac 120
agcattaaga agaacctgat tggggcgctg ctgttcgatt cgggggagac tgcggaggcg 180
accaggctga agcggactgc gcgccggagg tacaccagga ggaagaatcg gatctgctac 240
ctccaggaga ttttctcgaa tgagatggcc aaggtggacg attccttctt ccatcgcctg 300
gaggagtcgt tcctcgttga ggaggacaag aagcatgaga ggcatcccat tttcgggaat 360
atcgttgacg aggtggctta ccatgagaag tacccgacca tctaccatct gcggaagaag 420
ctcgtcgatt cgaccgataa ggccgacctg cggctgatct acctggccct cgcgcacatg 480
attaagttcc ggggccattt cctcatcgag ggcgacctca acccggacaa ctcggacgtg 540
gataagctct tcattcagct cgtgcagaca tacaaccagc tcttcgagga gaatcccatt 600
aacgcctcgg gggtcgacgc taaggctatt ctctcggctc ggctgtcgaa gtcgcgccgg 660
ctggagaatc tcattgccca gctcccaggc gagaagaaga acggcctctt cggcaacctg 720
attgccctgt cgctggggct cacaccgaat ttcaagtcga acttcgacct cgccgaggac 780
gctaagctcc agctcagcaa ggatacttac gatgatgacc tcgataacct gctcgcccag 840
attggggatc agtacgcgga tctgttcctc gcggccaaga atctcagcga tgctattctc 900
ctgtcggaca ttctccgcgt caacacagag attactaagg ccccactgtc ggcgagcatg 960
attaagaggt acgatgagca tcatcaggac ctgacactgc tcaaggcgct ggtccggcag 1020
cagctccccg agaagtacaa ggagattttc ttcgatcagt caaagaatgg gtacgcgggc 1080
tacattgatg gcggcgcgtc ccaggaggag ttctacaagt tcattaagcc catcctggag 1140
aagatggacg ggaccgagga gctgctggtg aagctcaatc gggaggacct gctccggaag 1200
cagcgcacat tcgacaatgg ctcgattcct caccagattc acctgggcga gctgcacgcc 1260
attctccgca ggcaggagga cttctacccg ttcctcaagg acaaccgcga gaagatcgag 1320
aagatcctga ccttccggat tccatactac gtggggccgc tcgcgcgggg gaactcccgg 1380
ttcgcgtgga tgactcgcaa gtccgaagaa acgattacac cgtggaattt cgaggaggtc 1440
gtcgacaagg gcgctagtgc gcagtcattc attgagagga tgaccaattt cgataagaac 1500
ctgcctaacg agaaggtgct gccgaagcat tcgctgctct acgagtactt caccgtttac 1560
aatgagctga ccaaggtgaa gtatgtgact gagggcatga ggaagccagc gttcctgagc 1620
ggcgagcaga agaaggctat cgtggacctg ctcttcaaga ctaaccggaa ggtgactgtg 1680
aagcagctca aggaggacta cttcaagaag attgagtgct tcgattccgt tgagattagc 1740
ggggtggagg atcggttcaa tgcttcgctc gggacatacc acgatctcct gaagatcatt 1800
aaggataagg acttcctcga caacgaggag aacgaggaca ttctcgaaga tattgtcctg 1860
accctcaccc tcttcgagga tcgggagatg atcgaggaga ggctcaagac atacgctcat 1920
ctgttcgatg ataaggtcat gaagcagctg aagcgcaggc ggtacacagg gtgggggcgg 1980
ctgagccgga agctgatcaa cgggattcgg gataagcagt ccgggaagac aattctcgac 2040
ttcctcaagt ccgacgggtt cgctaaccgg aacttcatgc agctcattca tgatgactcg 2100
ctgacattca aggaggatat tcagaaggcg caggtttcgg ggcagggcga ctcgctccac 2160
gagcatattg cgaatctggc gggctccccc gcgattaaga agggcattct gcaaaccgtc 2220
aaggtggttg atgagctggt caaggtcatg gggcggcata agccagagaa tattgtcatc 2280
gagatggcgc gggagaatca gaccacacag aaggggcaga agaactcacg ggagcggatg 2340
aagcgcatcg aggagggcat caaggagctg gggtcgcaga tcctgaagga gcatcccgtg 2400
gagaacactc agctgcaaaa tgagaagctg tacctctact acctccagaa cgggagggac 2460
atgtatgtgg atcaggagct ggatattaat aggctgagcg attacgatgt cgaccacatt 2520
gtcccacagt cgttcctgaa ggacgacagc attgacaaca aggtgctgac ccgctcggat 2580
aagaacaggg gcaagagcga taatgttcca agcgaggagg ttgtgaagaa gatgaagaac 2640
tactggcggc agctcctgaa cgcgaagctc atcacacagc ggaagttcga caacctcacc 2700
aaggctgagc gcgggggcct gagcgagctg gacaaggcgg ggttcattaa gaggcagctg 2760
gtcgagacac ggcagattac aaagcatgtt gcgcagattc tcgattcccg gatgaacacc 2820
aagtacgatg agaacgataa gctgattcgg gaggtcaagg taattaccct gaagtccaag 2880
ctggtgtccg acttcaggaa ggacttccag ttctacaagg ttcgggagat caacaactac 2940
caccacgcgc atgatgccta cctcaacgcg gtcgtgggga ccgctctcat caagaagtac 3000
ccaaagctgg agtcagagtt cgtctacggg gattacaagg tttacgacgt gcggaagatg 3060
atcgctaaga gcgagcagga gattggcaag gctaccgcta agtacttctt ctactccaac 3120
atcatgaact tcttcaagac agagattacc ctcgcgaatg gcgagatccg gaagaggccc 3180
ctcatcgaga caaatgggga gacaggggag attgtctggg ataaggggcg ggatttcgcg 3240
accgtccgga aggtcctgtc gatgccccag gttaatattg tcaagaagac tgaggtccag 3300
actggcggct tctcaaagga gtcgattctc ccaaagagga actccgataa gctcattgct 3360
cggaagaagg attgggaccc caagaagtac gggggattcg actcccccac tgttgcttac 3420
tctgttctgg ttgttgctaa ggtggagaag gggaagtcga agaagctgaa gagcgtgaag 3480
gagctgctcg ggattacaat tatggagagg tcatccttcg agaagaatcc catcgacttc 3540
ctggaggcca agggctacaa ggaggtgaag aaggacctga ttattaagct gcccaagtac 3600
tcgctcttcg agctggagaa tgggcggaag cggatgctgg cgtccgcggg ggagctgcaa 3660
aaggggaacg agctggcgct cccctccaag tatgtgaact tcctctacct ggcgtcgcac 3720
tacgagaagc tgaaggggtc cccagaggat aatgagcaga agcagctctt cgtcgagcag 3780
cataagcact acctggacga gattatcgag cagattagcg agttctcgaa gcgggtcatc 3840
ctcgcggatg cgaacctgga taaggtgctc agcgcctaca ataagcaccg ggacaagccg 3900
attcgggagc aggcggagaa tattattcac ctcttcacac tcaccaacct cggggcacca 3960
gctgcgttca agtacttcga cactactatc gaccggaagc ggtacacctc gacgaaggag 4020
gtgctcgacg ccaccctcat tcaccagtcg atcacaggcc tgtacgagac acggattgac 4080
ctgtcccagc tcgggggcga c 4101
<210> 109
<211> 4101
<212> DNA
<213> 人工序列
<220>
<223> 合成的Cas9
<400> 109
gacaagaagt actccattgg cctggcgatt gggacaaact cggtggggtg ggccgtgatt 60
acggatgagt acaaggttcc aagcaagaag ttcaaggtcc tcgggaacac agatcggcat 120
tcgattaaga agaatctcat tggggcgctc ctcttcgact cgggggagac agcggaggct 180
accaggctca agcggacagc caggcggcgg tacacaaggc ggaagaatcg catctgctac 240
ctccaggaga ttttctcgaa tgagatggcg aaggtggacg acagcttctt ccatcggctg 300
gaggagtcct tcctggtgga ggaggataag aagcacgaga ggcatccaat tttcgggaac 360
atcgtggacg aggttgcgta ccatgagaag taccctacaa tctaccatct gcggaagaag 420
ctggttgact ccacagacaa ggcggacctg aggctgatct acctcgctct ggcccacatg 480
attaagttcc gcgggcattt cctgatcgag ggggacctga atcccgacaa ttcggatgtg 540
gacaagctct tcatccagct ggtgcagacc tacaaccagc tgttcgagga gaatcccatc 600
aatgcgtcgg gcgttgacgc taaggccatt ctgtccgcta ggctgtcgaa gagcaggagg 660
ctggagaacc tgatcgccca gctgccaggc gagaagaaga atgggctctt cgggaatctg 720
attgcgctct ccctggggct gacaccgaac ttcaagagca atttcgatct ggctgaggac 780
gcgaagctcc agctctcgaa ggacacttac gacgatgacc tcgataacct cctcgcgcag 840
atcggggacc agtacgctga tctcttcctc gccgctaaga acctctcgga tgctatcctg 900
ctctccgaca ttctccgggt taataccgag attacaaagg ccccactgtc ggcgtccatg 960
atcaagcggt acgatgagca tcatcaggat ctcaccctgc tcaaggccct cgtgcggcag 1020
cagctgcccg agaagtacaa ggagattttc ttcgaccaga gcaagaatgg gtacgctggc 1080
tacattgacg gcggggcctc acaggaggag ttctacaagt tcatcaagcc aatcctggag 1140
aagatggatg ggacagagga gctgctggtg aagctcaacc gggaggatct gctcaggaag 1200
cagcggacgt tcgacaacgg gtcgattccc catcagatcc acctggggga gctgcacgcg 1260
atcctgcgcc ggcaggagga tttctaccct ttcctgaagg ataatcggga gaagatcgag 1320
aagattctca ccttccggat tccctactac gtcgggccac tcgcgcgggg caatagcagg 1380
ttcgcctgga tgacacggaa gagcgaggag acaatcaccc cctggaactt cgaggaggtt 1440
gtcgacaagg gggcgtccgc ccagtcattc attgagcgga tgaccaattt cgacaagaat 1500
ctgccaaatg agaaggttct cccaaagcat agcctcctct acgagtactt cactgtttac 1560
aacgagctga ccaaggtgaa gtatgtgacc gagggcatgc ggaagcccgc gttcctgtcc 1620
ggcgagcaga agaaggccat tgtggacctc ctgttcaaga ccaatcgcaa ggtcacagtc 1680
aagcagctca aggaggatta cttcaagaag atcgagtgct tcgactcggt tgagattagc 1740
ggggtggagg atcggttcaa cgcgagcctc ggcacttacc acgacctcct gaagatcatc 1800
aaggataagg acttcctcga caacgaggag aacgaggata ttctggagga catcgtgctc 1860
accctgacgc tgttcgagga tcgggagatg atcgaggagc gcctgaagac ctacgctcat 1920
ctcttcgatg ataaggtcat gaagcagctg aagaggaggc ggtacaccgg gtggggccgc 1980
ctgagcagga agctcattaa cgggatcagg gacaagcaga gcggcaagac catcctggac 2040
ttcctcaaga gcgatggctt cgccaaccgg aatttcatgc agctcatcca cgacgactcc 2100
ctcaccttca aggaggacat tcagaaggct caggtcagcg gccagggcga ctcgctgcat 2160
gagcacatcg ctaacctggc gggcagccca gccatcaaga agggcatcct ccagacagtg 2220
aaggtcgtgg atgagctggt gaaggtcatg ggccggcata agcccgagaa tattgtgatt 2280
gagatggcgc gggagaatca gaccactcag aagggccaga agaactcgcg ggagcgcatg 2340
aagaggatcg aggaggggat taaggagctg ggcagccaga ttctcaagga gcaccccgtg 2400
gagaataccc agctccagaa cgagaagctg tacctctact acctccagaa tgggcgggac 2460
atgtatgttg atcaggagct ggacatcaat cgcctctcgg attacgacgt ggaccacatc 2520
gtgccccaga gcttcctgaa ggatgatagc atcgacaata aggtcctgac ccgctccgac 2580
aagaatcgcg gcaagagcga caacgtgccg agcgaggagg tcgtgaagaa gatgaagaac 2640
tactggcggc agctgctgaa cgcgaagctc attacacagc ggaagttcga taacctgacg 2700
aaggcggaga ggggcggcct ctccgagctg gacaaggcgg gcttcattaa gaggcagctc 2760
gtggagactc gccagatcac caagcacgtg gctcagatcc tcgatagccg gatgaatacg 2820
aagtacgatg agaatgacaa gctcatccgg gaggtgaagg taatcaccct gaagtcaaag 2880
ctcgttagcg atttccggaa ggacttccag ttctacaagg tgcgggagat taacaactac 2940
catcatgcgc acgatgcgta cctcaatgcg gtggtgggca cagccctgat taagaagtac 3000
cccaagctgg agagcgagtt cgtctacggg gactacaagg tgtacgatgt tcggaagatg 3060
atcgccaaga gcgagcagga gattgggaag gccaccgcta agtacttctt ctactcgaat 3120
attatgaatt tcttcaagac cgagatcaca ctcgctaatg gggagattcg gaagcggccc 3180
ctcatcgaga ctaacgggga gactggcgag attgtgtggg acaaggggcg cgacttcgct 3240
accgtgcgca aggtcctctc gatgccccag gttaatattg ttaagaagac agaggtgcag 3300
acgggcgggt tctccaagga gtctatcctg ccgaagcgga actcggacaa gctgatcgcc 3360
cgcaagaagg attgggaccc caagaagtac gggggattcg atagcccaac cgtggcttac 3420
agcgtcctgg tggtcgccaa ggttgagaag gggaagtcga agaagctcaa gagcgttaag 3480
gagctgctgg gcatcaccat catggagcgg tccagcttcg agaagaatcc tatcgacttc 3540
ctggaggcta aggggtacaa ggaggtcaag aaggacctga tcattaagct gcccaagtac 3600
tctctgttcg agctggagaa cgggaggaag cggatgctgg cgtctgctgg cgagctacag 3660
aagggcaatg agctggcgct cccctcgaag tatgtcaact tcctctacct ggcttcccat 3720
tacgagaagc tgaagggctc gcccgaggat aatgagcaga agcagctctt cgtggagcag 3780
cacaagcact acctcgacga gatcattgag cagatttcgg agttctcgaa gcgggtcatt 3840
ctcgcggacg cgaacctcga caaggtcctc tcggcgtaca acaagcaccg ggacaagccc 3900
atccgggagc aggccgagaa cattatccac ctcttcacac tgaccaacct cggcgctccc 3960
gccgcgttca agtacttcga caccaccatt gaccgcaaga gatacacatc caccaaggag 4020
gtgctggacg cgaccctcat ccaccagagc atcacaggcc tctacgagac acggatcgac 4080
ctctcgcagc tcgggggcga t 4101
<210> 110
<211> 4092
<212> DNA
<213> 人工序列
<220>
<223> 合成的Cas9
<400> 110
gacaagaagt actcgatcgg cctggcgatt ggcacaaaca gcgtggggtg ggctgtgatc 60
actgatgagt acaaggtgcc atcgaagaag ttcaaggtgc tggggaatac agaccggcat 120
tcgatcaaga agaatctcat tggcgctctc ctcttcgatt ccggcgagac tgctgaggcg 180
acccgcctga agcgcaccgc ccggcggcgc tacactcggc ggaagaatag gatttgctac 240
ctccaggaga ttttctcgaa tgagatggcc aaggtggatg acagcttctt ccaccgcctg 300
gaggagtcgt tcctggtcga ggaggacaag aagcatgagc ggcaccctat cttcgggaat 360
atcgttgatg aggtcgccta ccacgagaag taccccacta tctaccatct ccgcaagaag 420
ctcgtggaca gcacagataa ggccgacctc cgcctgatct acctcgccct cgcgcacatg 480
attaagttcc gggggcactt cctcattgag ggggatctga atcccgataa ctccgacgtg 540
gacaagctgt tcatccagct ggtgcagaca tacaaccagc tgttcgagga gaatcccatc 600
aacgcgagcg gcgtggacgc taaggccatt ctgtcggcta ggctctcgaa gtcgaggcgg 660
ctggagaacc tgattgcgca gctccccggc gagaagaaga acgggctgtt cgggaatctc 720
atcgccctct ccctcggcct cacaccaaac ttcaagagca atttcgacct ggctgaggac 780
gctaagctgc aactctcaaa ggatacatac gatgacgacc tggacaatct cctggctcag 840
atcggcgacc agtacgctga cctgttcctc gcggccaaga atctgtcgga cgcgattctc 900
ctcagcgaca tcctgcgcgt caataccgag attacgaagg ctccactgtc tgcgtcaatg 960
attaagcggt acgatgagca tcaccaggat ctgaccctcc tgaaggcgct cgtgcggcag 1020
cagctgcccg agaagtacaa ggagattttc ttcgatcaga gcaagaatgg ctacgccggc 1080
tacatcgacg ggggcgcgag ccaggaggag ttctacaagt tcatcaagcc catcctggag 1140
aagatggacg gcaccgagga gctactcgtg aagctcaatc gggaggatct cctccggaag 1200
cagcggacat tcgataacgg gtctatccca caccagatcc acctcggcga gctgcatgcg 1260
attctgcggc ggcaggagga tttctaccct ttcctgaagg acaaccggga gaagatcgag 1320
aagatcctca cattccggat tccatactac gtcggccccc tggcgagggg caatagccgg 1380
ttcgcgtgga tgacaaggaa gtccgaggag actattaccc cgtggaattt cgaggaggtg 1440
gttgacaagg gcgcttccgc gcagagcttc attgagcgga tgacaaactt cgacaagaat 1500
ctccccaacg agaaggtcct gccgaagcat agcctcctgt acgagtactt caccgtctac 1560
aatgagctaa ctaaggtcaa gtatgtgaca gagggcatga ggaagccagc cttcctctca 1620
ggcgagcaga agaaggccat tgtggacctc ctgttcaaga caaaccgcaa ggtgacagtg 1680
aagcagctga aggaggatta cttcaagaag attgagtgct tcgactcagt ggagatttca 1740
ggcgtggagg atcggttcaa cgcgagcctg gggacttacc acgacctgct gaagattatt 1800
aaggacaagg acttcctgga taacgaggag aatgaggaca tcctggagga tattgtgctc 1860
accctcaccc tgttcgagga cagggagatg attgaggaga ggctcaagac ctacgcgcac 1920
ctgttcgatg acaaggtcat gaagcagctg aagaggcggc gctacactgg gtggggccgc 1980
ctgtcgcgga agctgatcaa cggcattcgg gataagcagt ccgggaagac cattctggat 2040
ttcctgaagt cggacggctt cgccaacagg aatttcatgc agctgatcca cgacgactcc 2100
ctcaccttca aggaggacat tcagaaggcc caggttagcg gccaggggga ctcactccac 2160
gagcatattg ccaatctggc cggctctcca gctatcaaga agggcatcct gcaaacagtt 2220
aaggttgttg acgagctggt taaggtcatg gggcggcata agcccgagaa cattgtcatc 2280
gagatggctc gggagaacca gacaactcag aagggccaga agaactccag ggagcgcatg 2340
aagcggattg aggagggcat taaggagctg gggtcccaga tcctcaagga gcaccctgtc 2400
gagaacactc agctgcaaaa cgagaagctc tacctgtact acctccagaa cgggcgggat 2460
atgtatgtgg atcaggagct ggacatcaac aggctctccg actacgacgt ggatcacatt 2520
gtcccacagt ctttcctcaa ggatgattcc atcgacaaca aggtgctgac gcgcagcgac 2580
aagaataggg ggaagtcgga caacgttccg agcgaggagg tcgtgaagaa gatgaagaat 2640
tactggaggc agctcctgaa tgcgaagctg atcactcaga ggaagttcga caatctgaca 2700
aaggcggaga ggggcgggct ctcggagctg gataaggcgg gcttcatcaa gcggcagctc 2760
gttgaaaccc ggcagatcac caagcatgtc gcccagatcc tcgatagccg catgaacacc 2820
aagtacgatg agaacgacaa gctcattcgg gaggttaagg tcattacgct gaagtccaag 2880
ctcgtcagcg acttcaggaa ggatttccag ttctacaagg ttcgggagat taacaactac 2940
caccacgcgc atgatgcgta cctgaacgct gttgtcggca ctgctctcat caagaagtac 3000
ccaaagctgg agtccgagtt cgtctacggg gactacaagg tctacgatgt ccggaagatg 3060
atcgccaagt cggagcagga gatcgggaag gctactgcga agtacttctt ctacagcaac 3120
attatgaatt tcttcaagac ggagattacg ctggcgaacg gggagattag gaagaggccc 3180
ctcattgaga ctaatgggga gacaggcgag attgtttggg acaagggccg cgacttcgcg 3240
actgtgcgga aggtcctgtc catgccacag gtgaatattg ttaagaagac agaggtgcag 3300
actgggggct tctcgaagga gagcattctc ccaaagcgga acagcgataa gctcatcgcg 3360
cgcaagaagg attgggaccc taagaagtac ggcggcttcg attctcccac tgtggcctac 3420
tccgttctcg tggttgccaa ggttgagaag gggaagtcga agaagctgaa gtcggtcaag 3480
gagctgctcg ggattacaat catggagcgg agcagcttcg agaagaaccc tattgatttc 3540
ctggaggcca agggctacaa ggaggttaag aaggatctca ttatcaagct ccctaagtac 3600
tctctgttcg agctggagaa tggccggaag aggatgctgg cctcggctgg cgagctacag 3660
aaggggaatg agctggccct cccgtcgaag tatgtgaatt tcctgtacct cgcgtcgcac 3720
tacgagaagc tcaagggcag cccggaggat aatgagcaga agcagctctt cgtggagcag 3780
cataagcact acctggacga gatcattgag cagatcagcg agttctcgaa gcgggttatt 3840
ctggctgatg ctaacctgga caaggttctg agcgcctaca ataagcatcg cgacaagccg 3900
attcgcgagc aggcggagaa tattatccac ctgttcaccc tcactaacct cggggctccc 3960
gcggccttca agtacttcga taccacaata gataggaagc ggtacacctc gacgaaggag 4020
gtcctcgacg ccacactcat ccatcagtcg attacaggcc tgtacgagac acggattgac 4080
ctctcgcagc tg 4092
<210> 111
<211> 4101
<212> DNA
<213> 人工序列
<220>
<223> 合成的Cas9
<400> 111
gacaagaagt attccatagg cctggctatc ggcaccaaca gcgtgggctg ggccgtcatc 60
accgacgagt acaaagtgcc gagtaaaaag ttcaaagtgc tcggcaacac cgaccgccac 120
tccataaaga aaaacctgat cggggcgctc ctgttcgaca gcggcgagac ggcggaggcc 180
acccgcttga aacgcacggc ccgacggcgc tacacgcggc gcaagaaccg gatctgttac 240
ctacaggaga ttttctctaa cgagatggcg aaggtggacg actcgttctt tcaccgcctc 300
gaagagtcct tcctcgtgga ggaggacaag aaacacgagc gccacccgat cttcggcaac 360
atcgtggacg aggtggccta ccacgagaag tacccgacca tctaccacct ccggaagaaa 420
ctcgtggaca gcacggacaa ggccgacctg aggctcatct acctcgccct ggcgcacatg 480
attaagttcc ggggccactt cctgatcgag ggcgacctga acccggacaa cagcgacgtg 540
gacaagctgt tcatccagct agtccagacc tacaaccagc ttttcgagga aaaccccatc 600
aacgccagcg gggtggacgc gaaggcgatc ctgtccgccc ggctgagcaa gtcccggcgg 660
ctggagaacc tcatcgcgca gttgcccggc gagaagaaga acgggctgtt cgggaacctg 720
atcgccctct ccctggggct caccccgaac ttcaagtcca acttcgacct cgccgaggac 780
gccaaactac agctgagcaa ggacacctac gacgacgacc tcgacaacct gctggcccag 840
atcggggacc agtacgcaga cctgttcctc gccgccaaga acctctccga cgccatcctg 900
ctgtcggaca tcctgcgggt gaacacggag atcacgaagg ccccgctctc ggcctcgatg 960
attaaacgct acgacgagca ccaccaggac ttgaccctcc tcaaggcgct ggtccgccag 1020
cagcttcccg agaagtacaa ggaaatcttt ttcgatcaga gcaagaacgg gtacgccggg 1080
tacatcgacg gcggggcgtc ccaggaggag ttctacaagt tcatcaagcc catcctggag 1140
aaaatggacg ggaccgagga gctgctcgtg aagctcaacc gcgaagattt gctccgcaag 1200
cagcgcacgt tcgacaacgg gtcgatcccg caccagatcc acctgggcga gctgcacgcg 1260
atcctcaggc gtcaggaaga cttctacccc ttcctcaagg acaaccgcga gaagatagag 1320
aagattctga ccttcagaat tccttattac gtgggcccgc tggctcgggg caactcgcgc 1380
ttcgcctgga tgacgcgcaa gtccgaggag accatcaccc cgtggaactt cgaggaggtg 1440
gtggataagg gtgcctcggc ccagtccttc atcgagcgga tgaccaactt cgacaagaac 1500
ctgccgaacg agaaggtgct ccccaagcac agcctgctct acgaatattt cacggtgtac 1560
aacgagctga cgaaggtcaa gtacgtgacc gagggaatga ggaaacctgc attcctctcc 1620
ggggagcaga agaaagccat agtcgacctc ctgttcaaga ccaaccggaa ggtcaccgtc 1680
aagcagctca aggaggacta cttcaagaag atcgagtgct tcgattcagt ggagatcagc 1740
ggcgtcgagg accggttcaa cgccagcctg ggcacctacc acgacctgct caagatcatc 1800
aaggacaagg acttcctcga caacgaggag aacgaggaca tcctggagga catcgtgctg 1860
accctgacgc tcttcgagga ccgcgagatg atcgaggagc gcctcaagac ctacgcccac 1920
ctgttcgacg acaaggtgat gaagcagctc aagcggcgga gatatactgg gtggggccgc 1980
ctctcccgga agctcattaa cggtatcagg gataagcagt ccgggaagac gatcctcgac 2040
ttcctcaagt cggacgggtt cgccaaccgc aacttcatgc agctcatcca cgacgactcc 2100
ctgacgttca aggaggacat ccagaaggcc caagtgtctg gtcaaggtga ctcgctccac 2160
gagcacatcg ccaacctcgc gggcagcccg gccatcaaga agggaatact ccagaccgtc 2220
aaggtggtgg acgagctggt gaaggtcatg ggccgccaca agccggagaa catcgtcatc 2280
gagatggcgc gggagaacca gaccacgcag aaggggcaga aaaatagccg tgagcgcatg 2340
aagcgcatcg aggaggggat taaggagttg ggcagccaga tcctcaagga gcaccctgtg 2400
gagaacacgc agttgcaaaa cgagaagctc tacctgtact acctccagaa cgggagggat 2460
atgtacgtgg accaagaact ggacatcaac cgcctgtccg actacgacgt ggaccacatc 2520
gtgccgcaga gcttcctcaa ggacgacagc atcgacaaca aggtgctcac ccggtccgac 2580
aagaatcggg gcaagtccga caacgtgccc agcgaggagg tcgtcaaaaa gatgaaaaac 2640
tactggcgac aactactgaa cgccaagctc atcacccagc gcaagttcga caacctcaca 2700
aaagccgagc gcggcgggtt gagcgagctg gacaaggccg ggttcatcaa gcgccagctc 2760
gtcgagacgc gccagatcac gaagcacgtc gcgcagatac tcgacagccg gatgaacacc 2820
aagtacgacg agaacgacaa gctcatccgg gaggtgaagg tcatcaccct caagtcgaag 2880
ctcgtgagcg acttccgcaa ggacttccag ttctacaagg tccgggagat caacaactac 2940
caccacgccc acgatgctta tcttaacgcc gtggtgggga cggccctcat taagaaatac 3000
ccgaagctgg agtcggagtt cgtgtacggc gactacaagg tgtacgacgt caggaagatg 3060
atcgccaagt ccgaacagga gatcgggaag gccacggcga aatacttctt ctacagcaac 3120
atcatgaact tcttcaagac cgagatcacc ctcgccaacg gcgagatccg caagcgcccg 3180
ctcatcgaga cgaacgggga gaccggcgag atcgtctggg acaaggggcg cgacttcgcc 3240
actgtgcgga aggtgctgtc gatgccccag gtcaacatcg tcaagaagac ggaggtccag 3300
acgggcgggt tcagcaagga gagcatcctg ccgaagcgca acagcgacaa gctgatcgcc 3360
cgcaaaaagg actgggatcc aaaaaagtac ggcggcttcg acagccccac cgtcgcctac 3420
agcgtcctcg tcgtcgctaa agtcgagaag ggcaagtcca aaaagctcaa gagcgtcaag 3480
gagctgctcg ggatcaccat catggagcgg tccagcttcg agaagaaccc aattgatttc 3540
ctggaggcga agggctacaa ggaggtcaag aaagacctca tcataaagct gccgaagtac 3600
tcactcttcg agctggagaa cgggcgcaag cggatgctgg cgtcggccgg agagctccaa 3660
aagggcaacg agctggcgct gccgagcaag tacgtgaact tcctctacct ggcgtcccac 3720
tacgagaagc tcaagggcag tccagaggat aacgagcaga agcagctatt cgtggagcag 3780
cacaagcact acctggacga gatcatcgag cagatcagcg agttctccaa gcgcgtcatc 3840
ctggcggacg ccaacctgga caaggtgctg tccgcgtaca acaagcaccg cgacaagccg 3900
atccgcgagc aagccgagaa catcatccac ctgttcaccc tcacgaacct cggggcaccc 3960
gccgccttca aatatttcga cacgaccatc gaccgcaagc gctacaccag cacgaaggag 4020
gtgctcgacg ccaccctgat ccaccagagc atcaccgggc tgtacgagac ccgcatcgac 4080
ctctcgcagc tcggcgggga c 4101
<210> 112
<211> 4101
<212> DNA
<213> 人工序列
<220>
<223> 合成的Cas9
<400> 112
gacaagaagt acagtattgg attggccatc gggacgaaca gcgtgggctg ggccgtcatc 60
accgacgagt acaaggtgcc atccaagaag tttaaggttc tggggaatac cgaccgccac 120
tcgatcaaga aaaatctcat cggggcgctg cttttcgaca gcggcgagac ggcggaagcg 180
acgcggctca agcggacggc tcgtcgccgt tacacccggc gtaagaaccg catctgttac 240
ctccaggaga tattcagcaa cgagatggcg aaggtggacg actccttttt ccaccgtctt 300
gaggagtcct tcctggtcga ggaggacaag aagcacgagc gccacccgat cttcgggaac 360
atcgtggacg aggtggccta ccacgagaag taccccacga tctaccacct ccgcaaaaaa 420
ctcgtggact caactgacaa ggccgatttg aggcttatct acctcgccct cgcccacatg 480
attaagttcc gtgggcactt cctaatcgag ggtgacctca accccgacaa ctctgacgtg 540
gacaagctgt tcatccagct tgtgcagacc tacaatcagc tctttgagga gaatccgatc 600
aacgcatctg gtgtggacgc aaaggccatc ctcagcgcgc ggctgagcaa gtctaggcgg 660
ttggagaacc tgatcgccca actgcccggc gagaagaaaa atggcctctt cggcaacctg 720
atcgccctgt cgctggggct cacgccgaac ttcaagagta actttgacct ggcggaggac 780
gctaagctcc agctatctaa ggacacatac gacgacgacc tggacaacct gctggcccag 840
atcggcgacc agtacgccga cctcttccta gccgccaaga acctgtccga cgccatcctc 900
ctcagcgaca tcctgcgcgt gaacacggag atcacgaagg ctccgctcag cgcctccatg 960
attaagcggt acgacgagca ccaccaagac ctaactttac tcaaagccct cgtgcggcag 1020
cagcttcccg agaagtacaa agagatattt tttgatcagt ccaagaacgg ttatgcgggc 1080
tacatcgacg gcggcgcgag ccaggaggag ttctacaagt tcatcaagcc catcctggag 1140
aagatggacg gcacggagga gctgctcgtg aagctcaacc gtgaagacct cctgcgaaag 1200
cagcgaacct tcgacaacgg ttcgatcccg caccagatcc acctcgggga gctgcacgcc 1260
atcctgaggc gacaggagga cttctaccct ttcctaaagg acaaccgcga gaagattgaa 1320
aaaatcctga cgtttcgcat accctactac gtcggcccgc tggcgcgcgg caactcccgg 1380
ttcgcctgga tgacccgtaa gagcgaggag acgatcaccc cgtggaactt cgaggaggtc 1440
gtggacaagg gcgcgagcgc gcagagcttc atcgagcgca tgaccaactt cgacaagaac 1500
ctcccgaacg agaaggtgct cccaaagcac tccctcctgt acgagtattt caccgtgtac 1560
aacgagttga caaaggtgaa gtacgtgacg gagggaatgc ggaagcctgc gttcctctcg 1620
ggcgagcaga agaaggcaat cgtggacctg ctcttcaaga ccaaccggaa ggtgacggtg 1680
aagcagctca aggaggacta cttcaaaaaa atcgagtgct tcgactccgt ggagataagc 1740
ggcgtggagg accgattcaa cgcctccctc ggcacctacc acgacctcct taagatcatc 1800
aaggacaagg acttcctgga caacgaggag aacgaggaca tcctggagga catcgtgctc 1860
accctgaccc tcttcgagga ccgggagatg atcgaggagc gcctcaagac gtacgcccac 1920
ttgttcgacg acaaggtgat gaagcagctc aagcggcggc gatacaccgg gtggggccgc 1980
ctatcccgca aacttatcaa cggcatccgc gacaagcagt ccggcaagac gatcctggat 2040
ttcctcaagt cggacgggtt cgccaaccgg aacttcatgc agctcatcca cgacgacagc 2100
ctcacgttca aggaggacat ccagaaggcc caagtgagcg gtcaagggga cagcctccac 2160
gagcacattg cgaaccttgc tgggagccct gcgatcaaga aggggatatt gcaaaccgtg 2220
aaggtcgtgg acgagttggt gaaggtcatg gggcgacaca agcccgagaa catcgtgatc 2280
gagatggcca gggaaaatca gaccacgcag aagggccaaa aaaacagccg cgagcggatg 2340
aagcggatcg aggagggcat caaggagctg gggtcgcaga tcctcaagga gcacccggtg 2400
gagaacacgc agctccagaa cgagaagctg tacctctatt acctacagaa cgggcgggat 2460
atgtacgtgg accaggagct agacatcaac cgcctgtccg actacgacgt ggaccatatc 2520
gtcccgcagt cgttcttgaa ggacgacagc atcgacaaca aggtgctcac aagatcggat 2580
aagaatcgag gcaagtccga caacgtgccc tcggaggagg tggtcaagaa aatgaaaaac 2640
tactggcggc agttgctgaa cgccaagctc attacgcagc ggaagttcga caacctgacg 2700
aaggctgaac gtggtgggct cagcgagcta gacaaggcgg ggttcatcaa gcggcagctc 2760
gtcgagaccc ggcagatcac caagcacgtg gcgcagatcc tggactcgcg catgaacacc 2820
aagtacgacg agaacgacaa gctcatccgt gaggtgaagg tcatcaccct taagtctaag 2880
ctggtcagtg acttccgcaa ggacttccag ttctacaagg tccgggagat caacaactac 2940
caccacgcgc acgacgccta cctcaacgcg gtggtgggga cggcgcttat taagaaatat 3000
cccaagctgg aaagcgagtt cgtttacggc gactacaagg tgtacgacgt ccgcaagatg 3060
atcgcaaagt cggaacagga aatcggaaag gcgacggcca aatatttctt ttactccaac 3120
atcatgaatt tttttaagac ggagatcacc ctggcgaacg gggagatccg caagcggccc 3180
ctcatcgaga ccaacgggga gacgggcgag atcgtctggg acaagggccg ggacttcgcc 3240
accgtgcgga aggtgctttc tatgcctcaa gtcaatatcg tcaaaaagac agaggtgcag 3300
accggcgggt tcagcaagga gtctatcctg ccgaagcgca actcggacaa gctcatcgcg 3360
cgcaagaaag actgggaccc caaaaaatat ggcgggttcg actcgccgac cgtcgcctac 3420
agcgtcctcg tggtggctaa ggtcgagaag ggcaagagca aaaagctaaa gtcggtgaag 3480
gagctgctgg gcatcaccat catggagcgc tcgtctttcg agaagaatcc aatcgacttc 3540
ctagaggcga aggggtacaa ggaggtcaaa aaggatctta tcatcaaact gccgaagtac 3600
agtctgttcg agctggagaa cgggcggaag cggatgctgg ctagtgcggg cgagttgcag 3660
aagggcaacg agttggcact gccctccaag tacgtgaact tcctgtacct ggcctcccac 3720
tacgagaagc tcaaggggag ccccgaggac aacgagcaga agcagctatt cgtcgagcag 3780
cacaagcact acctggacga gatcatcgag cagatcagtg agttctccaa gcgggtcatc 3840
ctcgcggacg ccaacctgga caaggtgctg agcgcgtaca acaagcacag ggacaagcca 3900
atcagggaac aggccgagaa catcatccac ctgttcaccc tgaccaacct gggtgcaccg 3960
gctgccttca agtactttga cacgaccatc gaccggaagc gctacacctc cacgaaggag 4020
gtgctggacg ccacgctgat ccaccagagc atcaccgggc tctacgagac acggatcgac 4080
ctgagccagc ttggcgggga c 4101
<210> 113
<211> 4092
<212> DNA
<213> 人工序列
<220>
<223> 合成的Cas9
<400> 113
gacaaaaagt attccattgg actcgctatc ggcacgaaca gcgtcgggtg ggcggtcatc 60
actgacgagt acaaggtgcc gagcaagaag tttaaggtgc tgggaaacac cgacaggcac 120
tcgatcaaga aaaatcttat cggggcccta ctcttcgact ccggagaaac cgccgaggcc 180
acccggttga agcgcacggc ccgccgtcgc tacaccaggc gcaagaaccg gatctgctac 240
ctccaggaga tattcagcaa tgagatggcg aaggtggacg actcgttttt tcacaggcta 300
gaggagtctt tcctcgtgga ggaggacaag aaacacgagc gccaccccat cttcggcaac 360
atcgtggatg aggtggcata tcacgagaag tacccaacca tctaccacct ccgcaaaaag 420
ctcgtggact ctaccgacaa ggccgacctc cgtctgatct acctcgcgct ggcccacatg 480
attaagttcc gaggacactt tctgatcgag ggcgacctga acccagacaa cagcgacgtg 540
gacaagctgt tcatccaact tgtccagacc tacaatcagc tcttcgagga gaaccctatc 600
aacgcctcgg gcgtggacgc gaaggccatc ctgtccgccc gcctgagcaa gtcgcggcgg 660
ctggagaacc tgatcgccca gctccccggc gaaaaaaaga acggcctctt cggcaacctc 720
atcgcgttgt cgctggggct caccccgaac ttcaagtcca acttcgacct ggccgaggac 780
gctaaactcc agctctcgaa ggatacctac gacgacgacc tcgacaacct gctggcccag 840
atcggcgacc agtacgcgga ccttttcctg gcggccaaga acctgagcga cgcgatcctc 900
cttagcgaca tactccgtgt gaacaccgag atcacgaagg ccccgctctc cgcgtccatg 960
attaagcgct acgacgagca ccaccaagac cttaccctgc ttaaggcgct ggtcaggcag 1020
cagttaccgg agaagtacaa ggagatcttt tttgatcaat ctaagaacgg ttacgccggg 1080
tacatcgacg gcggcgcgtc ccaggaggag ttctacaagt tcatcaagcc gatcttggag 1140
aaaatggacg ggaccgagga gctgctcgtg aagctcaacc gcgaagacct cctccgcaag 1200
cagcgcacct tcgacaacgg gagcatcccg caccagatcc acctgggaga gctgcacgcg 1260
atcctgcgga gacaagagga cttctacccc ttcctcaagg acaaccggga gaagattgaa 1320
aaaatactta cttttcgtat cccgtactac gtcgggcccc ttgcgagggg caactccaga 1380
ttcgcgtgga tgacccgcaa gtccgaggag accatcaccc cgtggaactt cgaggaggtg 1440
gtggacaagg gcgcgtcggc ccagtcgttc atcgagcgca tgaccaactt cgacaagaac 1500
cttccgaacg agaaggtgct cccgaagcac agcctgctct acgaatattt tactgtgtac 1560
aacgagctga cgaaggtcaa gtacgttacg gaggggatga ggaagcccgc cttcctctcc 1620
ggcgagcaga agaaagccat tgtggatctc ctgttcaaga ccaaccgcaa ggtgacggtg 1680
aaacagctca aagaggacta cttcaagaag atcgagtgct tcgactccgt agagatcagc 1740
ggggtcgagg accgcttcaa cgcctcgctg ggcacgtacc acgacctgct aaagattatc 1800
aaggacaaag acttcctaga caatgaggag aacgaggaca ttctggagga catcgtgctg 1860
actctgacgc tgttcgaaga ccgcgagatg atcgaggagc ggcttaagac gtacgcccac 1920
ctgttcgacg acaaggtgat gaagcagttg aaacggcggc gctacaccgg gtggggccgc 1980
ctctcccgca agctcatcaa cggcatccgc gacaagcagt cggggaagac gatcctggac 2040
ttcctcaaga gcgacggctt cgccaaccga aacttcatgc agctaatcca cgacgacagc 2100
ctgacgttca aggaggacat ccagaaggcc caagtgagcg gccagggaga ctcgctacac 2160
gagcatatcg ccaacctggc tggcagcccg gcgattaaga aaggaatcct ccaaaccgtc 2220
aaagtggtgg acgagctggt gaaggtgatg ggccgccaca agcccgagaa cattgtgatc 2280
gagatggcgc gggagaacca gacgacgcag aagggccaaa aaaatagcag ggaaaggatg 2340
aagcgaatag aggaggggat caaggagctg gggagccaga ttctcaaaga gcacccggtc 2400
gagaacacac agctccagaa cgagaagctg tacctctact acctccaaaa cggccgcgat 2460
atgtacgtgg accaggaact agacatcaac cggctgagcg actatgacgt ggaccacatc 2520
gtgccgcagt ccttcctcaa ggacgactcg attgacaaca aagtgctcac tagatccgac 2580
aagaacagag gcaagagcga taacgtcccg tcggaggagg tcgtcaagaa aatgaaaaac 2640
tactggcggc agctcctaaa cgccaagctc atcacgcagc gtaagttcga caacctgacg 2700
aaggcggagc ggggcgggct gagcgagctg gacaaagcgg ggttcatcaa gcggcagctc 2760
gttgagacgc ggcagatcac aaagcacgtc gcgcaaatcc tcgactcccg catgaacacc 2820
aagtacgacg agaacgacaa gctcatccgg gaggtgaagg tcattaccct taaatcgaag 2880
ctcgtcagcg actttcgtaa ggacttccag ttctacaagg tcagagagat caacaactac 2940
caccacgccc acgacgccta tctgaacgcc gtggtgggca ccgcgcttat taagaagtac 3000
cccaagctgg agtccgagtt cgtgtacggc gactacaagg tttatgacgt caggaagatg 3060
atcgccaagt cggaacagga gatcggaaaa gctaccgcca aatatttctt ctatagcaac 3120
atcatgaact tcttcaaaac cgagatcacc ctcgccaacg gcgagatccg gaagcgcccg 3180
ctcatcgaga ccaacgggga gaccggggag atcgtctggg acaaggggcg ggacttcgct 3240
actgtccgaa aggtgctctc catgccacaa gtgaatatcg tcaagaaaac agaggtgcag 3300
accggagggt tcagtaagga gtccatcctg cccaagcgga actccgacaa gctaattgct 3360
cgcaaaaagg attgggatcc taaaaaatat ggcggcttcg actcgcccac ggtcgcctac 3420
tctgtgctgg tcgtggcgaa ggtggagaag ggcaagtcca agaagctcaa gagcgtcaag 3480
gagctgctgg ggatcacgat catggagcgt agttcgtttg agaagaatcc catcgacttc 3540
ctggaggcta agggctacaa ggaggtcaaa aaggacctca tcattaagct gccgaagtac 3600
agcctcttcg agctggagaa cgggcggaag cgtatgctcg cctccgctgg ggagttacaa 3660
aaggggaacg agctggcgct gccgtctaag tacgtcaact tcctgtacct ggcctcccac 3720
tacgagaagc tcaaggggtc gccggaggac aacgagcaga agcagctctt cgtagagcag 3780
cacaagcact acctggacga gatcatcgag cagatttcag agttctcaaa gcgggtcatc 3840
ctcgccgacg ccaacctgga caaggtgctc tcggcctaca acaagcaccg ggacaagccg 3900
atccgcgaac aggccgaaaa catcatccac ctgttcacgc tcaccaacct cggtgccccg 3960
gcggccttca agtactttga cacgaccatc gaccggaagc gctatacctc gacgaaggag 4020
gtgctggacg ccaccctgat ccaccagtcc atcaccgggc tttacgagac ccggatcgac 4080
ctctcgcagc ta 4092
<210> 114
<211> 4101
<212> DNA
<213> 人工序列
<220>
<223> 合成的Cas9
<400> 114
gacaagaagt atagtattgg actcgccatc ggaaccaact ctgtggggtg ggctgttatt 60
acagatgaat ataaggtgcc atccaaaaag tttaaagttc tgggcaatac tgatagacac 120
tcaatcaaga agaatctgat aggtgcactt ctgtttgata gtggagagac tgccgaggca 180
accagactta aaaggactgc aagaagaaga tataccagaa gaaagaatag gatttgctat 240
ttgcaggaaa tcttcagcaa cgaaatggcc aaggttgatg actcattttt ccataggttg 300
gaggagagtt ttcttgtgga ggaagataag aagcacgaaa gacacccaat tttcgggaat 360
atagtggacg aggtggctta tcatgagaag tatcccacta tctaccacct gagaaagaaa 420
cttgtggact caaccgataa ggctgatctt aggcttatat acttggccct tgcacatatg 480
atcaaattca ggggccattt tcttatcgaa ggcgatctta atcccgataa ctcagatgtg 540
gacaagctgt ttatacaact tgtgcaaacc tacaatcaac tcttcgagga gaatcccatt 600
aacgcctccg gcgtggatgc aaaagccata ctgtcagcca gactgagcaa aagtaggaga 660
ctggagaatc ttatagccca actgcccggt gaaaagaaga atgggctctt cggaaatctg 720
atcgctcttt cattggggtt gacacccaac tttaagagta actttgactt ggcagaagat 780
gcaaagttgc agctcagtaa agacacatat gacgatgacc ttgacaatct cttggcacaa 840
ataggggatc aatacgctga ccttttcctc gctgccaaga acctcagcga cgctatactg 900
ttgtccgaca ttcttagggt taataccgaa attacaaagg cccctcttag tgcaagtatg 960
atcaaaaggt atgatgagca tcaccaagac cttacactgc tgaaggctct ggttagacag 1020
caactccctg aaaagtataa ggaaatattc ttcgaccaaa gtaagaacgg gtacgccggt 1080
tatattgatg ggggcgcaag tcaagaagaa ttttacaaat tcatcaagcc aattcttgaa 1140
aagatggacg ggactgagga attgctggtg aaactgaata gagaggacct tcttagaaaa 1200
cagaggacat ttgacaatgg gtccatccca caccagattc atctggggga actccacgca 1260
atattgagga gacaagaaga cttttaccca ttccttaagg ataatagaga gaaaatcgaa 1320
aaaatcctga ctttcaggat tccttactat gttgggccac tggccagggg gaactcaaga 1380
ttcgcttgga tgacaaggaa gtcagaagaa accataaccc cttggaattt tgaagaggtg 1440
gttgataagg gggcatcagc ccagtctttc atagagagga tgaccaactt tgataaaaat 1500
cttccaaatg agaaggtttt gccaaaacat agtcttttgt acgagtactt tactgtttat 1560
aacgaattga ccaaggtgaa gtatgtgacc gagggaatga ggaagccagc atttttgtcc 1620
ggggagcaaa agaaagcaat cgttgatctt ctcttcaaga ccaacagaaa agtgaccgtg 1680
aaacaactga aggaagacta cttcaaaaag atagaatgtt tcgattcagt ggaaattagc 1740
ggtgttgaag acaggttcaa tgcttcattg ggtacttacc acgacctgtt gaagataatc 1800
aaagacaagg actttctcga taatgaggag aacgaagaca tcttggaaga cattgtgctt 1860
acactcactt tgtttgagga cagggaaatg attgaggaaa gactcaaaac ttacgctcat 1920
ttgtttgatg ataaggttat gaaacaacta aaaagaagaa ggtacaccgg ctggggaaga 1980
ttgagtagga aactgatcaa cggtattaga gataaacaat ccggaaagac tatcctcgat 2040
ttccttaaga gtgatggctt tgcaaatagg aattttatgc agctgattca tgacgactca 2100
cttaccttca aagaagacat ccaaaaagct caggtgtctg ggcaaggcga cagtctgcat 2160
gaacatatag ctaacttggc tgggagtccc gccatcaaga aggggatact tcaaacagtt 2220
aaagttgtgg acgaattggt gaaggtaatg ggaaggcaca agcctgaaaa tatagtgata 2280
gaaatggcaa gggaaaatca aacaacccag aagggacaga agaacagtag ggaaaggatg 2340
aaaaggatag aagaggggat caaagagctt ggtagccaga tcctcaagga acatccagtg 2400
gagaataccc aacttcaaaa cgagaaactc tatttgtact acttgcagaa cggaagagat 2460
atgtatgtgg accaagagct tgatattaac aggctgagcg attatgacgt tgaccacata 2520
gtgccccaat cattcctcaa ggatgactct attgataata aggtgctgac aaggagtgac 2580
aagaatagag ggaaatccga caacgttcca tccgaggaag ttgtgaagaa gatgaagaac 2640
tactggaggc agttgctgaa cgctaagctc attacccaga ggaaattcga taacctgacc 2700
aaagcagaga gaggcgggct gagcgaactc gataaagcag gtttcatcaa gagacaactc 2760
gtggagacta ggcaaattac taagcacgtg gctcaaatac tcgacagcag gatgaacaca 2820
aagtacgacg agaacgacaa gctcattaga gaggttaagg ttattactct gaaaagtaaa 2880
ttggttagcg atttcagaaa ggatttccaa ttctataagg ttagagagat caacaattat 2940
catcatgcac atgatgccta tctgaatgct gtggttggta cagcccttat caagaagtac 3000
cctaagctag agagcgagtt tgtgtacgga gattataagg tgtatgatgt gaggaaaatg 3060
atcgctaaaa gtgagcaaga gattggaaag gctaccgcca aatacttctt ttattccaat 3120
attatgaatt tcttcaagac agaaatcacc ctggctaacg gcgagataag gaagaggccg 3180
cttatcgaaa ctaatgggga gacaggcgaa atagtgtggg acaaagggag ggatttcgca 3240
actgtgagga aggttttgag catgcctcag gtgaatatcg ttaagaaaac cgaagttcaa 3300
actggagggt tctctaagga aagcattctc cccaagagga actccgacaa gctgattgct 3360
agaaagaaag actgggaccc caagaagtat ggcggattcg actcacccac tgtggcatat 3420
agcgttctcg tggtggcaaa ggttgaaaag ggtaaatcca aaaaactcaa atccgtgaag 3480
gaactccttg gcataactat tatggaaagg agtagctttg aaaagaatcc catcgacttt 3540
ctcgaagcta agggctataa ggaagttaag aaggacctta taatcaaact tccaaaatac 3600
tccctttttg agttggaaaa cggcagaaag agaatgttgg ccagtgccgg ggagcttcaa 3660
aagggcaacg aactggctct gcctagcaaa tatgtgaact ttttgtatct ggcatcacac 3720
tacgagaaac ttaaaggctc tcctgaggac aacgagcaaa aacagctctt tgttgaacag 3780
cataagcact acctcgacga gattattgag cagatcagcg agttctcaaa gagagttatt 3840
ctggctgacg ctaatcttga caaggttttg tccgcttaca acaaacacag ggataagcca 3900
atcagggagc aggcagaaaa cataatccat ctctttaccc tgacaaacct cggtgccccc 3960
gctgctttca agtattttga tactaccatt gacaggaaga gatatacttc cactaaggaa 4020
gtgctcgacg caaccctcat acaccaaagt atcacaggcc tctatgaaac taggatagat 4080
ttgtctcaac ttgggggcga t 4101
<210> 115
<211> 4101
<212> DNA
<213> 人工序列
<220>
<223> 合成的Cas9
<400> 115
gacaaaaagt attccatcgg gcttgctatc ggaaccaact ctgtggggtg ggcagttatt 60
accgacgaat acaaggtgcc cagcaagaag tttaaggttc tggggaacac agatagacat 120
agcataaaga aaaacctgat aggcgcactg ttgttcgact ccggggaaac agccgaagct 180
accaggctga agagaactgc aagaagaagg tacaccagaa gaaaaaacag aatatgttat 240
ctccaagaga ttttctctaa cgagatggcc aaggtggacg actcattctt tcacagactg 300
gaagaatctt tccttgtgga agaagataag aaacacgaga ggcaccctat ttttggcaat 360
atcgtggatg aggtggctta ccacgaaaaa taccctacaa tataccacct caggaaaaaa 420
ttggttgata gtacagacaa ggccgacctc aggctcatct atttggccct ggcccatatg 480
attaaattca gggggcactt tctcatcgag ggagatttga accccgacaa cagtgatgtt 540
gataagctct ttattcagct cgtgcagact tacaatcagt tgtttgagga aaaccccatt 600
aatgcttccg gggtggacgc caaggcaatc ctttctgcaa gactctcaaa gtcaaggaga 660
ctcgaaaatc tgatagcaca gcttccagga gagaagaaga acgggctctt tggaaacctg 720
atcgctctgt cactcggact cacacccaat ttcaaaagca attttgattt ggcagaggac 780
gctaagctgc aactcagtaa ggatacctac gacgatgact tggataatct gctcgcacaa 840
attggggacc agtatgcaga cctgtttctc gcagctaaga acttgagtga cgccatattg 900
ctcagtgaca tcctcagggt taataccgag attacaaaag ctccactctc tgcaagcatg 960
atcaagaggt atgacgagca ccatcaagac ctgacactcc ttaaggcgtt ggttaggcag 1020
caacttcctg aaaagtataa ggaaatcttc ttcgatcaaa gcaaaaacgg ctacgccggc 1080
tatatagacg ggggagcatc ccaagaagaa ttttataagt tcataaaacc tatattggag 1140
aagatggacg ggacagagga attgctcgtg aaactgaaca gggaggatct cctcaggaag 1200
caaaggacct tcgacaatgg ctccatccca catcagattc acctcggcga actgcacgca 1260
atactgagaa gacaagagga cttttatcct ttcctgaagg acaacaggga gaaaatcgag 1320
aaaatcttga cattcagaat cccatactac gttgggcctc tggccagagg taacagtagg 1380
ttcgcctgga tgactaggaa atcagaggag actattacac cctggaactt tgaagaagtt 1440
gttgataagg gagcttcagc acaatcattc atcgaaagaa tgacaaactt tgacaaaaat 1500
ctgcctaatg agaaagtgct cccaaaacat tccctgctgt atgagtattt taccgtttat 1560
aacgagctta ccaaggtgaa atacgttact gaaggtatga gaaagccagc ttttctttca 1620
ggggagcaaa agaaggctat cgtggatctt ctctttaaga ccaacagaaa ggttaccgtg 1680
aagcagctta aggaagacta ctttaaaaag atcgagtgtt ttgactcagt ggaaataagc 1740
ggtgttgaag atagattcaa cgcatccttg ggaacttatc atgatcttct taagataatc 1800
aaggataaag actttctcga caacgaggaa aacgaagata tactggagga catagttctg 1860
acacttactt tgttcgagga tagggagatg atcgaggaaa gactgaaaac atatgctcac 1920
cttttcgacg acaaagttat gaaacaactc aagagaagga gatatacagg gtgggggaga 1980
ttgagcagga aactgattaa tggtatcaga gacaaacagt caggaaaaac aatactcgac 2040
tttttgaaat cagacgggtt cgcaaatagg aatttcatgc agcttataca cgacgattca 2100
cttactttta aagaggacat tcaaaaggct caagttagtg gacaaggtga ctccctccac 2160
gaacacatcg caaatctcgc tggcagccct gcaattaaga agggtatact ccagacagtt 2220
aaggttgttg acgagctggt taaagtgatg ggaagacaca aacccgagaa catagtgata 2280
gagatggcca gggaaaacca aaccactcaa aaagggcaga aaaattccag agagaggatg 2340
aaaaggattg aagaaggtat caaggagctg ggtagccaaa ttctgaaaga acatcctgtg 2400
gaaaacactc aactccagaa tgagaaactc tatctgtact atctgcaaaa tgggagagat 2460
atgtatgtgg accaggaact ggacataaac aggctctcag attacgatgt ggatcatatc 2520
gtgccacagt cctttcttaa ggatgatagc atcgacaata aggtgcttac caggtccgac 2580
aagaacaggg gaaagtcaga taacgtgcct tctgaagaag ttgttaaaaa gatgaagaac 2640
tactggagac agctgcttaa cgctaagctc ataacacaga ggaagtttga caacttgacc 2700
aaggccgaga gaggcggact ctcagaattg gataaggcag ggttcataaa aaggcagctg 2760
gtggaaacaa ggcagataac taaacatgtg gctcagatcc tcgatagtag gatgaataca 2820
aaatacgatg agaacgacaa gctcataagg gaggttaaag tgataactct gaaatccaaa 2880
ctggttagcg attttaggaa ggatttccag ttttacaaag ttagggagat caacaattat 2940
catcacgccc acgatgccta cttgaacgca gttgtgggta ctgcacttat caaaaagtac 3000
cctaagctgg aatccgagtt tgtttatgga gactataagg tgtacgacgt tagaaaaatg 3060
attgcaaagt cagagcagga gatagggaaa gccactgcaa aatatttctt ttatagcaat 3120
atcatgaatt tctttaagac agaaatcaca ctggccaatg gggaaataag gaagaggccc 3180
ctgatcgaaa ctaatggcga gacaggggag attgtgtggg ataaaggtag ggactttgca 3240
acagtgagga aagtgctgag catgccccaa gttaatatcg ttaaaaagac cgaggttcaa 3300
acagggggct ttagtaagga aagcattttg cccaagagga atagtgacaa attgattgct 3360
aggaaaaaag attgggaccc caaaaagtat ggcggatttg atagccccac tgttgcttac 3420
tccgtgctcg tggttgcaaa ggtggagaag ggaaagagca agaaactgaa gtcagttaag 3480
gaactccttg gtatcactat catggaaaga agctcctttg agaagaaccc tattgacttc 3540
ctggaggcta aagggtacaa agaggttaag aaagacctta tcattaaatt gcccaaatat 3600
agtcttttcg agcttgaaaa cggaagaaag aggatgcttg catccgctgg cgaattgcaa 3660
aagggcaatg agcttgctct cccttccaag tatgtgaact tcctttatct tgcctcacac 3720
tatgaaaaac tcaaaggttc acccgaagac aacgaacaaa agcaactatt tgtggaacaa 3780
cacaagcact acctggacga aatcattgag caaatttctg agttttcaaa aagggtaatc 3840
ttggctgacg caaatctcga caaagttttg tcagcttaca acaaacatag agataagcca 3900
attagagagc aagctgagaa tatcatccat ctgtttaccc tgactaacct tggagcgcct 3960
gctgctttta aatatttcga caccacaatc gacaggaaga ggtacactag cactaaggaa 4020
gttctcgacg ccaccctcat ccaccagagt attacaggcc tgtacgagac aagaattgat 4080
ctttctcaac ttggtggtga c 4101
<210> 116
<211> 4101
<212> DNA
<213> 人工序列
<220>
<223> 合成的Cas9
<400> 116
gataagaagt actcaatcgg tctggcaatc ggaaccaact ctgtgggttg ggcagtgatt 60
acagatgagt ataaggtgcc aagcaaaaaa ttcaaggtgc tgggtaatac cgacagacac 120
agcattaaga agaatttgat tggagcactc ctctttgact caggggaaac agcagaggca 180
acaaggctga agaggacagc aaggcggagg tacacaaggc ggaaaaacag gatatgctac 240
ctccaggaaa tctttagcaa cgagatggct aaagtggatg atagcttttt ccatagactc 300
gaagaatcct ttcttgttga agaggacaaa aagcatgaaa ggcatcccat cttcggcaat 360
atagttgatg aggttgcata ccatgagaag taccccacaa tctaccacct cagaaagaaa 420
cttgtggact ccacagataa agcagacctg aggctcatat acctcgcact cgcacacatg 480
atcaagttca gagggcactt tctcatcgaa ggtgacctga atccagataa ttcagatgtg 540
gataaactgt ttatacagct ggtgcaaaca tacaaccaac ttttcgagga aaacccaatc 600
aatgcctccg gtgttgatgc aaaggccatc ctgtcagcaa gactcagcaa aagcaggcgg 660
ctcgaaaacc tcatcgccca gcttcccggt gaaaagaaga acgggctctt tggtaatctc 720
atcgcattga gccttggtct tactccaaac ttcaagagca attttgatct ggcagaggat 780
gctaaactgc aactctcaaa ggacacatat gacgatgacc ttgacaatct gttggcccag 840
atcggggacc aatatgcaga cctcttcctg gccgcaaaga atctgtcaga tgcaatcctc 900
ttgtccgaca tactgagagt taacactgag atcacaaagg cacctctgtc cgcctccatg 960
attaagagat acgatgagca tcaccaggat ctgactttgc tcaaagccct cgttagacag 1020
cagttgccag aaaagtacaa agaaatattc tttgatcaat caaaaaacgg atatgcaggg 1080
tacatcgacg gtggggcaag ccaggaagag ttctacaaat tcatcaaacc tatcctggaa 1140
aagatggatg ggacagaaga gctgctggtt aagctgaata gggaagacct cctcagaaag 1200
cagaggacat ttgataacgg gagcatccct catcaaatcc acctcggtga actccatgct 1260
atcctgagaa ggcaggaaga cttttatcca tttttgaagg acaataggga gaaaatcgaa 1320
aaaatcctga cattcagaat cccatactac gttggtcctc tggcaagagg taacagtagg 1380
ttcgcatgga tgacaaggaa aagcgaggag acaatcacac cctggaattt tgaggaagtt 1440
gttgacaagg gtgccagcgc acaatccttt atcgaaagaa tgacaaattt cgacaagaat 1500
ctgcctaacg aaaaggttct cccaaagcat tcactcctgt acgaatattt tacagtttat 1560
aacgaactga ctaaagttaa atacgttacc gagggtatga ggaagccagc attcctttcc 1620
ggggaacaga agaaagctat tgtggacctc ctgttcaaga caaatagaaa agtgacagtt 1680
aagcaactca aagaggatta cttcaaaaag atcgaatgtt ttgactctgt ggagatcagc 1740
ggggtggagg atagattcaa cgccagcctg ggtacatatc atgatctcct gaaaatcatt 1800
aaagacaagg acttccttga caacgaggag aacgaggaca ttctggaaga cattgttctg 1860
accctcacac tctttgagga tagggagatg attgaggaaa gactgaagac ctacgcccac 1920
ctctttgacg ataaagtgat gaaacagctc aagagaagaa ggtatacagg ttgggggaga 1980
ctgagcagga agttgatcaa tgggattagg gacaaacagt ccgggaaaac aatcctcgat 2040
tttctgaagt cagacggttt cgcaaacaga aattttatgc agctcattca cgatgacagc 2100
ttgacattca aggaagacat ccaaaaggct caagtgagcg gccaagggga tagcctccac 2160
gagcatattg caaatctggc aggttcacca gccatcaaaa agggcatact tcagacagtt 2220
aaggttgtgg acgaattggt taaagttatg ggcaggcata agccagagaa tatcgttatc 2280
gaaatggcaa gggagaacca aacaactcaa aaagggcaga aaaatagcag agagaggatg 2340
aaaagaatcg aggaagggat caaggaactt gggtcccaaa tcctcaagga gcacccagtt 2400
gaaaatactc aactgcaaaa cgagaagctc tatctctact atctccaaaa cgggagggat 2460
atgtatgttg accaggagct ggatattaac agactgtcag attatgatgt tgatcatatc 2520
gtgccccagt cattcctgaa ggacgattcc atcgacaaca aagttctcac aaggtccgat 2580
aaaaacaggg gcaagtccga taacgttcca agcgaagaag tggtgaaaaa gatgaaaaac 2640
tattggagac aacttctgaa tgcaaagttg attactcaga gaaagtttga caacctcaca 2700
aaagcagaaa gaggcgggct tagcgaactc gataaggcag ggtttatcaa aagacagctg 2760
gttgagacaa ggcagatcac aaaacatgtg gcacagatcc ttgactcaag gatgaatacc 2820
aagtatgatg agaatgataa gttgatcagg gaggttaaag ttatcacact caaatccaaa 2880
ctggtgtcag acttcaggaa agactttcaa ttttataagg tgagggagat caataactac 2940
caccatgcac atgacgccta cctgaacgca gtggtgggta cagcattgat taaaaaatac 3000
cctaagctgg agtctgagtt tgtgtacggg gactacaagg tgtacgacgt gaggaaaatg 3060
atagccaagt ccgagcagga gatcgggaaa gcaacagcta agtatttctt ttacagtaat 3120
atcatgaatt tctttaaaac tgagattact ctggcaaacg gggagatcag gaaaagaccc 3180
ctcatcgaga ctaatggtga aacaggtgag atcgtttggg acaaggggag ggattttgct 3240
actgttagaa aagttctgag tatgccacaa gtgaatattg tgaaaaagac agaagttcag 3300
acaggtgggt tctccaaaga atccatcctg cccaagagaa attcagacaa gctcatcgca 3360
agaaagaagg actgggaccc taagaagtac ggaggatttg acagccccac cgtggcctat 3420
tccgtgcttg ttgtggcaaa ggtggagaaa gggaagagca aaaaactgaa atccgtgaaa 3480
gaactgctgg gaattaccat catggaaaga agctcctttg agaagaaccc aatcgacttc 3540
ctggaagcaa aaggatataa ggaagtgaaa aaggacctca ttatcaagct cccaaaatac 3600
tcacttttcg agttggagaa cggtagaaag aggatgctgg caagcgcagg ggaacttcag 3660
aaaggcaatg agctggcatt gccatcaaag tatgtgaact tcctctactt ggccagccat 3720
tacgagaaac ttaaaggtag cccagaagat aacgagcaaa aacagctctt tgtggaacag 3780
cataagcatt atctggatga gatcatagaa caaatctcag agttttccaa gagagttatc 3840
ctcgcagatg caaacctgga taaggttctc tcagcctata ataagcatag agacaagcca 3900
attagagagc aagcagagaa cattatccac ttgttcactc ttacaaacct gggggcacca 3960
gccgccttca aatatttcga tacaacaata gacagaaaga ggtataccag caccaaagaa 4020
gttctcgacg ccacactgat ccatcaatca atcacaggcc tttacgaaac taggatcgac 4080
ttgtcacaac tgggtgggga t 4101
<210> 117
<211> 3307
<212> DNA
<213> 人工序列
<220>
<223> 合成的Cas9
<400> 117
gagcaaggac acctacgacg acgacttgga caacctattg gcccagatag gtgaccagta 60
tgcagacctc ttccttgcgg ccaagaactt gagtgacgct atactgctca gtgacatcct 120
gagggtgaac actgagatca ctaaggcccc tctctctgcc tcaatgatta agcgttacga 180
cgagcatcac caggatctca ccctgcttaa ggcccttgtt cggcagcagc tccctgagaa 240
gtacaaggag atattttttg accagtctaa gaacggctac gccggttaca ttgacggtgg 300
ggcaagccag gaggagttct acaagttcat caagccgatc cttgagaaga tggacggcac 360
cgaggagcta cttgtcaagt tgaaccggga agacctgctc cggaaacagc gtacattcga 420
caacggcagc atccctcacc agatccacct gggcgaacta cacgccatcc tccgacgtca 480
ggaggacttc tatccattct tgaaagataa cagggaaaaa atcgaaaaaa tacttacgtt 540
tcgaatacct tactacgtgg ggccccttgc tcggggaaac tccagattcg catggatgac 600
caggaagtca gaggagacca tcacaccctg gaactttgag gaggtggttg acaaaggtgc 660
ttctgcccag tccttcattg agcggatgac taacttcgac aagaacctgc ccaacgagaa 720
ggtgctgcca aagcacagcc tgctctacga atactttact gtgtacaatg agctgacgaa 780
ggtgaagtac gtgacagagg ggatgcggaa gcccgctttc ctgagcggcg agcaaaaaaa 840
agcaatcgtg gacctactgt tcaagaccaa ccgaaaggtg acagtgaagc agctcaagga 900
ggactacttc aaaaaaatcg agtgcttcga ctctgttgag ataagcggcg tggaggaccg 960
attcaacgcc tcattgggaa cctatcacga cctgctcaag atcattaagg acaaggactt 1020
cctggataat gaggagaatg aggacatcct ggaggatatt gtgctgaccc ttactctatt 1080
cgaggacagg gagatgatcg aggagcgact caagacctac gctcacctgt tcgacgacaa 1140
ggttatgaag caattgaagc gtaggcgata cacggggtgg ggaagactct cccgaaaact 1200
gataaacggc atcagggaca agcagtcagg gaagacgatc ttggacttcc tgaaatccga 1260
cgggttcgcc aaccgcaact tcatgcagct cattcacgac gactcactaa cgttcaaaga 1320
ggacattcag aaggctcaag tcagtggaca aggcgactcc ctgcacgagc acattgcaaa 1380
ccttgcgggc tccccggcga ttaaaaaggg cattctccaa acggttaagg tggtggacga 1440
gctggtgaag gtgatgggcc gacacaagcc tgagaacatc gtgatcgaga tggccaggga 1500
gaaccagact acccagaagg gtcagaagaa ctctcgggaa cgtatgaagc gtattgagga 1560
ggggattaag gagttgggct ctcaaatcct caaggagcac cctgtggaga acactcagct 1620
ccaaaacgag aagctgtacc tgtactacct gcaaaacggg cgcgatatgt acgtggatca 1680
ggagttggac atcaacaggc ttagcgatta cgacgtggac cacatcgtgc cacagtcatt 1740
cttaaaggac gacagcatcg acaacaaggt tctgacgagg agcgacaaga atcgagggaa 1800
aagtgacaat gttccatccg aggaggtggt caagaaaatg aagaactatt ggcgtcagct 1860
tctgaacgcc aagctcatca cccagcggaa attcgacaac ctgactaagg ctgagcgagg 1920
cggactctcc gagcttgaca aggctggctt catcaagcgg cagttggtcg aaacccgaca 1980
gataacgaag cacgttgccc agatacttga ctcccgtatg aacaccaagt acgacgagaa 2040
cgacaagctc atcagggagg tgaaggtcat tacccttaag tccaaactcg tcagcgactt 2100
tcgtaaggac ttccagttct acaaggtgcg cgagatcaat aactaccacc acgcacacga 2160
cgcctacctg aacgcagtgg ttggaaccgc gttgattaaa aagtacccca agttggagtc 2220
ggagttcgtt tacggggact acaaggtgta cgacgttcgg aagatgatcg ccaagtctga 2280
acaggagatc gggaaagcaa ccgccaagta tttcttctat agcaacatca tgaacttctt 2340
taaaaccgag atcacacttg ccaatggcga gatccgtaag aggccgctga tcgagacaaa 2400
tggggagact ggcgagatcg tgtgggacaa gggccgcgac ttcgcaaccg ttcggaaagt 2460
cttgtccatg cctcaagtca acatcgtcaa gaagactgag gtgcaaacag gcgggttctc 2520
gaaggagtcc atactgccca agaggaactc agacaagctc atagcacgca aaaaagactg 2580
ggatccaaag aaatacggcg ggttcgactc gccgacagtc gcatactccg tgttagtggt 2640
ggctaaagtg gaaaagggga agtccaagaa gctcaagtcc gtcaaggagt tgctcgggat 2700
caccattatg gaacggtcct cattcgagaa gaatcccatt gacttcctag aggcgaaggg 2760
ctacaaagag gtcaaaaagg acctaattat taagctcccc aagtattcac tcttcgaact 2820
tgaaaatggt cgtaagcgga tgttggcaag cgctggagag cttcagaagg ggaacgagct 2880
tgcactgcct tccaagtacg tgaacttcct gtacctcgcc tctcattacg agaagttgaa 2940
gggctcaccg gaggacaacg agcagaagca gttgttcgtg gagcagcaca agcactacct 3000
cgacgagatc attgagcaga taagtgagtt cagcaaacgg gtgatccttg ccgacgctaa 3060
cctggacaag gtgctgagcg cctacaacaa gcacagagac aagccgatcc gagagcaagc 3120
ggagaacatc atacacctgt tcaccctcac gaacctcggg gctcccgcag ccttcaaata 3180
ttttgacacg accatcgacc gtaaacgcta cactagcacg aaggaggtgc tggacgctac 3240
ccttatccac cagtccatca ccggcctgta cgagacgaga atcgacttgt cgcagctcgg 3300
tggtgac 3307
<210> 118
<211> 4101
<212> DNA
<213> 人工序列
<220>
<223> 合成的Cas9
<400> 118
gacaaaaaat actcaattgg tctggcaatt gggaccaaca gtgtcggatg ggccgtgatt 60
accgacgagt acaaggtgcc gtccaaaaaa ttcaaggtgc ttgggaacac cgaccgccac 120
tcgatcaaga aaaacctaat cggtgcgttg cttttcgaca gtggggagac cgccgaggca 180
acacgcttaa aacgcacagc taggaggaga tatacacggc gcaagaaccg aatatgctac 240
ttacaggaga tattctccaa tgagatggcg aaggtggacg actctttctt ccatcggctt 300
gaggaatcct tcctggtcga ggaggacaag aagcacgagc gacacccgat attcgggaac 360
atcgttgatg aggtggcgta ccacgagaag tacccaacga tataccactt acgcaagaag 420
ctcgtggact ctacggacaa ggccgacttg cgccttatct acttggcact ggcccacatg 480
attaagttcc gaggccactt ccttatcgag ggtgacctga accccgataa ctccgacgtg 540
gacaagctct tcatccaact cgtccagaca tacaaccagc tattcgagga gaatcctatc 600
aacgcctctg gggtggacgc taaagctatc ctctcagccc gcctgtcaaa gtcgaggagg 660
ttggagaacc taatcgccca gcttccaggc gagaagaaaa atgggctgtt cggaaacctt 720
atcgcactct cactgggcct aaccccgaac ttcaagtcca acttcgacct ggcagaggac 780
gcgaaattgc agttgtcgaa agacacctat gacgatgacc tggacaacct gttggcccag 840
ataggggacc agtacgccga cctgttccta gcggccaaga acctgtccga cgccatcttg 900
ctgtcggata tactgcgggt gaacaccgag atcactaaag cacctctctc cgccagcatg 960
attaagcgtt acgacgagca ccaccaagat ttgaccctgc taaaggcact tgtacggcag 1020
cagcttcccg agaagtacaa ggagatcttt ttcgaccaaa gcaagaacgg ctacgccggg 1080
tacatcgacg gaggtgccag ccaggaggag ttctacaagt tcattaagcc catcctggag 1140
aagatggacg ggactgagga actacttgtg aagctgaacc gggaagactt actacggaag 1200
cagcgtacct tcgacaacgg ttctatccca catcagatcc atcttgggga gttgcacgcg 1260
atcctgcgac gccaggagga cttttacccc ttcctgaaag acaaccgcga gaaaatcgag 1320
aagatactga ccttcagaat accttactac gtcggacccc ttgcgcgagg caactcaaga 1380
ttcgcgtgga tgaccaggaa atcagaggag accatcacac cctggaattt cgaggaggtg 1440
gttgacaagg gtgcctccgc ccagtccttt atcgaacgaa tgaccaactt cgacaagaac 1500
ttgcccaacg agaaggtgct ccccaaacac agcctcctct acgaatattt cacagtgtac 1560
aacgagctta ctaaagttaa gtatgttact gagggcatga ggaaacccgc cttcctgtca 1620
ggcgagcaga agaaagctat tgtggacctc cttttcaaga ccaaccggaa ggtgacagtg 1680
aagcagctca aggaggacta cttcaagaag atagagtgct tcgacagcgt ggagatcagc 1740
ggggtggagg acagattcaa tgcctctctc ggaacatacc acgacttgct taagatcatc 1800
aaggacaagg acttcctcga caacgaggaa aacgaggata ttctggagga tattgttctg 1860
actcttaccc tgttcgagga ccgggagatg atcgaggagc gtctcaagac ctacgcccac 1920
ctgttcgacg acaaagttat gaagcagctc aagcgtcgga gatataccgg atggggccgt 1980
ctgtctcgga agctcatcaa cgggatcagg gacaagcagt cagggaagac gatcttagac 2040
ttccttaagt ctgacggctt cgccaacagg aacttcatgc agttgatcca cgacgacagc 2100
cttaccttca aggaggacat ccagaaggcc caagtgagtg gccagggtga cagcctccac 2160
gagcatattg ctaatcttgc gggttcccca gcgattaaaa agggcatact tcaaaccgtt 2220
aaggtggtgg acgagcttgt caaggtgatg gggcgacaca agcccgagaa catcgtgatc 2280
gagatggcca gggagaacca gaccacccag aaggggcaga agaatagccg agaacgcatg 2340
aagcgcatcg aggaggggat taaggagcta gggagccaga tcctcaagga acatcccgtc 2400
gagaacaccc agctccagaa cgagaagcta tacctctact acttgcaaaa cgggagggat 2460
atgtacgtgg atcaggagtt ggacattaac cgcctaagcg actacgacgt agatcacatc 2520
gtgcctcagt cattcctcaa agacgacagc attgacaaca aagtcttgac ccgatccgac 2580
aagaaccgag gaaaatccga caatgtgccc tcagaggagg tcgtcaagaa aatgaagaac 2640
tattggaggc agctacttaa cgccaaactc ataacccagc ggaagttcga caacctgaca 2700
aaggctgagc ggggtgggct cagcgagctt gacaaggctg gcttcatcaa gcggcagttg 2760
gtggagacaa gacagataac gaagcacgtg gctcagatcc tggactctcg catgaacacg 2820
aagtacgacg agaacgacaa attgatccgc gaggtcaagg ttattacgct caagagcaaa 2880
cttgtcagcg atttccgcaa ggacttccag ttctacaagg tgagggagat taacaactac 2940
caccatgcac atgatgccta cttgaacgca gtggtgggga ccgcgcttat taaaaagtac 3000
cctaagttgg agtcagagtt cgtttatggg gactacaagg tgtacgacgt ccggaagatg 3060
attgcaaagt ctgaacagga aatcgggaag gccaccgcca aatatttctt ctacagtaac 3120
attatgaatt tttttaagac tgaaattact ctcgcaaacg gcgagatcag gaagcgtccc 3180
ctcatcgaga caaacgggga gaccggggag atagtctggg acaaggggcg ggacttcgct 3240
acggtgagga aggtgctctc gatgccacaa gtgaacatcg tcaaaaagac agaggtgcag 3300
accggtggct tctcaaagga gtcaatcctg ccaaaacgta acagcgacaa gctcatcgcc 3360
cgcaagaaag actgggaccc taagaagtat ggtgggttcg actcaccgac ggtcgcatac 3420
tccgttctgg tcgtggcaaa ggtggaaaag ggcaagtcca aaaaactgaa atccgtgaag 3480
gagttgcttg gcattaccat catggaacgc agcagcttcg agaagaaccc cattgacttc 3540
ctggaggcta aagggtacaa ggaggtcaag aaagatttaa ttattaagct acctaagtac 3600
agcttgttcg agctggagaa cggccgaaaa cgaatgctcg catccgccgg ggaacttcaa 3660
aagggcaacg agcttgcgct gccctccaag tacgtgaact tcctgtactt ggcatcccac 3720
tacgagaaac tcaagggtag cccagaggac aacgagcaga agcagctatt cgtggagcag 3780
cacaagcact acctcgacga gataatcgag cagatcagtg agttcagtaa gcgggtgata 3840
ctcgcggacg ccaacttgga caaggtgctt agtgcctaca acaagcaccg tgacaagccc 3900
atccgagaac aggctgagaa catcatccac cttttcactc tgacaaacct cggtgctccc 3960
gccgccttca aatacttcga cactaccatc gacaggaagc gctacacatc tacgaaggaa 4020
gttcttgacg ctacgcttat tcatcagtct atcacagggc tgtacgagac aaggatcgac 4080
cttagccaac tcggcgggga t 4101
<210> 119
<211> 4101
<212> DNA
<213> 人工序列
<220>
<223> 合成的Cas9
<400> 119
gacaagaagt acagcatcgg cctggacatc ggcaccaact ctgtgggctg ggccgtgatc 60
accgacgagt acaaggtgcc cagcaagaaa ttcaaggtgc tgggcaacac cgaccggcac 120
agcatcaaga agaacctgat cggagccctg ctgttcgaca gcggcgaaac agccgaggcc 180
acccggctga agagaaccgc cagaagaaga tacaccagac ggaagaaccg gatctgctat 240
ctgcaagaga tcttcagcaa cgagatggcc aaggtggacg acagcttctt ccacagactg 300
gaagagtcct tcctggtgga agaggataag aagcacgagc ggcaccccat cttcggcaac 360
atcgtggacg aggtggccta ccacgagaag taccccacca tctaccacct gagaaagaaa 420
ctggtggaca gcaccgacaa ggccgacctg cggctgatct atctggccct ggcccacatg 480
atcaagttcc ggggccactt cctgatcgag ggcgacctga accccgacaa cagcgacgtg 540
gacaagctgt tcatccagct ggtgcagacc tacaaccagc tgttcgagga aaaccccatc 600
aacgccagcg gcgtggacgc caaggccatc ctgtctgcca gactgagcaa gagcagacgg 660
ctggaaaatc tgatcgccca gctgcccggc gagaagaaga atggcctgtt cggaaacctg 720
attgccctga gcctgggcct gacccccaac ttcaagagca acttcgacct ggccgaggat 780
gccaaactgc agctgagcaa ggacacctac gacgacgacc tggacaacct gctggcccag 840
atcggcgacc agtacgccga cctgtttctg gccgccaaga acctgtccga cgccatcctg 900
ctgagcgaca tcctgagagt gaacaccgag atcaccaagg cccccctgag cgcctctatg 960
atcaagagat acgacgagca ccaccaggac ctgaccctgc tgaaagctct cgtgcggcag 1020
cagctgcctg agaagtacaa agagattttc ttcgaccaga gcaagaacgg ctacgccggc 1080
tacattgacg gcggagccag ccaggaagag ttctacaagt tcatcaagcc catcctggaa 1140
aagatggacg gcaccgagga actgctcgtg aagctgaaca gagaggacct gctgcggaag 1200
cagcggacct tcgacaacgg cagcatcccc caccagatcc acctgggaga gctgcacgcc 1260
attctgcggc ggcaggaaga tttttaccca ttcctgaagg acaaccggga aaagatcgag 1320
aagatcctga ccttccgcat cccctactac gtgggccctc tggccagggg aaacagcaga 1380
ttcgcctgga tgaccagaaa gagcgaggaa accatcaccc cctggaactt cgaggaagtg 1440
gtggacaagg gcgcttccgc ccagagcttc atcgagcgga tgaccaactt cgataagaac 1500
ctgcccaacg agaaggtgct gcccaagcac agcctgctgt acgagtactt caccgtgtat 1560
aacgagctga ccaaagtgaa atacgtgacc gagggaatga gaaagcccgc cttcctgagc 1620
ggcgagcaga aaaaggccat cgtggacctg ctgttcaaga ccaaccggaa agtgaccgtg 1680
aagcagctga aagaggacta cttcaagaaa atcgagtgct tcgactccgt ggaaatctcc 1740
ggcgtggaag atcggttcaa cgcctccctg ggcacatacc acgatctgct gaaaattatc 1800
aaggacaagg acttcctgga caatgaggaa aacgaggaca ttctggaaga tatcgtgctg 1860
accctgacac tgtttgagga cagagagatg atcgaggaac ggctgaaaac ctatgcccac 1920
ctgttcgacg acaaagtgat gaagcagctg aagcggcgga gatacaccgg ctggggcagg 1980
ctgagccgga agctgatcaa cggcatccgg gacaagcagt ccggcaagac aatcctggat 2040
ttcctgaagt ccgacggctt cgccaacaga aacttcatgc agctgatcca cgacgacagc 2100
ctgaccttta aagaggacat ccagaaagcc caggtgtccg gccagggcga tagcctgcac 2160
gagcacattg ccaatctggc cggcagcccc gccattaaga agggcatcct gcagacagtg 2220
aaggtggtgg acgagctcgt gaaagtgatg ggccggcaca agcccgagaa catcgtgatc 2280
gaaatggcca gagagaacca gaccacccag aagggacaga agaacagccg cgagagaatg 2340
aagcggatcg aagagggcat caaagagctg ggcagccaga tcctgaaaga acaccccgtg 2400
gaaaacaccc agctgcagaa cgagaagctg tacctgtact acctgcagaa tgggcgggat 2460
atgtacgtgg accaggaact ggacatcaac cggctgtccg actacgatgt ggaccatatc 2520
gtgcctcaga gctttctgaa ggacgactcc atcgacaaca aggtgctgac cagaagcgac 2580
aagaaccggg gcaagagcga caacgtgccc tccgaagagg tcgtgaagaa gatgaagaac 2640
tactggcggc agctgctgaa cgccaagctg attacccaga gaaagttcga caatctgacc 2700
aaggccgaga gaggcggcct gagcgaactg gataaggccg gcttcatcaa gagacagctg 2760
gtggaaaccc ggcagatcac aaagcacgtg gcacagatcc tggactcccg gatgaacact 2820
aagtacgacg agaatgacaa gctgatccgg gaagtgaaag tgatcaccct gaagtccaag 2880
ctggtgtccg atttccggaa ggatttccag ttttacaaag tgcgcgagat caacaactac 2940
caccacgccc acgacgccta cctgaacgcc gtcgtgggaa ccgccctgat caaaaagtac 3000
cctaagctgg aaagcgagtt cgtgtacggc gactacaagg tgtacgacgt gcggaagatg 3060
atcgccaaga gcgagcagga aatcggcaag gctaccgcca agtacttctt ctacagcaac 3120
atcatgaact ttttcaagac cgagattacc ctggccaacg gcgagatccg gaagcggcct 3180
ctgatcgaga caaacggcga aaccggggag atcgtgtggg ataagggccg ggattttgcc 3240
accgtgcgga aagtgctgag catgccccaa gtgaatatcg tgaaaaagac cgaggtgcag 3300
acaggcggct tcagcaaaga gtctatcctg cccaagagga acagcgataa gctgatcgcc 3360
agaaagaagg actgggaccc taagaagtac ggcggcttcg acagccccac cgtggcctat 3420
tctgtgctgg tggtggccaa agtggaaaag ggcaagtcca agaaactgaa gagtgtgaaa 3480
gagctgctgg ggatcaccat catggaaaga agcagcttcg agaagaatcc catcgacttt 3540
ctggaagcca agggctacaa agaagtgaaa aaggacctga tcatcaagct gcctaagtac 3600
tccctgttcg agctggaaaa cggccggaag agaatgctgg cctctgccgg cgaactgcag 3660
aagggaaacg aactggccct gccctccaaa tatgtgaact tcctgtacct ggccagccac 3720
tatgagaagc tgaagggctc ccccgaggat aatgagcaga aacagctgtt tgtggaacag 3780
cacaagcact acctggacga gatcatcgag cagatcagcg agttctccaa gagagtgatc 3840
ctggccgacg ctaatctgga caaagtgctg tccgcctaca acaagcaccg ggataagccc 3900
atcagagagc aggccgagaa tatcatccac ctgtttaccc tgaccaatct gggagcccct 3960
gccgccttca agtactttga caccaccatc gaccggaaga ggtacaccag caccaaagag 4020
gtgctggacg ccaccctgat ccaccagagc atcaccggcc tgtacgagac acggatcgac 4080
ctgtctcagc tgggaggtga c 4101
<210> 120
<211> 4101
<212> DNA
<213> 人工序列
<220>
<223> 合成的eCas9
<400> 120
gacaagaagt acagcatcgg cctggacatc ggcaccaact ctgtgggctg ggccgtgatc 60
accgacgagt acaaggtgcc cagcaagaaa ttcaaggtgc tgggcaacac cgaccggcac 120
agcatcaaga agaacctgat cggagccctg ctgttcgaca gcggcgaaac agccgaggcc 180
acccggctga agagaaccgc cagaagaaga tacaccagac ggaagaaccg gatctgctat 240
ctgcaagaga tcttcagcaa cgagatggcc aaggtggacg acagcttctt ccacagactg 300
gaagagtcct tcctggtgga agaggataag aagcacgagc ggcaccccat cttcggcaac 360
atcgtggacg aggtggccta ccacgagaag taccccacca tctaccacct gagaaagaaa 420
ctggtggaca gcaccgacaa ggccgacctg cggctgatct atctggccct ggcccacatg 480
atcaagttcc ggggccactt cctgatcgag ggcgacctga accccgacaa cagcgacgtg 540
gacaagctgt tcatccagct ggtgcagacc tacaaccagc tgttcgagga aaaccccatc 600
aacgccagcg gcgtggacgc caaggccatc ctgtctgcca gactgagcaa gagcagacgg 660
ctggaaaatc tgatcgccca gctgcccggc gagaagaaga atggcctgtt cggaaacctg 720
attgccctga gcctgggcct gacccccaac ttcaagagca acttcgacct ggccgaggat 780
gccaaactgc agctgagcaa ggacacctac gacgacgacc tggacaacct gctggcccag 840
atcggcgacc agtacgccga cctgtttctg gccgccaaga acctgtccga cgccatcctg 900
ctgagcgaca tcctgagagt gaacaccgag atcaccaagg cccccctgag cgcctctatg 960
atcaagagat acgacgagca ccaccaggac ctgaccctgc tgaaagctct cgtgcggcag 1020
cagctgcctg agaagtacaa agagattttc ttcgaccaga gcaagaacgg ctacgccggc 1080
tacattgacg gcggagccag ccaggaagag ttctacaagt tcatcaagcc catcctggaa 1140
aagatggacg gcaccgagga actgctcgtg aagctgaaca gagaggacct gctgcggaag 1200
cagcggacct tcgacaacgg cagcatcccc caccagatcc acctgggaga gctgcacgcc 1260
attctgcggc ggcaggaaga tttttaccca ttcctgaagg acaaccggga aaagatcgag 1320
aagatcctga ccttccgcat cccctactac gtgggccctc tggccagggg aaacagcaga 1380
ttcgcctgga tgaccagaaa gagcgaggaa accatcaccc cctggaactt cgaggaagtg 1440
gtggacaagg gcgcttccgc ccagagcttc atcgagcgga tgaccaactt cgataagaac 1500
ctgcccaacg agaaggtgct gcccaagcac agcctgctgt acgagtactt caccgtgtat 1560
aacgagctga ccaaagtgaa atacgtgacc gagggaatga gaaagcccgc cttcctgagc 1620
ggcgagcaga aaaaggccat cgtggacctg ctgttcaaga ccaaccggaa agtgaccgtg 1680
aagcagctga aagaggacta cttcaagaaa atcgagtgct tcgactccgt ggaaatctcc 1740
ggcgtggaag atcggttcaa cgcctccctg ggcacatacc acgatctgct gaaaattatc 1800
aaggacaagg acttcctgga caatgaggaa aacgaggaca ttctggaaga tatcgtgctg 1860
accctgacac tgtttgagga cagagagatg atcgaggaac ggctgaaaac ctatgcccac 1920
ctgttcgacg acaaagtgat gaagcagctg aagcggcgga gatacaccgg ctggggcagg 1980
ctgagccgga agctgatcaa cggcatccgg gacaagcagt ccggcaagac aatcctggat 2040
ttcctgaagt ccgacggctt cgccaacaga aacttcatgc agctgatcca cgacgacagc 2100
ctgaccttta aagaggacat ccagaaagcc caggtgtccg gccagggcga tagcctgcac 2160
gagcacattg ccaatctggc cggcagcccc gccattaaga agggcatcct gcagacagtg 2220
aaggtggtgg acgagctcgt gaaagtgatg ggccggcaca agcccgagaa catcgtgatc 2280
gaaatggcca gagagaacca gaccacccag aagggacaga agaacagccg cgagagaatg 2340
aagcggatcg aagagggcat caaagagctg ggcagccaga tcctgaaaga acaccccgtg 2400
gaaaacaccc agctgcagaa cgagaagctg tacctgtact acctgcagaa tgggcgggat 2460
atgtacgtgg accaggaact ggacatcaac cggctgtccg actacgatgt ggaccatatc 2520
gtgcctcaga gctttctggc cgacgactcc atcgacaaca aggtgctgac cagaagcgac 2580
aagaaccggg gcaagagcga caacgtgccc tccgaagagg tcgtgaagaa gatgaagaac 2640
tactggcggc agctgctgaa cgccaagctg attacccaga gaaagttcga caatctgacc 2700
aaggccgaga gaggcggcct gagcgaactg gataaggccg gcttcatcaa gagacagctg 2760
gtggaaaccc ggcagatcac aaagcacgtg gcacagatcc tggactcccg gatgaacact 2820
aagtacgacg agaatgacaa gctgatccgg gaagtgaaag tgatcaccct gaagtccaag 2880
ctggtgtccg atttccggaa ggatttccag ttttacaaag tgcgcgagat caacaactac 2940
caccacgccc acgacgccta cctgaacgcc gtcgtgggaa ccgccctgat caaaaagtac 3000
cctgccctgg aaagcgagtt cgtgtacggc gactacaagg tgtacgacgt gcggaagatg 3060
atcgccaaga gcgagcagga aatcggcaag gctaccgcca agtacttctt ctacagcaac 3120
atcatgaact ttttcaagac cgagattacc ctggccaacg gcgagatccg gaaggcccct 3180
ctgatcgaga caaacggcga aaccggggag atcgtgtggg ataagggccg ggattttgcc 3240
accgtgcgga aagtgctgag catgccccaa gtgaatatcg tgaaaaagac cgaggtgcag 3300
acaggcggct tcagcaaaga gtctatcctg cccaagagga acagcgataa gctgatcgcc 3360
agaaagaagg actgggaccc taagaagtac ggcggcttcg acagccccac cgtggcctat 3420
tctgtgctgg tggtggccaa agtggaaaag ggcaagtcca agaaactgaa gagtgtgaaa 3480
gagctgctgg ggatcaccat catggaaaga agcagcttcg agaagaatcc catcgacttt 3540
ctggaagcca agggctacaa agaagtgaaa aaggacctga tcatcaagct gcctaagtac 3600
tccctgttcg agctggaaaa cggccggaag agaatgctgg cctctgccgg cgaactgcag 3660
aagggaaacg aactggccct gccctccaaa tatgtgaact tcctgtacct ggccagccac 3720
tatgagaagc tgaagggctc ccccgaggat aatgagcaga aacagctgtt tgtggaacag 3780
cacaagcact acctggacga gatcatcgag cagatcagcg agttctccaa gagagtgatc 3840
ctggccgacg ctaatctgga caaagtgctg tccgcctaca acaagcaccg ggataagccc 3900
atcagagagc aggccgagaa tatcatccac ctgtttaccc tgaccaatct gggagcccct 3960
gccgccttca agtactttga caccaccatc gaccggaaga ggtacaccag caccaaagag 4020
gtgctggacg ccaccctgat ccaccagagc atcaccggcc tgtacgagac acggatcgac 4080
ctgtctcagc tgggaggtga c 4101
<210> 121
<211> 4101
<212> DNA
<213> 人工序列
<220>
<223> 合成的nCas9(D10A)
<400> 121
gacaagaagt acagcatcgg cctggccatc ggcaccaact ctgtgggctg ggccgtgatc 60
accgacgagt acaaggtgcc cagcaagaaa ttcaaggtgc tgggcaacac cgaccggcac 120
agcatcaaga agaacctgat cggagccctg ctgttcgaca gcggcgaaac agccgaggcc 180
acccggctga agagaaccgc cagaagaaga tacaccagac ggaagaaccg gatctgctat 240
ctgcaagaga tcttcagcaa cgagatggcc aaggtggacg acagcttctt ccacagactg 300
gaagagtcct tcctggtgga agaggataag aagcacgagc ggcaccccat cttcggcaac 360
atcgtggacg aggtggccta ccacgagaag taccccacca tctaccacct gagaaagaaa 420
ctggtggaca gcaccgacaa ggccgacctg cggctgatct atctggccct ggcccacatg 480
atcaagttcc ggggccactt cctgatcgag ggcgacctga accccgacaa cagcgacgtg 540
gacaagctgt tcatccagct ggtgcagacc tacaaccagc tgttcgagga aaaccccatc 600
aacgccagcg gcgtggacgc caaggccatc ctgtctgcca gactgagcaa gagcagacgg 660
ctggaaaatc tgatcgccca gctgcccggc gagaagaaga atggcctgtt cggaaacctg 720
attgccctga gcctgggcct gacccccaac ttcaagagca acttcgacct ggccgaggat 780
gccaaactgc agctgagcaa ggacacctac gacgacgacc tggacaacct gctggcccag 840
atcggcgacc agtacgccga cctgtttctg gccgccaaga acctgtccga cgccatcctg 900
ctgagcgaca tcctgagagt gaacaccgag atcaccaagg cccccctgag cgcctctatg 960
atcaagagat acgacgagca ccaccaggac ctgaccctgc tgaaagctct cgtgcggcag 1020
cagctgcctg agaagtacaa agagattttc ttcgaccaga gcaagaacgg ctacgccggc 1080
tacattgacg gcggagccag ccaggaagag ttctacaagt tcatcaagcc catcctggaa 1140
aagatggacg gcaccgagga actgctcgtg aagctgaaca gagaggacct gctgcggaag 1200
cagcggacct tcgacaacgg cagcatcccc caccagatcc acctgggaga gctgcacgcc 1260
attctgcggc ggcaggaaga tttttaccca ttcctgaagg acaaccggga aaagatcgag 1320
aagatcctga ccttccgcat cccctactac gtgggccctc tggccagggg aaacagcaga 1380
ttcgcctgga tgaccagaaa gagcgaggaa accatcaccc cctggaactt cgaggaagtg 1440
gtggacaagg gcgcttccgc ccagagcttc atcgagcgga tgaccaactt cgataagaac 1500
ctgcccaacg agaaggtgct gcccaagcac agcctgctgt acgagtactt caccgtgtat 1560
aacgagctga ccaaagtgaa atacgtgacc gagggaatga gaaagcccgc cttcctgagc 1620
ggcgagcaga aaaaggccat cgtggacctg ctgttcaaga ccaaccggaa agtgaccgtg 1680
aagcagctga aagaggacta cttcaagaaa atcgagtgct tcgactccgt ggaaatctcc 1740
ggcgtggaag atcggttcaa cgcctccctg ggcacatacc acgatctgct gaaaattatc 1800
aaggacaagg acttcctgga caatgaggaa aacgaggaca ttctggaaga tatcgtgctg 1860
accctgacac tgtttgagga cagagagatg atcgaggaac ggctgaaaac ctatgcccac 1920
ctgttcgacg acaaagtgat gaagcagctg aagcggcgga gatacaccgg ctggggcagg 1980
ctgagccgga agctgatcaa cggcatccgg gacaagcagt ccggcaagac aatcctggat 2040
ttcctgaagt ccgacggctt cgccaacaga aacttcatgc agctgatcca cgacgacagc 2100
ctgaccttta aagaggacat ccagaaagcc caggtgtccg gccagggcga tagcctgcac 2160
gagcacattg ccaatctggc cggcagcccc gccattaaga agggcatcct gcagacagtg 2220
aaggtggtgg acgagctcgt gaaagtgatg ggccggcaca agcccgagaa catcgtgatc 2280
gaaatggcca gagagaacca gaccacccag aagggacaga agaacagccg cgagagaatg 2340
aagcggatcg aagagggcat caaagagctg ggcagccaga tcctgaaaga acaccccgtg 2400
gaaaacaccc agctgcagaa cgagaagctg tacctgtact acctgcagaa tgggcgggat 2460
atgtacgtgg accaggaact ggacatcaac cggctgtccg actacgatgt ggaccatatc 2520
gtgcctcaga gctttctgaa ggacgactcc atcgacaaca aggtgctgac cagaagcgac 2580
aagaaccggg gcaagagcga caacgtgccc tccgaagagg tcgtgaagaa gatgaagaac 2640
tactggcggc agctgctgaa cgccaagctg attacccaga gaaagttcga caatctgacc 2700
aaggccgaga gaggcggcct gagcgaactg gataaggccg gcttcatcaa gagacagctg 2760
gtggaaaccc ggcagatcac aaagcacgtg gcacagatcc tggactcccg gatgaacact 2820
aagtacgacg agaatgacaa gctgatccgg gaagtgaaag tgatcaccct gaagtccaag 2880
ctggtgtccg atttccggaa ggatttccag ttttacaaag tgcgcgagat caacaactac 2940
caccacgccc acgacgccta cctgaacgcc gtcgtgggaa ccgccctgat caaaaagtac 3000
cctaagctgg aaagcgagtt cgtgtacggc gactacaagg tgtacgacgt gcggaagatg 3060
atcgccaaga gcgagcagga aatcggcaag gctaccgcca agtacttctt ctacagcaac 3120
atcatgaact ttttcaagac cgagattacc ctggccaacg gcgagatccg gaagcggcct 3180
ctgatcgaga caaacggcga aaccggggag atcgtgtggg ataagggccg ggattttgcc 3240
accgtgcgga aagtgctgag catgccccaa gtgaatatcg tgaaaaagac cgaggtgcag 3300
acaggcggct tcagcaaaga gtctatcctg cccaagagga acagcgataa gctgatcgcc 3360
agaaagaagg actgggaccc taagaagtac ggcggcttcg acagccccac cgtggcctat 3420
tctgtgctgg tggtggccaa agtggaaaag ggcaagtcca agaaactgaa gagtgtgaaa 3480
gagctgctgg ggatcaccat catggaaaga agcagcttcg agaagaatcc catcgacttt 3540
ctggaagcca agggctacaa agaagtgaaa aaggacctga tcatcaagct gcctaagtac 3600
tccctgttcg agctggaaaa cggccggaag agaatgctgg cctctgccgg cgaactgcag 3660
aagggaaacg aactggccct gccctccaaa tatgtgaact tcctgtacct ggccagccac 3720
tatgagaagc tgaagggctc ccccgaggat aatgagcaga aacagctgtt tgtggaacag 3780
cacaagcact acctggacga gatcatcgag cagatcagcg agttctccaa gagagtgatc 3840
ctggccgacg ctaatctgga caaagtgctg tccgcctaca acaagcaccg ggataagccc 3900
atcagagagc aggccgagaa tatcatccac ctgtttaccc tgaccaatct gggagcccct 3960
gccgccttca agtactttga caccaccatc gaccggaaga ggtacaccag caccaaagag 4020
gtgctggacg ccaccctgat ccaccagagc atcaccggcc tgtacgagac acggatcgac 4080
ctgtctcagc tgggaggtga c 4101
<210> 122
<211> 4101
<212> DNA
<213> 人工序列
<220>
<223> 合成的nCas9(H840A)
<400> 122
gacaagaagt acagcatcgg cctggacatc ggcaccaact ctgtgggctg ggccgtgatc 60
accgacgagt acaaggtgcc cagcaagaaa ttcaaggtgc tgggcaacac cgaccggcac 120
agcatcaaga agaacctgat cggagccctg ctgttcgaca gcggcgaaac agccgaggcc 180
acccggctga agagaaccgc cagaagaaga tacaccagac ggaagaaccg gatctgctat 240
ctgcaagaga tcttcagcaa cgagatggcc aaggtggacg acagcttctt ccacagactg 300
gaagagtcct tcctggtgga agaggataag aagcacgagc ggcaccccat cttcggcaac 360
atcgtggacg aggtggccta ccacgagaag taccccacca tctaccacct gagaaagaaa 420
ctggtggaca gcaccgacaa ggccgacctg cggctgatct atctggccct ggcccacatg 480
atcaagttcc ggggccactt cctgatcgag ggcgacctga accccgacaa cagcgacgtg 540
gacaagctgt tcatccagct ggtgcagacc tacaaccagc tgttcgagga aaaccccatc 600
aacgccagcg gcgtggacgc caaggccatc ctgtctgcca gactgagcaa gagcagacgg 660
ctggaaaatc tgatcgccca gctgcccggc gagaagaaga atggcctgtt cggaaacctg 720
attgccctga gcctgggcct gacccccaac ttcaagagca acttcgacct ggccgaggat 780
gccaaactgc agctgagcaa ggacacctac gacgacgacc tggacaacct gctggcccag 840
atcggcgacc agtacgccga cctgtttctg gccgccaaga acctgtccga cgccatcctg 900
ctgagcgaca tcctgagagt gaacaccgag atcaccaagg cccccctgag cgcctctatg 960
atcaagagat acgacgagca ccaccaggac ctgaccctgc tgaaagctct cgtgcggcag 1020
cagctgcctg agaagtacaa agagattttc ttcgaccaga gcaagaacgg ctacgccggc 1080
tacattgacg gcggagccag ccaggaagag ttctacaagt tcatcaagcc catcctggaa 1140
aagatggacg gcaccgagga actgctcgtg aagctgaaca gagaggacct gctgcggaag 1200
cagcggacct tcgacaacgg cagcatcccc caccagatcc acctgggaga gctgcacgcc 1260
attctgcggc ggcaggaaga tttttaccca ttcctgaagg acaaccggga aaagatcgag 1320
aagatcctga ccttccgcat cccctactac gtgggccctc tggccagggg aaacagcaga 1380
ttcgcctgga tgaccagaaa gagcgaggaa accatcaccc cctggaactt cgaggaagtg 1440
gtggacaagg gcgcttccgc ccagagcttc atcgagcgga tgaccaactt cgataagaac 1500
ctgcccaacg agaaggtgct gcccaagcac agcctgctgt acgagtactt caccgtgtat 1560
aacgagctga ccaaagtgaa atacgtgacc gagggaatga gaaagcccgc cttcctgagc 1620
ggcgagcaga aaaaggccat cgtggacctg ctgttcaaga ccaaccggaa agtgaccgtg 1680
aagcagctga aagaggacta cttcaagaaa atcgagtgct tcgactccgt ggaaatctcc 1740
ggcgtggaag atcggttcaa cgcctccctg ggcacatacc acgatctgct gaaaattatc 1800
aaggacaagg acttcctgga caatgaggaa aacgaggaca ttctggaaga tatcgtgctg 1860
accctgacac tgtttgagga cagagagatg atcgaggaac ggctgaaaac ctatgcccac 1920
ctgttcgacg acaaagtgat gaagcagctg aagcggcgga gatacaccgg ctggggcagg 1980
ctgagccgga agctgatcaa cggcatccgg gacaagcagt ccggcaagac aatcctggat 2040
ttcctgaagt ccgacggctt cgccaacaga aacttcatgc agctgatcca cgacgacagc 2100
ctgaccttta aagaggacat ccagaaagcc caggtgtccg gccagggcga tagcctgcac 2160
gagcacattg ccaatctggc cggcagcccc gccattaaga agggcatcct gcagacagtg 2220
aaggtggtgg acgagctcgt gaaagtgatg ggccggcaca agcccgagaa catcgtgatc 2280
gaaatggcca gagagaacca gaccacccag aagggacaga agaacagccg cgagagaatg 2340
aagcggatcg aagagggcat caaagagctg ggcagccaga tcctgaaaga acaccccgtg 2400
gaaaacaccc agctgcagaa cgagaagctg tacctgtact acctgcagaa tgggcgggat 2460
atgtacgtgg accaggaact ggacatcaac cggctgtccg actacgatgt ggacgccatc 2520
gtgcctcaga gctttctgaa ggacgactcc atcgacaaca aggtgctgac cagaagcgac 2580
aagaaccggg gcaagagcga caacgtgccc tccgaagagg tcgtgaagaa gatgaagaac 2640
tactggcggc agctgctgaa cgccaagctg attacccaga gaaagttcga caatctgacc 2700
aaggccgaga gaggcggcct gagcgaactg gataaggccg gcttcatcaa gagacagctg 2760
gtggaaaccc ggcagatcac aaagcacgtg gcacagatcc tggactcccg gatgaacact 2820
aagtacgacg agaatgacaa gctgatccgg gaagtgaaag tgatcaccct gaagtccaag 2880
ctggtgtccg atttccggaa ggatttccag ttttacaaag tgcgcgagat caacaactac 2940
caccacgccc acgacgccta cctgaacgcc gtcgtgggaa ccgccctgat caaaaagtac 3000
cctaagctgg aaagcgagtt cgtgtacggc gactacaagg tgtacgacgt gcggaagatg 3060
atcgccaaga gcgagcagga aatcggcaag gctaccgcca agtacttctt ctacagcaac 3120
atcatgaact ttttcaagac cgagattacc ctggccaacg gcgagatccg gaagcggcct 3180
ctgatcgaga caaacggcga aaccggggag atcgtgtggg ataagggccg ggattttgcc 3240
accgtgcgga aagtgctgag catgccccaa gtgaatatcg tgaaaaagac cgaggtgcag 3300
acaggcggct tcagcaaaga gtctatcctg cccaagagga acagcgataa gctgatcgcc 3360
agaaagaagg actgggaccc taagaagtac ggcggcttcg acagccccac cgtggcctat 3420
tctgtgctgg tggtggccaa agtggaaaag ggcaagtcca agaaactgaa gagtgtgaaa 3480
gagctgctgg ggatcaccat catggaaaga agcagcttcg agaagaatcc catcgacttt 3540
ctggaagcca agggctacaa agaagtgaaa aaggacctga tcatcaagct gcctaagtac 3600
tccctgttcg agctggaaaa cggccggaag agaatgctgg cctctgccgg cgaactgcag 3660
aagggaaacg aactggccct gccctccaaa tatgtgaact tcctgtacct ggccagccac 3720
tatgagaagc tgaagggctc ccccgaggat aatgagcaga aacagctgtt tgtggaacag 3780
cacaagcact acctggacga gatcatcgag cagatcagcg agttctccaa gagagtgatc 3840
ctggccgacg ctaatctgga caaagtgctg tccgcctaca acaagcaccg ggataagccc 3900
atcagagagc aggccgagaa tatcatccac ctgtttaccc tgaccaatct gggagcccct 3960
gccgccttca agtactttga caccaccatc gaccggaaga ggtacaccag caccaaagag 4020
gtgctggacg ccaccctgat ccaccagagc atcaccggcc tgtacgagac acggatcgac 4080
ctgtctcagc tgggaggtga c 4101
<210> 123
<211> 339
<212> PRT
<213> 智人
<400> 123
Met Ala Met Gln Met Gln Leu Glu Ala Asn Ala Asp Thr Ser Val Glu
1 5 10 15
Glu Glu Ser Phe Gly Pro Gln Pro Ile Ser Arg Leu Glu Gln Cys Gly
20 25 30
Ile Asn Ala Asn Asp Val Lys Lys Leu Glu Glu Ala Gly Phe His Thr
35 40 45
Val Glu Ala Val Ala Tyr Ala Pro Lys Lys Glu Leu Ile Asn Ile Lys
50 55 60
Gly Ile Ser Glu Ala Lys Ala Asp Lys Ile Leu Ala Glu Ala Ala Lys
65 70 75 80
Leu Val Pro Met Gly Phe Thr Thr Ala Thr Glu Phe His Gln Arg Arg
85 90 95
Ser Glu Ile Ile Gln Ile Thr Thr Gly Ser Lys Glu Leu Asp Lys Leu
100 105 110
Leu Gln Gly Gly Ile Glu Thr Gly Ser Ile Thr Glu Met Phe Gly Glu
115 120 125
Phe Arg Thr Gly Lys Thr Gln Ile Cys His Thr Leu Ala Val Thr Cys
130 135 140
Gln Leu Pro Ile Asp Arg Gly Gly Gly Glu Gly Lys Ala Met Tyr Ile
145 150 155 160
Asp Thr Glu Gly Thr Phe Arg Pro Glu Arg Leu Leu Ala Val Ala Glu
165 170 175
Arg Tyr Gly Leu Ser Gly Ser Asp Val Leu Asp Asn Val Ala Tyr Ala
180 185 190
Arg Ala Phe Asn Thr Asp His Gln Thr Gln Leu Leu Tyr Gln Ala Ser
195 200 205
Ala Met Met Val Glu Ser Arg Tyr Ala Leu Leu Ile Val Asp Ser Ala
210 215 220
Thr Ala Leu Tyr Arg Thr Asp Tyr Ser Gly Arg Gly Glu Leu Ser Ala
225 230 235 240
Arg Gln Met His Leu Ala Arg Phe Leu Arg Met Leu Leu Arg Leu Ala
245 250 255
Asp Glu Phe Gly Val Ala Val Val Ile Thr Asn Gln Val Val Ala Gln
260 265 270
Val Asp Gly Ala Ala Met Phe Ala Ala Asp Pro Lys Lys Pro Ile Gly
275 280 285
Gly Asn Ile Ile Ala His Ala Ser Thr Thr Arg Leu Tyr Leu Arg Lys
290 295 300
Gly Arg Gly Glu Thr Arg Ile Cys Lys Ile Tyr Asp Ser Pro Cys Leu
305 310 315 320
Pro Glu Ala Glu Ala Met Phe Ala Ile Asn Ala Asp Gly Val Gly Asp
325 330 335
Ala Lys Asp
<210> 124
<211> 418
<212> PRT
<213> 智人
<400> 124
Met Ser Gly Thr Glu Glu Ala Ile Leu Gly Gly Arg Asp Ser His Pro
1 5 10 15
Ala Ala Gly Gly Gly Ser Val Leu Cys Phe Gly Gln Cys Gln Tyr Thr
20 25 30
Ala Glu Glu Tyr Gln Ala Ile Gln Lys Ala Leu Arg Gln Arg Leu Gly
35 40 45
Pro Glu Tyr Ile Ser Ser Arg Met Ala Gly Gly Gly Gln Lys Val Cys
50 55 60
Tyr Ile Glu Gly His Arg Val Ile Asn Leu Ala Asn Glu Met Phe Gly
65 70 75 80
Tyr Asn Gly Trp Ala His Ser Ile Thr Gln Gln Asn Val Asp Phe Val
85 90 95
Asp Leu Asn Asn Gly Lys Phe Tyr Val Gly Val Cys Ala Phe Val Arg
100 105 110
Val Gln Leu Lys Asp Gly Ser Tyr His Glu Asp Val Gly Tyr Gly Val
115 120 125
Ser Glu Gly Leu Lys Ser Lys Ala Leu Ser Leu Glu Lys Ala Arg Lys
130 135 140
Glu Ala Val Thr Asp Gly Leu Lys Arg Ala Leu Arg Ser Phe Gly Asn
145 150 155 160
Ala Leu Gly Asn Cys Ile Leu Asp Lys Asp Tyr Leu Arg Ser Leu Asn
165 170 175
Lys Leu Pro Arg Gln Leu Pro Leu Glu Val Asp Leu Thr Lys Ala Lys
180 185 190
Arg Gln Asp Leu Glu Pro Ser Val Glu Glu Ala Arg Tyr Asn Ser Cys
195 200 205
Arg Pro Asn Met Ala Leu Gly His Pro Gln Leu Gln Gln Val Thr Ser
210 215 220
Pro Ser Arg Pro Ser His Ala Val Ile Pro Ala Asp Gln Asp Cys Ser
225 230 235 240
Ser Arg Ser Leu Ser Ser Ser Ala Val Glu Ser Glu Ala Thr His Gln
245 250 255
Arg Lys Leu Arg Gln Lys Gln Leu Gln Gln Gln Phe Arg Glu Arg Met
260 265 270
Glu Lys Gln Gln Val Arg Val Ser Thr Pro Ser Ala Glu Lys Ser Glu
275 280 285
Ala Ala Pro Pro Ala Pro Pro Val Thr His Ser Thr Pro Val Thr Val
290 295 300
Ser Glu Pro Leu Leu Glu Lys Asp Phe Leu Ala Gly Val Thr Gln Glu
305 310 315 320
Leu Ile Lys Thr Leu Glu Asp Asn Ser Glu Lys Trp Ala Val Thr Pro
325 330 335
Asp Ala Gly Asp Gly Val Val Lys Pro Ser Ser Arg Ala Asp Pro Ala
340 345 350
Gln Thr Ser Asp Thr Leu Ala Leu Asn Asn Gln Met Val Thr Gln Asn
355 360 365
Arg Thr Pro His Ser Val Cys His Gln Lys Pro Gln Ala Lys Ser Gly
370 375 380
Ser Trp Asp Leu Gln Thr Tyr Ser Ala Asp Gln Arg Thr Thr Gly Asn
385 390 395 400
Trp Glu Ser His Arg Lys Ser Gln Asp Met Lys Lys Arg Lys Tyr Asp
405 410 415
Pro Ser
<210> 125
<211> 348
<212> PRT
<213> 枯草芽孢杆菌（Bacillus subtilis）
<400> 125
Met Ser Asp Arg Gln Ala Ala Leu Asp Met Ala Leu Lys Gln Ile Glu
1 5 10 15
Lys Gln Phe Gly Lys Gly Ser Ile Met Lys Leu Gly Glu Lys Thr Asp
20 25 30
Thr Arg Ile Ser Thr Val Pro Ser Gly Ser Leu Ala Leu Asp Thr Ala
35 40 45
Leu Gly Ile Gly Gly Tyr Pro Arg Gly Arg Ile Ile Glu Val Tyr Gly
50 55 60
Pro Glu Ser Ser Gly Lys Thr Thr Val Ala Leu His Ala Ile Ala Glu
65 70 75 80
Val Gln Gln Gln Gly Gly Gln Ala Ala Phe Ile Asp Ala Glu His Ala
85 90 95
Leu Asp Pro Val Tyr Ala Gln Lys Leu Gly Val Asn Ile Glu Glu Leu
100 105 110
Leu Leu Ser Gln Pro Asp Thr Gly Glu Gln Ala Leu Glu Ile Ala Glu
115 120 125
Ala Leu Val Arg Ser Gly Ala Val Asp Ile Val Val Val Asp Ser Val
130 135 140
Ala Ala Leu Val Pro Lys Ala Glu Ile Glu Gly Asp Met Gly Asp Ser
145 150 155 160
His Val Gly Leu Gln Ala Arg Leu Met Ser Gln Ala Leu Arg Lys Leu
165 170 175
Ser Gly Ala Ile Asn Lys Ser Lys Thr Ile Ala Ile Phe Ile Asn Gln
180 185 190
Ile Arg Glu Lys Val Gly Val Met Phe Gly Asn Pro Glu Thr Thr Pro
195 200 205
Gly Gly Arg Ala Leu Lys Phe Tyr Ser Ser Val Arg Leu Glu Val Arg
210 215 220
Arg Ala Glu Gln Leu Lys Gln Gly Asn Asp Val Met Gly Asn Lys Thr
225 230 235 240
Lys Ile Lys Val Val Lys Asn Lys Val Ala Pro Pro Phe Arg Thr Ala
245 250 255
Glu Val Asp Ile Met Tyr Gly Glu Gly Ile Ser Lys Glu Gly Glu Ile
260 265 270
Ile Asp Leu Gly Thr Glu Leu Asp Ile Val Gln Lys Ser Gly Ser Trp
275 280 285
Tyr Ser Tyr Glu Glu Glu Arg Leu Gly Gln Gly Arg Glu Asn Ala Lys
290 295 300
Gln Phe Leu Lys Glu Asn Lys Asp Ile Met Leu Met Ile Gln Glu Gln
305 310 315 320
Ile Arg Glu His Tyr Gly Leu Asp Asn Asn Gly Val Val Gln Gln Gln
325 330 335
Ala Glu Glu Thr Gln Glu Glu Leu Glu Phe Glu Glu
340 345
<210> 126
<211> 353
<212> PRT
<213> 大肠杆菌（Escherichia coli）
<400> 126
Met Ala Ile Asp Glu Asn Lys Gln Lys Ala Leu Ala Ala Ala Leu Gly
1 5 10 15
Gln Ile Glu Lys Gln Phe Gly Lys Gly Ser Ile Met Arg Leu Gly Glu
20 25 30
Asp Arg Ser Met Asp Val Glu Thr Ile Ser Thr Gly Ser Leu Ser Leu
35 40 45
Asp Ile Ala Leu Gly Ala Gly Gly Leu Pro Met Gly Arg Ile Val Glu
50 55 60
Ile Tyr Gly Pro Glu Ser Ser Gly Lys Thr Thr Leu Thr Leu Gln Val
65 70 75 80
Ile Ala Ala Ala Gln Arg Glu Gly Lys Thr Cys Ala Phe Ile Asp Ala
85 90 95
Glu His Ala Leu Asp Pro Ile Tyr Ala Arg Lys Leu Gly Val Asp Ile
100 105 110
Asp Asn Leu Leu Cys Ser Gln Pro Asp Thr Gly Glu Gln Ala Leu Glu
115 120 125
Ile Cys Asp Ala Leu Ala Arg Ser Gly Ala Val Asp Val Ile Val Val
130 135 140
Asp Ser Val Ala Ala Leu Thr Pro Lys Ala Glu Ile Glu Gly Glu Ile
145 150 155 160
Gly Asp Ser His Met Gly Leu Ala Ala Arg Met Met Ser Gln Ala Met
165 170 175
Arg Lys Leu Ala Gly Asn Leu Lys Gln Ser Asn Thr Leu Leu Ile Phe
180 185 190
Ile Asn Gln Ile Arg Met Lys Ile Gly Val Met Phe Gly Asn Pro Glu
195 200 205
Thr Thr Thr Gly Gly Asn Ala Leu Lys Phe Tyr Ala Ser Val Arg Leu
210 215 220
Asp Ile Arg Arg Ile Gly Ala Val Lys Glu Gly Glu Asn Val Val Gly
225 230 235 240
Ser Glu Thr Arg Val Lys Val Val Lys Asn Lys Ile Ala Ala Pro Phe
245 250 255
Lys Gln Ala Glu Phe Gln Ile Leu Tyr Gly Glu Gly Ile Asn Phe Tyr
260 265 270
Gly Glu Leu Val Asp Leu Gly Val Lys Glu Lys Leu Ile Glu Lys Ala
275 280 285
Gly Ala Trp Tyr Ser Tyr Lys Gly Glu Lys Ile Gly Gln Gly Lys Ala
290 295 300
Asn Ala Thr Ala Trp Leu Lys Asp Asn Pro Glu Thr Ala Lys Glu Ile
305 310 315 320
Glu Lys Lys Val Arg Glu Leu Leu Leu Ser Asn Pro Asn Ser Thr Pro
325 330 335
Asp Phe Ser Val Asp Asp Ser Glu Gly Val Ala Glu Thr Asn Glu Asp
340 345 350
Phe
<210> 127
<211> 162
<212> PRT
<213> 埃希氏菌噬菌体P1
<400> 127
Met Ala Gln Arg Gly Val Asn Lys Val Ile Leu Ile Gly Thr Leu Gly
1 5 10 15
Gln Asp Pro Glu Ile Arg Tyr Ile Pro Asn Gly Gly Ala Val Gly Arg
20 25 30
Leu Ser Ile Ala Thr Asn Glu Ser Trp Arg Asp Lys Gln Thr Gly Gln
35 40 45
Gln Lys Glu Gln Thr Glu Trp His Lys Val Val Leu Phe Gly Lys Leu
50 55 60
Ala Glu Ile Ala Ser Glu Tyr Leu Arg Lys Gly Ser Gln Val Tyr Ile
65 70 75 80
Glu Gly Lys Leu Lys Thr Arg Lys Trp Thr Asp Asp Ala Gly Val Glu
85 90 95
Arg Tyr Thr Thr Glu Ile Ile Val Ser Gln Gly Gly Thr Met Gln Met
100 105 110
Ile Gly Ala Arg Arg Asp Asp Ser Gln Ser Ser Asn Gly Trp Gly Gln
115 120 125
Ser Asn Gln Pro Gln Asn His Gln Gln Tyr Ser Gly Gly Gly Lys Pro
130 135 140
Gln Ser Asn Ala Asn Asn Glu Pro Pro Met Asp Phe Asp Asp Asp Ile
145 150 155 160
Pro Phe
<210> 128
<211> 36
<212> PRT
<213> 智人
<400> 128
Ala Leu Asp Phe Leu Ser Arg Leu Pro Leu Pro Pro Pro Val Ser Pro
1 5 10 15
Ile Cys Thr Phe Val Ser Pro Ala Ala Gln Lys Ala Phe Gln Pro Pro
20 25 30
Arg Ser Cys Gly
35
<210> 129
<211> 866
<212> PRT
<213> 大肠杆菌
<400> 129
Met Ser Thr Lys Pro Leu Phe Leu Leu Arg Lys Ala Lys Lys Ser Ser
1 5 10 15
Gly Glu Pro Asp Val Val Leu Trp Ala Ser Asn Asp Phe Glu Ser Thr
20 25 30
Cys Ala Thr Leu Asp Tyr Leu Ile Val Lys Ser Gly Lys Lys Leu Ser
35 40 45
Ser Tyr Phe Lys Ala Val Ala Thr Asn Phe Pro Val Val Asn Asp Leu
50 55 60
Pro Ala Glu Gly Glu Ile Asp Phe Thr Trp Ser Glu Arg Tyr Gln Leu
65 70 75 80
Ser Lys Asp Ser Met Thr Trp Glu Leu Lys Pro Gly Ala Ala Pro Asp
85 90 95
Asn Ala His Tyr Gln Gly Asn Thr Asn Val Asn Gly Glu Asp Met Thr
100 105 110
Glu Ile Glu Glu Asn Met Leu Leu Pro Ile Ser Gly Gln Glu Leu Pro
115 120 125
Ile Arg Trp Leu Ala Gln His Gly Ser Glu Lys Pro Val Thr His Val
130 135 140
Ser Arg Asp Gly Leu Gln Ala Leu His Ile Ala Arg Ala Glu Glu Leu
145 150 155 160
Pro Ala Val Thr Ala Leu Ala Val Ser His Lys Thr Ser Leu Leu Asp
165 170 175
Pro Leu Glu Ile Arg Glu Leu His Lys Leu Val Arg Asp Thr Asp Lys
180 185 190
Val Phe Pro Asn Pro Gly Asn Ser Asn Leu Gly Leu Ile Thr Ala Phe
195 200 205
Phe Glu Ala Tyr Leu Asn Ala Asp Tyr Thr Asp Arg Gly Leu Leu Thr
210 215 220
Lys Glu Trp Met Lys Gly Asn Arg Val Ser His Ile Thr Arg Thr Ala
225 230 235 240
Ser Gly Ala Asn Ala Gly Gly Gly Asn Leu Thr Asp Arg Gly Glu Gly
245 250 255
Phe Val His Asp Leu Thr Ser Leu Ala Arg Asp Val Ala Thr Gly Val
260 265 270
Leu Ala Arg Ser Met Asp Leu Asp Ile Tyr Asn Leu His Pro Ala His
275 280 285
Ala Lys Arg Ile Glu Glu Ile Ile Ala Glu Asn Lys Pro Pro Phe Ser
290 295 300
Val Phe Arg Asp Lys Phe Ile Thr Met Pro Gly Gly Leu Asp Tyr Ser
305 310 315 320
Arg Ala Ile Val Val Ala Ser Val Lys Glu Ala Pro Ile Gly Ile Glu
325 330 335
Val Ile Pro Ala His Val Thr Glu Tyr Leu Asn Lys Val Leu Thr Glu
340 345 350
Thr Asp His Ala Asn Pro Asp Pro Glu Ile Val Asp Ile Ala Cys Gly
355 360 365
Arg Ser Ser Ala Pro Met Pro Gln Arg Val Thr Glu Glu Gly Lys Gln
370 375 380
Asp Asp Glu Glu Lys Pro Gln Pro Ser Gly Thr Thr Ala Val Glu Gln
385 390 395 400
Gly Glu Ala Glu Thr Met Glu Pro Asp Ala Thr Glu His His Gln Asp
405 410 415
Thr Gln Pro Leu Asp Ala Gln Ser Gln Val Asn Ser Val Asp Ala Lys
420 425 430
Tyr Gln Glu Leu Arg Ala Glu Leu His Glu Ala Arg Lys Asn Ile Pro
435 440 445
Ser Lys Asn Pro Val Asp Asp Asp Lys Leu Leu Ala Ala Ser Arg Gly
450 455 460
Glu Phe Val Asp Gly Ile Ser Asp Pro Asn Asp Pro Lys Trp Val Lys
465 470 475 480
Gly Ile Gln Thr Arg Asp Cys Val Tyr Gln Asn Gln Pro Glu Thr Glu
485 490 495
Lys Thr Ser Pro Asp Met Asn Gln Pro Glu Pro Val Val Gln Gln Glu
500 505 510
Pro Glu Ile Ala Cys Asn Ala Cys Gly Gln Thr Gly Gly Asp Asn Cys
515 520 525
Pro Asp Cys Gly Ala Val Met Gly Asp Ala Thr Tyr Gln Glu Thr Phe
530 535 540
Asp Glu Glu Ser Gln Val Glu Ala Lys Glu Asn Asp Pro Glu Glu Met
545 550 555 560
Glu Gly Ala Glu His Pro His Asn Glu Asn Ala Gly Ser Asp Pro His
565 570 575
Arg Asp Cys Ser Asp Glu Thr Gly Glu Val Ala Asp Pro Val Ile Val
580 585 590
Glu Asp Ile Glu Pro Gly Ile Tyr Tyr Gly Ile Ser Asn Glu Asn Tyr
595 600 605
His Ala Gly Pro Gly Ile Ser Lys Ser Gln Leu Asp Asp Ile Ala Asp
610 615 620
Thr Pro Ala Leu Tyr Leu Trp Arg Lys Asn Ala Pro Val Asp Thr Thr
625 630 635 640
Lys Thr Lys Thr Leu Asp Leu Gly Thr Ala Phe His Cys Arg Val Leu
645 650 655
Glu Pro Glu Glu Phe Ser Asn Arg Phe Ile Val Ala Pro Glu Phe Asn
660 665 670
Arg Arg Thr Asn Ala Gly Lys Glu Glu Glu Lys Ala Phe Leu Met Glu
675 680 685
Cys Ala Ser Thr Gly Lys Thr Val Ile Thr Ala Glu Glu Gly Arg Lys
690 695 700
Ile Glu Leu Met Tyr Gln Ser Val Met Ala Leu Pro Leu Gly Gln Trp
705 710 715 720
Leu Val Glu Ser Ala Gly His Ala Glu Ser Ser Ile Tyr Trp Glu Asp
725 730 735
Pro Glu Thr Gly Ile Leu Cys Arg Cys Arg Pro Asp Lys Ile Ile Pro
740 745 750
Glu Phe His Trp Ile Met Asp Val Lys Thr Thr Ala Asp Ile Gln Arg
755 760 765
Phe Lys Thr Ala Tyr Tyr Asp Tyr Arg Tyr His Val Gln Asp Ala Phe
770 775 780
Tyr Ser Asp Gly Tyr Glu Ala Gln Phe Gly Val Gln Pro Thr Phe Val
785 790 795 800
Phe Leu Val Ala Ser Thr Thr Ile Glu Cys Gly Arg Tyr Pro Val Glu
805 810 815
Ile Phe Met Met Gly Glu Glu Ala Lys Leu Ala Gly Gln Gln Glu Tyr
820 825 830
His Arg Asn Leu Arg Thr Leu Ser Asp Cys Leu Asn Thr Asp Glu Trp
835 840 845
Pro Ala Ile Lys Thr Leu Ser Leu Pro Arg Trp Ala Lys Glu Tyr Ala
850 855 860
Asn Asp
865
<210> 130
<211> 577
<212> PRT
<213> 大肠杆菌
<400> 130
Met Lys Gln Gln Ile Gln Leu Arg Arg Arg Glu Val Asp Glu Thr Ala
1 5 10 15
Asp Leu Pro Ala Glu Leu Pro Pro Leu Leu Arg Arg Leu Tyr Ala Ser
20 25 30
Arg Gly Val Arg Ser Ala Gln Glu Leu Glu Arg Ser Val Lys Gly Met
35 40 45
Leu Pro Trp Gln Gln Leu Ser Gly Val Glu Lys Ala Val Glu Ile Leu
50 55 60
Tyr Asn Ala Phe Arg Glu Gly Thr Arg Ile Ile Val Val Gly Asp Phe
65 70 75 80
Asp Ala Asp Gly Ala Thr Ser Thr Ala Leu Ser Val Leu Ala Met Arg
85 90 95
Ser Leu Gly Cys Ser Asn Ile Asp Tyr Leu Val Pro Asn Arg Phe Glu
100 105 110
Asp Gly Tyr Gly Leu Ser Pro Glu Val Val Asp Gln Ala His Ala Arg
115 120 125
Gly Ala Gln Leu Ile Val Thr Val Asp Asn Gly Ile Ser Ser His Ala
130 135 140
Gly Val Glu His Ala Arg Ser Leu Gly Ile Pro Val Ile Val Thr Asp
145 150 155 160
His His Leu Pro Gly Asp Thr Leu Pro Ala Ala Glu Ala Ile Ile Asn
165 170 175
Pro Asn Leu Arg Asp Cys Asn Phe Pro Ser Lys Ser Leu Ala Gly Val
180 185 190
Gly Val Ala Phe Tyr Leu Met Leu Ala Leu Arg Thr Phe Leu Arg Asp
195 200 205
Gln Gly Trp Phe Asp Glu Arg Asn Ile Ala Ile Pro Asn Leu Ala Glu
210 215 220
Leu Leu Asp Leu Val Ala Leu Gly Thr Val Ala Asp Val Val Pro Leu
225 230 235 240
Asp Ala Asn Asn Arg Ile Leu Thr Trp Gln Gly Met Ser Arg Ile Arg
245 250 255
Ala Gly Lys Cys Arg Pro Gly Ile Lys Ala Leu Leu Glu Val Ala Asn
260 265 270
Arg Asp Ala Gln Lys Leu Ala Ala Ser Asp Leu Gly Phe Ala Leu Gly
275 280 285
Pro Arg Leu Asn Ala Ala Gly Arg Leu Asp Asp Met Ser Val Gly Val
290 295 300
Ala Leu Leu Leu Cys Asp Asn Ile Gly Glu Ala Arg Val Leu Ala Asn
305 310 315 320
Glu Leu Asp Ala Leu Asn Gln Thr Arg Lys Glu Ile Glu Gln Gly Met
325 330 335
Gln Ile Glu Ala Leu Thr Leu Cys Glu Lys Leu Glu Arg Ser Arg Asp
340 345 350
Thr Leu Pro Gly Gly Leu Ala Met Tyr His Pro Glu Trp His Gln Gly
355 360 365
Val Val Gly Ile Leu Ala Ser Arg Ile Lys Glu Arg Phe His Arg Pro
370 375 380
Val Ile Ala Phe Ala Pro Ala Gly Asp Gly Thr Leu Lys Gly Ser Gly
385 390 395 400
Arg Ser Ile Gln Gly Leu His Met Arg Asp Ala Leu Glu Arg Leu Asp
405 410 415
Thr Leu Tyr Pro Gly Met Met Leu Lys Phe Gly Gly His Ala Met Ala
420 425 430
Ala Gly Leu Ser Leu Glu Glu Asp Lys Phe Lys Leu Phe Gln Gln Arg
435 440 445
Phe Gly Glu Leu Val Thr Glu Trp Leu Asp Pro Ser Leu Leu Gln Gly
450 455 460
Glu Val Val Ser Asp Gly Pro Leu Ser Pro Ala Glu Met Thr Met Glu
465 470 475 480
Val Ala Gln Leu Leu Arg Asp Ala Gly Pro Trp Gly Gln Met Phe Pro
485 490 495
Glu Pro Leu Phe Asp Gly His Phe Arg Leu Leu Gln Gln Arg Leu Val
500 505 510
Gly Glu Arg His Leu Lys Val Met Val Glu Pro Val Gly Gly Gly Pro
515 520 525
Leu Leu Asp Gly Ile Ala Phe Asn Val Asp Thr Ala Leu Trp Pro Asp
530 535 540
Asn Gly Val Arg Glu Val Gln Leu Ala Tyr Lys Leu Asp Ile Asn Glu
545 550 555 560
Phe Arg Gly Asn Arg Ser Leu Gln Ile Ile Ile Asp Asn Ile Trp Pro
565 570 575
Ile
<210> 131
<211> 291
<212> PRT
<213> 埃希氏菌噬菌体T5
<400> 131
Met Ser Lys Ser Trp Gly Lys Phe Ile Glu Glu Glu Glu Ala Glu Met
1 5 10 15
Ala Ser Arg Arg Asn Leu Met Ile Val Asp Gly Thr Asn Leu Gly Phe
20 25 30
Arg Phe Lys His Asn Asn Ser Lys Lys Pro Phe Ala Ser Ser Tyr Val
35 40 45
Ser Thr Ile Gln Ser Leu Ala Lys Ser Tyr Ser Ala Arg Thr Thr Ile
50 55 60
Val Leu Gly Asp Lys Gly Lys Ser Val Phe Arg Leu Glu His Leu Pro
65 70 75 80
Glu Tyr Lys Gly Asn Arg Asp Glu Lys Tyr Ala Gln Arg Thr Glu Glu
85 90 95
Glu Lys Ala Leu Asp Glu Gln Phe Phe Glu Tyr Leu Lys Asp Ala Phe
100 105 110
Glu Leu Cys Lys Thr Thr Phe Pro Thr Phe Thr Ile Arg Gly Val Glu
115 120 125
Ala Asp Asp Met Ala Ala Tyr Ile Val Lys Leu Ile Gly His Leu Tyr
130 135 140
Asp His Val Trp Leu Ile Ser Thr Asp Gly Asp Trp Asp Thr Leu Leu
145 150 155 160
Thr Asp Lys Val Ser Arg Phe Ser Phe Thr Thr Arg Arg Glu Tyr His
165 170 175
Leu Arg Asp Met Tyr Glu His His Asn Val Asp Asp Val Glu Gln Phe
180 185 190
Ile Ser Leu Lys Ala Ile Met Gly Asp Leu Gly Asp Asn Ile Arg Gly
195 200 205
Val Glu Gly Ile Gly Ala Lys Arg Gly Tyr Asn Ile Ile Arg Glu Phe
210 215 220
Gly Asn Val Leu Asp Ile Ile Asp Gln Leu Pro Leu Pro Gly Lys Gln
225 230 235 240
Lys Tyr Ile Gln Asn Leu Asn Ala Ser Glu Glu Leu Leu Phe Arg Asn
245 250 255
Leu Ile Leu Val Asp Leu Pro Thr Tyr Cys Val Asp Ala Ile Ala Ala
260 265 270
Val Gly Gln Asp Val Leu Asp Lys Phe Thr Lys Asp Ile Leu Glu Ile
275 280 285
Ala Glu Gln
290
<210> 132
<211> 300
<212> PRT
<213> 埃希氏菌噬菌体T7
<400> 132
Met Ala Leu Leu Asp Leu Lys Gln Phe Tyr Glu Leu Arg Glu Gly Cys
1 5 10 15
Asp Asp Lys Gly Ile Leu Val Met Asp Gly Asp Trp Leu Val Phe Gln
20 25 30
Ala Met Ser Ala Ala Glu Phe Asp Ala Ser Trp Glu Glu Glu Ile Trp
35 40 45
His Arg Cys Cys Asp His Ala Lys Ala Arg Gln Ile Leu Glu Asp Ser
50 55 60
Ile Lys Ser Tyr Glu Thr Arg Lys Lys Ala Trp Ala Gly Ala Pro Ile
65 70 75 80
Val Leu Ala Phe Thr Asp Ser Val Asn Trp Arg Lys Glu Leu Val Asp
85 90 95
Pro Asn Tyr Lys Ala Asn Arg Lys Ala Val Lys Lys Pro Val Gly Tyr
100 105 110
Phe Glu Phe Leu Asp Ala Leu Phe Glu Arg Glu Glu Phe Tyr Cys Ile
115 120 125
Arg Glu Pro Met Leu Glu Gly Asp Asp Val Met Gly Val Ile Ala Ser
130 135 140
Asn Pro Ser Ala Phe Gly Ala Arg Lys Ala Val Ile Ile Ser Cys Asp
145 150 155 160
Lys Asp Phe Lys Thr Ile Pro Asn Cys Asp Phe Leu Trp Cys Thr Thr
165 170 175
Gly Asn Ile Leu Thr Gln Thr Glu Glu Ser Ala Asp Trp Trp His Leu
180 185 190
Phe Gln Thr Ile Lys Gly Asp Ile Thr Asp Gly Tyr Ser Gly Ile Ala
195 200 205
Gly Trp Gly Asp Thr Ala Glu Asp Phe Leu Asn Asn Pro Phe Ile Thr
210 215 220
Glu Pro Lys Thr Ser Val Leu Lys Ser Gly Lys Asn Lys Gly Gln Glu
225 230 235 240
Val Thr Lys Trp Val Lys Arg Asp Pro Glu Pro His Glu Thr Leu Trp
245 250 255
Asp Cys Ile Lys Ser Ile Gly Ala Lys Ala Gly Met Thr Glu Glu Asp
260 265 270
Ile Ile Lys Gln Gly Gln Met Ala Arg Ile Leu Arg Phe Asn Glu Tyr
275 280 285
Asn Phe Ile Asp Lys Glu Ile Tyr Leu Trp Arg Pro
290 295 300
<210> 133
<211> 226
<212> PRT
<213> 埃希氏菌噬菌体lambda
<400> 133
Met Thr Pro Asp Ile Ile Leu Gln Arg Thr Gly Ile Asp Val Arg Ala
1 5 10 15
Val Glu Gln Gly Asp Asp Ala Trp His Lys Leu Arg Leu Gly Val Ile
20 25 30
Thr Ala Ser Glu Val His Asn Val Ile Ala Lys Pro Arg Ser Gly Lys
35 40 45
Lys Trp Pro Asp Met Lys Met Ser Tyr Phe His Thr Leu Leu Ala Glu
50 55 60
Val Cys Thr Gly Val Ala Pro Glu Val Asn Ala Lys Ala Leu Ala Trp
65 70 75 80
Gly Lys Gln Tyr Glu Asn Asp Ala Arg Thr Leu Phe Glu Phe Thr Ser
85 90 95
Gly Val Asn Val Thr Glu Ser Pro Ile Ile Tyr Arg Asp Glu Ser Met
100 105 110
Arg Thr Ala Cys Ser Pro Asp Gly Leu Cys Ser Asp Gly Asn Gly Leu
115 120 125
Glu Leu Lys Cys Pro Phe Thr Ser Arg Asp Phe Met Lys Phe Arg Leu
130 135 140
Gly Gly Phe Glu Ala Ile Lys Ser Ala Tyr Met Ala Gln Val Gln Tyr
145 150 155 160
Ser Met Trp Val Thr Arg Lys Asn Ala Trp Tyr Phe Ala Asn Tyr Asp
165 170 175
Pro Arg Met Lys Arg Glu Gly Leu His Tyr Val Val Ile Glu Arg Asp
180 185 190
Glu Lys Tyr Met Ala Ser Phe Asp Glu Ile Val Pro Glu Phe Ile Glu
195 200 205
Lys Met Asp Glu Ala Leu Ala Glu Ile Gly Phe Val Phe Gly Glu Gln
210 215 220
Trp Arg
225
<210> 134
<211> 475
<212> PRT
<213> 大肠杆菌
<400> 134
Met Met Asn Asp Gly Lys Gln Gln Ser Thr Phe Leu Phe His Asp Tyr
1 5 10 15
Glu Thr Phe Gly Thr His Pro Ala Leu Asp Arg Pro Ala Gln Phe Ala
20 25 30
Ala Ile Arg Thr Asp Ser Glu Phe Asn Val Ile Gly Glu Pro Glu Val
35 40 45
Phe Tyr Cys Lys Pro Ala Asp Asp Tyr Leu Pro Gln Pro Gly Ala Val
50 55 60
Leu Ile Thr Gly Ile Thr Pro Gln Glu Ala Arg Ala Lys Gly Glu Asn
65 70 75 80
Glu Ala Ala Phe Ala Ala Arg Ile His Ser Leu Phe Thr Val Pro Lys
85 90 95
Thr Cys Ile Leu Gly Tyr Asn Asn Val Arg Phe Asp Asp Glu Val Thr
100 105 110
Arg Asn Ile Phe Tyr Arg Asn Phe Tyr Asp Pro Tyr Ala Trp Ser Trp
115 120 125
Gln His Asp Asn Ser Arg Trp Asp Leu Leu Asp Val Met Arg Ala Cys
130 135 140
Tyr Ala Leu Arg Pro Glu Gly Ile Asn Trp Pro Glu Asn Asp Asp Gly
145 150 155 160
Leu Pro Ser Phe Arg Leu Glu His Leu Thr Lys Ala Asn Gly Ile Glu
165 170 175
His Ser Asn Ala His Asp Ala Met Ala Asp Val Tyr Ala Thr Ile Ala
180 185 190
Met Ala Lys Leu Val Lys Thr Arg Gln Pro Arg Leu Phe Asp Tyr Leu
195 200 205
Phe Thr His Arg Asn Lys His Lys Leu Met Ala Leu Ile Asp Val Pro
210 215 220
Gln Met Lys Pro Leu Val His Val Ser Gly Met Phe Gly Ala Trp Arg
225 230 235 240
Gly Asn Thr Ser Trp Val Ala Pro Leu Ala Trp His Pro Glu Asn Arg
245 250 255
Asn Ala Val Ile Met Val Asp Leu Ala Gly Asp Ile Ser Pro Leu Leu
260 265 270
Glu Leu Asp Ser Asp Thr Leu Arg Glu Arg Leu Tyr Thr Ala Lys Thr
275 280 285
Asp Leu Gly Asp Asn Ala Ala Val Pro Val Lys Leu Val His Ile Asn
290 295 300
Lys Cys Pro Val Leu Ala Gln Ala Asn Thr Leu Arg Pro Glu Asp Ala
305 310 315 320
Asp Arg Leu Gly Ile Asn Arg Gln His Cys Leu Asp Asn Leu Lys Ile
325 330 335
Leu Arg Glu Asn Pro Gln Val Arg Glu Lys Val Val Ala Ile Phe Ala
340 345 350
Glu Ala Glu Pro Phe Thr Pro Ser Asp Asn Val Asp Ala Gln Leu Tyr
355 360 365
Asn Gly Phe Phe Ser Asp Ala Asp Arg Ala Ala Met Lys Ile Val Leu
370 375 380
Glu Thr Glu Pro Arg Asn Leu Pro Ala Leu Asp Ile Thr Phe Val Asp
385 390 395 400
Lys Arg Ile Glu Lys Leu Leu Phe Asn Tyr Arg Ala Arg Asn Phe Pro
405 410 415
Gly Thr Leu Asp Tyr Ala Glu Gln Gln Arg Trp Leu Glu His Arg Arg
420 425 430
Gln Val Phe Thr Pro Glu Phe Leu Gln Gly Tyr Ala Asp Glu Leu Gln
435 440 445
Met Leu Val Gln Gln Tyr Ala Asp Asp Lys Glu Lys Val Ala Leu Leu
450 455 460
Lys Ala Leu Trp Gln Tyr Ala Glu Glu Ile Val
465 470 475
<210> 135
<211> 846
<212> PRT
<213> 智人
<400> 135
Met Gly Ile Gln Gly Leu Leu Gln Phe Ile Lys Glu Ala Ser Glu Pro
1 5 10 15
Ile His Val Arg Lys Tyr Lys Gly Gln Val Val Ala Val Asp Thr Tyr
20 25 30
Cys Trp Leu His Lys Gly Ala Ile Ala Cys Ala Glu Lys Leu Ala Lys
35 40 45
Gly Glu Pro Thr Asp Arg Tyr Val Gly Phe Cys Met Lys Phe Val Asn
50 55 60
Met Leu Leu Ser His Gly Ile Lys Pro Ile Leu Val Phe Asp Gly Cys
65 70 75 80
Thr Leu Pro Ser Lys Lys Glu Val Glu Arg Ser Arg Arg Glu Arg Arg
85 90 95
Gln Ala Asn Leu Leu Lys Gly Lys Gln Leu Leu Arg Glu Gly Lys Val
100 105 110
Ser Glu Ala Arg Glu Cys Phe Thr Arg Ser Ile Asn Ile Thr His Ala
115 120 125
Met Ala His Lys Val Ile Lys Ala Ala Arg Ser Gln Gly Val Asp Cys
130 135 140
Leu Val Ala Pro Tyr Glu Ala Asp Ala Gln Leu Ala Tyr Leu Asn Lys
145 150 155 160
Ala Gly Ile Val Gln Ala Ile Ile Thr Glu Asp Ser Asp Leu Leu Ala
165 170 175
Phe Gly Cys Lys Lys Val Ile Leu Lys Met Asp Gln Phe Gly Asn Gly
180 185 190
Leu Glu Ile Asp Gln Ala Arg Leu Gly Met Cys Arg Gln Leu Gly Asp
195 200 205
Val Phe Thr Glu Glu Lys Phe Arg Tyr Met Cys Ile Leu Ser Gly Cys
210 215 220
Asp Tyr Leu Ser Ser Leu Arg Gly Ile Gly Leu Ala Lys Ala Cys Lys
225 230 235 240
Val Leu Arg Leu Ala Asn Asn Pro Asp Ile Val Lys Val Ile Lys Lys
245 250 255
Ile Gly His Tyr Leu Lys Met Asn Ile Thr Val Pro Glu Asp Tyr Ile
260 265 270
Asn Gly Phe Ile Arg Ala Asn Asn Thr Phe Leu Tyr Gln Leu Val Phe
275 280 285
Asp Pro Ile Lys Arg Lys Leu Ile Pro Leu Asn Ala Tyr Glu Asp Asp
290 295 300
Val Asp Pro Glu Thr Leu Ser Tyr Ala Gly Gln Tyr Val Asp Asp Ser
305 310 315 320
Ile Ala Leu Gln Ile Ala Leu Gly Asn Lys Asp Ile Asn Thr Phe Glu
325 330 335
Gln Ile Asp Asp Tyr Asn Pro Asp Thr Ala Met Pro Ala His Ser Arg
340 345 350
Ser His Ser Trp Asp Asp Lys Thr Cys Gln Lys Ser Ala Asn Val Ser
355 360 365
Ser Ile Trp His Arg Asn Tyr Ser Pro Arg Pro Glu Ser Gly Thr Val
370 375 380
Ser Asp Ala Pro Gln Leu Lys Glu Asn Pro Ser Thr Val Gly Val Glu
385 390 395 400
Arg Val Ile Ser Thr Lys Gly Leu Asn Leu Pro Arg Lys Ser Ser Ile
405 410 415
Val Lys Arg Pro Arg Ser Ala Glu Leu Ser Glu Asp Asp Leu Leu Ser
420 425 430
Gln Tyr Ser Leu Ser Phe Thr Lys Lys Thr Lys Lys Asn Ser Ser Glu
435 440 445
Gly Asn Lys Ser Leu Ser Phe Ser Glu Val Phe Val Pro Asp Leu Val
450 455 460
Asn Gly Pro Thr Asn Lys Lys Ser Val Ser Thr Pro Pro Arg Thr Arg
465 470 475 480
Asn Lys Phe Ala Thr Phe Leu Gln Arg Lys Asn Glu Glu Ser Gly Ala
485 490 495
Val Val Val Pro Gly Thr Arg Ser Arg Phe Phe Cys Ser Ser Asp Ser
500 505 510
Thr Asp Cys Val Ser Asn Lys Val Ser Ile Gln Pro Leu Asp Glu Thr
515 520 525
Ala Val Thr Asp Lys Glu Asn Asn Leu His Glu Ser Glu Tyr Gly Asp
530 535 540
Gln Glu Gly Lys Arg Leu Val Asp Thr Asp Val Ala Arg Asn Ser Ser
545 550 555 560
Asp Asp Ile Pro Asn Asn His Ile Pro Gly Asp His Ile Pro Asp Lys
565 570 575
Ala Thr Val Phe Thr Asp Glu Glu Ser Tyr Ser Phe Glu Ser Ser Lys
580 585 590
Phe Thr Arg Thr Ile Ser Pro Pro Thr Leu Gly Thr Leu Arg Ser Cys
595 600 605
Phe Ser Trp Ser Gly Gly Leu Gly Asp Phe Ser Arg Thr Pro Ser Pro
610 615 620
Ser Pro Ser Thr Ala Leu Gln Gln Phe Arg Arg Lys Ser Asp Ser Pro
625 630 635 640
Thr Ser Leu Pro Glu Asn Asn Met Ser Asp Val Ser Gln Leu Lys Ser
645 650 655
Glu Glu Ser Ser Asp Asp Glu Ser His Pro Leu Arg Glu Glu Ala Cys
660 665 670
Ser Ser Gln Ser Gln Glu Ser Gly Glu Phe Ser Leu Gln Ser Ser Asn
675 680 685
Ala Ser Lys Leu Ser Gln Cys Ser Ser Lys Asp Ser Asp Ser Glu Glu
690 695 700
Ser Asp Cys Asn Ile Lys Leu Leu Asp Ser Gln Ser Asp Gln Thr Ser
705 710 715 720
Lys Leu Arg Leu Ser His Phe Ser Lys Lys Asp Thr Pro Leu Arg Asn
725 730 735
Lys Val Pro Gly Leu Tyr Lys Ser Ser Ser Ala Asp Ser Leu Ser Thr
740 745 750
Thr Lys Ile Lys Pro Leu Gly Pro Ala Arg Ala Ser Gly Leu Ser Lys
755 760 765
Lys Pro Ala Ser Ile Gln Lys Arg Lys His His Asn Ala Glu Asn Lys
770 775 780
Pro Gly Leu Gln Ile Lys Leu Asn Glu Leu Trp Lys Asn Phe Gly Phe
785 790 795 800
Lys Lys Asp Ser Glu Lys Leu Pro Pro Cys Lys Lys Pro Leu Ser Pro
805 810 815
Val Arg Asp Asn Ile Gln Leu Thr Pro Glu Ala Glu Glu Asp Ile Phe
820 825 830
Asn Lys Pro Glu Cys Gly Arg Val Gln Arg Ala Ile Phe Gln
835 840 845
<210> 136
<211> 23
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 136
gctcagcagg cacctgcctc agc 23
<210> 137
<211> 23
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 137
ctgatggtcc atgtctgtta ctc 23
<210> 138
<211> 27
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 138
tttggctcac tcctgctcgg tgaattt 27
<210> 139
<211> 27
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 139
tttcgcgctt gttccaatca gtacgca 27
<210> 140
<211> 20
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 140
gagtccgagc agaagaagaa 20
<210> 141
<211> 20
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 141
gcattttcag gaggaagcga 20
<210> 142
<211> 20
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 142
gtcatcttag tcattacctg 20
<210> 143
<211> 20
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 143
ggaatccctt ctgcagcacc 20
<210> 144
<211> 22
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 144
acgtactgat gttaacagct ga 22
<210> 145
<211> 22
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 145
ggtcagctgt taacatcagt ac 22
<210> 146
<211> 22
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 146
tccagcccgc tggccctgta aa 22
<210> 147
<211> 174
<212> PRT
<213> 埃希氏菌噬菌体Mu
<400> 147
Met Ala Lys Pro Ala Lys Arg Ile Lys Ser Ala Ala Ala Ala Tyr Val
1 5 10 15
Pro Gln Asn Arg Asp Ala Val Ile Thr Asp Ile Lys Arg Ile Gly Asp
20 25 30
Leu Gln Arg Glu Ala Ser Arg Leu Glu Thr Glu Met Asn Asp Ala Ile
35 40 45
Ala Glu Ile Thr Glu Lys Phe Ala Ala Arg Ile Ala Pro Ile Lys Thr
50 55 60
Asp Ile Glu Thr Leu Ser Lys Gly Val Gln Gly Trp Cys Glu Ala Asn
65 70 75 80
Arg Asp Glu Leu Thr Asn Gly Gly Lys Val Lys Thr Ala Asn Leu Val
85 90 95
Thr Gly Asp Val Ser Trp Arg Val Arg Pro Pro Ser Val Ser Ile Arg
100 105 110
Gly Met Asp Ala Val Met Glu Thr Leu Glu Arg Leu Gly Leu Gln Arg
115 120 125
Phe Ile Arg Thr Lys Gln Glu Ile Asn Lys Glu Ala Ile Leu Leu Glu
130 135 140
Pro Lys Ala Val Ala Gly Val Ala Gly Ile Thr Val Lys Ser Gly Ile
145 150 155 160
Glu Asp Phe Ser Ile Ile Pro Phe Glu Gln Glu Ala Gly Ile
165 170
<210> 148
<211> 1228
<212> PRT
<213> 人工序列
<220>
<223> 合成的LbCas12a H759A
<400> 148
Met Ser Lys Leu Glu Lys Phe Thr Asn Cys Tyr Ser Leu Ser Lys Thr
1 5 10 15
Leu Arg Phe Lys Ala Ile Pro Val Gly Lys Thr Gln Glu Asn Ile Asp
20 25 30
Asn Lys Arg Leu Leu Val Glu Asp Glu Lys Arg Ala Glu Asp Tyr Lys
35 40 45
Gly Val Lys Lys Leu Leu Asp Arg Tyr Tyr Leu Ser Phe Ile Asn Asp
50 55 60
Val Leu His Ser Ile Lys Leu Lys Asn Leu Asn Asn Tyr Ile Ser Leu
65 70 75 80
Phe Arg Lys Lys Thr Arg Thr Glu Lys Glu Asn Lys Glu Leu Glu Asn
85 90 95
Leu Glu Ile Asn Leu Arg Lys Glu Ile Ala Lys Ala Phe Lys Gly Asn
100 105 110
Glu Gly Tyr Lys Ser Leu Phe Lys Lys Asp Ile Ile Glu Thr Ile Leu
115 120 125
Pro Glu Phe Leu Asp Asp Lys Asp Glu Ile Ala Leu Val Asn Ser Phe
130 135 140
Asn Gly Phe Thr Thr Ala Phe Thr Gly Phe Phe Asp Asn Arg Glu Asn
145 150 155 160
Met Phe Ser Glu Glu Ala Lys Ser Thr Ser Ile Ala Phe Arg Cys Ile
165 170 175
Asn Glu Asn Leu Thr Arg Tyr Ile Ser Asn Met Asp Ile Phe Glu Lys
180 185 190
Val Asp Ala Ile Phe Asp Lys His Glu Val Gln Glu Ile Lys Glu Lys
195 200 205
Ile Leu Asn Ser Asp Tyr Asp Val Glu Asp Phe Phe Glu Gly Glu Phe
210 215 220
Phe Asn Phe Val Leu Thr Gln Glu Gly Ile Asp Val Tyr Asn Ala Ile
225 230 235 240
Ile Gly Gly Phe Val Thr Glu Ser Gly Glu Lys Ile Lys Gly Leu Asn
245 250 255
Glu Tyr Ile Asn Leu Tyr Asn Gln Lys Thr Lys Gln Lys Leu Pro Lys
260 265 270
Phe Lys Pro Leu Tyr Lys Gln Val Leu Ser Asp Arg Glu Ser Leu Ser
275 280 285
Phe Tyr Gly Glu Gly Tyr Thr Ser Asp Glu Glu Val Leu Glu Val Phe
290 295 300
Arg Asn Thr Leu Asn Lys Asn Ser Glu Ile Phe Ser Ser Ile Lys Lys
305 310 315 320
Leu Glu Lys Leu Phe Lys Asn Phe Asp Glu Tyr Ser Ser Ala Gly Ile
325 330 335
Phe Val Lys Asn Gly Pro Ala Ile Ser Thr Ile Ser Lys Asp Ile Phe
340 345 350
Gly Glu Trp Asn Val Ile Arg Asp Lys Trp Asn Ala Glu Tyr Asp Asp
355 360 365
Ile His Leu Lys Lys Lys Ala Val Val Thr Glu Lys Tyr Glu Asp Asp
370 375 380
Arg Arg Lys Ser Phe Lys Lys Ile Gly Ser Phe Ser Leu Glu Gln Leu
385 390 395 400
Gln Glu Tyr Ala Asp Ala Asp Leu Ser Val Val Glu Lys Leu Lys Glu
405 410 415
Ile Ile Ile Gln Lys Val Asp Glu Ile Tyr Lys Val Tyr Gly Ser Ser
420 425 430
Glu Lys Leu Phe Asp Ala Asp Phe Val Leu Glu Lys Ser Leu Lys Lys
435 440 445
Asn Asp Ala Val Val Ala Ile Met Lys Asp Leu Leu Asp Ser Val Lys
450 455 460
Ser Phe Glu Asn Tyr Ile Lys Ala Phe Phe Gly Glu Gly Lys Glu Thr
465 470 475 480
Asn Arg Asp Glu Ser Phe Tyr Gly Asp Phe Val Leu Ala Tyr Asp Ile
485 490 495
Leu Leu Lys Val Asp His Ile Tyr Asp Ala Ile Arg Asn Tyr Val Thr
500 505 510
Gln Lys Pro Tyr Ser Lys Asp Lys Phe Lys Leu Tyr Phe Gln Asn Pro
515 520 525
Gln Phe Met Gly Gly Trp Asp Lys Asp Lys Glu Thr Asp Tyr Arg Ala
530 535 540
Thr Ile Leu Arg Tyr Gly Ser Lys Tyr Tyr Leu Ala Ile Met Asp Lys
545 550 555 560
Lys Tyr Ala Lys Cys Leu Gln Lys Ile Asp Lys Asp Asp Val Asn Gly
565 570 575
Asn Tyr Glu Lys Ile Asn Tyr Lys Leu Leu Pro Gly Pro Asn Lys Met
580 585 590
Leu Pro Lys Val Phe Phe Ser Lys Lys Trp Met Ala Tyr Tyr Asn Pro
595 600 605
Ser Glu Asp Ile Gln Lys Ile Tyr Lys Asn Gly Thr Phe Lys Lys Gly
610 615 620
Asp Met Phe Asn Leu Asn Asp Cys His Lys Leu Ile Asp Phe Phe Lys
625 630 635 640
Asp Ser Ile Ser Arg Tyr Pro Lys Trp Ser Asn Ala Tyr Asp Phe Asn
645 650 655
Phe Ser Glu Thr Glu Lys Tyr Lys Asp Ile Ala Gly Phe Tyr Arg Glu
660 665 670
Val Glu Glu Gln Gly Tyr Lys Val Ser Phe Glu Ser Ala Ser Lys Lys
675 680 685
Glu Val Asp Lys Leu Val Glu Glu Gly Lys Leu Tyr Met Phe Gln Ile
690 695 700
Tyr Asn Lys Asp Phe Ser Asp Lys Ser His Gly Thr Pro Asn Leu His
705 710 715 720
Thr Met Tyr Phe Lys Leu Leu Phe Asp Glu Asn Asn His Gly Gln Ile
725 730 735
Arg Leu Ser Gly Gly Ala Glu Leu Phe Met Arg Arg Ala Ser Leu Lys
740 745 750
Lys Glu Glu Leu Val Val Ala Pro Ala Asn Ser Pro Ile Ala Asn Lys
755 760 765
Asn Pro Asp Asn Pro Lys Lys Thr Thr Thr Leu Ser Tyr Asp Val Tyr
770 775 780
Lys Asp Lys Arg Phe Ser Glu Asp Gln Tyr Glu Leu His Ile Pro Ile
785 790 795 800
Ala Ile Asn Lys Cys Pro Lys Asn Ile Phe Lys Ile Asn Thr Glu Val
805 810 815
Arg Val Leu Leu Lys His Asp Asp Asn Pro Tyr Val Ile Gly Ile Asp
820 825 830
Arg Gly Glu Arg Asn Leu Leu Tyr Ile Val Val Val Asp Gly Lys Gly
835 840 845
Asn Ile Val Glu Gln Tyr Ser Leu Asn Glu Ile Ile Asn Asn Phe Asn
850 855 860
Gly Ile Arg Ile Lys Thr Asp Tyr His Ser Leu Leu Asp Lys Lys Glu
865 870 875 880
Lys Glu Arg Phe Glu Ala Arg Gln Asn Trp Thr Ser Ile Glu Asn Ile
885 890 895
Lys Glu Leu Lys Ala Gly Tyr Ile Ser Gln Val Val His Lys Ile Cys
900 905 910
Glu Leu Val Glu Lys Tyr Asp Ala Val Ile Ala Leu Glu Asp Leu Asn
915 920 925
Ser Gly Phe Lys Asn Ser Arg Val Lys Val Glu Lys Gln Val Tyr Gln
930 935 940
Lys Phe Glu Lys Met Leu Ile Asp Lys Leu Asn Tyr Met Val Asp Lys
945 950 955 960
Lys Ser Asn Pro Cys Ala Thr Gly Gly Ala Leu Lys Gly Tyr Gln Ile
965 970 975
Thr Asn Lys Phe Glu Ser Phe Lys Ser Met Ser Thr Gln Asn Gly Phe
980 985 990
Ile Phe Tyr Ile Pro Ala Trp Leu Thr Ser Lys Ile Asp Pro Ser Thr
995 1000 1005
Gly Phe Val Asn Leu Leu Lys Thr Lys Tyr Thr Ser Ile Ala Asp
1010 1015 1020
Ser Lys Lys Phe Ile Ser Ser Phe Asp Arg Ile Met Tyr Val Pro
1025 1030 1035
Glu Glu Asp Leu Phe Glu Phe Ala Leu Asp Tyr Lys Asn Phe Ser
1040 1045 1050
Arg Thr Asp Ala Asp Tyr Ile Lys Lys Trp Lys Leu Tyr Ser Tyr
1055 1060 1065
Gly Asn Arg Ile Arg Ile Phe Arg Asn Pro Lys Lys Asn Asn Val
1070 1075 1080
Phe Asp Trp Glu Glu Val Cys Leu Thr Ser Ala Tyr Lys Glu Leu
1085 1090 1095
Phe Asn Lys Tyr Gly Ile Asn Tyr Gln Gln Gly Asp Ile Arg Ala
1100 1105 1110
Leu Leu Cys Glu Gln Ser Asp Lys Ala Phe Tyr Ser Ser Phe Met
1115 1120 1125
Ala Leu Met Ser Leu Met Leu Gln Met Arg Asn Ser Ile Thr Gly
1130 1135 1140
Arg Thr Asp Val Asp Phe Leu Ile Ser Pro Val Lys Asn Ser Asp
1145 1150 1155
Gly Ile Phe Tyr Asp Ser Arg Asn Tyr Glu Ala Gln Glu Asn Ala
1160 1165 1170
Ile Leu Pro Lys Asn Ala Asp Ala Asn Gly Ala Tyr Asn Ile Ala
1175 1180 1185
Arg Lys Val Leu Trp Ala Ile Gly Gln Phe Lys Lys Ala Glu Asp
1190 1195 1200
Glu Lys Leu Asp Lys Val Lys Ile Ala Ile Ser Asn Lys Glu Trp
1205 1210 1215
Leu Glu Tyr Ala Gln Thr Ser Val Lys His
1220 1225
<210> 149
<211> 1307
<212> PRT
<213> 人工序列
<220>
<223> 合成的EnAsCas12a H800A
<400> 149
Met Thr Gln Phe Glu Gly Phe Thr Asn Leu Tyr Gln Val Ser Lys Thr
1 5 10 15
Leu Arg Phe Glu Leu Ile Pro Gln Gly Lys Thr Leu Lys His Ile Gln
20 25 30
Glu Gln Gly Phe Ile Glu Glu Asp Lys Ala Arg Asn Asp His Tyr Lys
35 40 45
Glu Leu Lys Pro Ile Ile Asp Arg Ile Tyr Lys Thr Tyr Ala Asp Gln
50 55 60
Cys Leu Gln Leu Val Gln Leu Asp Trp Glu Asn Leu Ser Ala Ala Ile
65 70 75 80
Asp Ser Tyr Arg Lys Glu Lys Thr Glu Glu Thr Arg Asn Ala Leu Ile
85 90 95
Glu Glu Gln Ala Thr Tyr Arg Asn Ala Ile His Asp Tyr Phe Ile Gly
100 105 110
Arg Thr Asp Asn Leu Thr Asp Ala Ile Asn Lys Arg His Ala Glu Ile
115 120 125
Tyr Lys Gly Leu Phe Lys Ala Glu Leu Phe Asn Gly Lys Val Leu Lys
130 135 140
Gln Leu Gly Thr Val Thr Thr Thr Glu His Glu Asn Ala Leu Leu Arg
145 150 155 160
Ser Phe Asp Lys Phe Thr Thr Tyr Phe Ser Gly Phe Tyr Arg Asn Arg
165 170 175
Lys Asn Val Phe Ser Ala Glu Asp Ile Ser Thr Ala Ile Pro His Arg
180 185 190
Ile Val Gln Asp Asn Phe Pro Lys Phe Lys Glu Asn Cys His Ile Phe
195 200 205
Thr Arg Leu Ile Thr Ala Val Pro Ser Leu Arg Glu His Phe Glu Asn
210 215 220
Val Lys Lys Ala Ile Gly Ile Phe Val Ser Thr Ser Ile Glu Glu Val
225 230 235 240
Phe Ser Phe Pro Phe Tyr Asn Gln Leu Leu Thr Gln Thr Gln Ile Asp
245 250 255
Leu Tyr Asn Gln Leu Leu Gly Gly Ile Ser Arg Glu Ala Gly Thr Glu
260 265 270
Lys Ile Lys Gly Leu Asn Glu Val Leu Asn Leu Ala Ile Gln Lys Asn
275 280 285
Asp Glu Thr Ala His Ile Ile Ala Ser Leu Pro His Arg Phe Ile Pro
290 295 300
Leu Phe Lys Gln Ile Leu Ser Asp Arg Asn Thr Leu Ser Phe Ile Leu
305 310 315 320
Glu Glu Phe Lys Ser Asp Glu Glu Val Ile Gln Ser Phe Cys Lys Tyr
325 330 335
Lys Thr Leu Leu Arg Asn Glu Asn Val Leu Glu Thr Ala Glu Ala Leu
340 345 350
Phe Asn Glu Leu Asn Ser Ile Asp Leu Thr His Ile Phe Ile Ser His
355 360 365
Lys Lys Leu Glu Thr Ile Ser Ser Ala Leu Cys Asp His Trp Asp Thr
370 375 380
Leu Arg Asn Ala Leu Tyr Glu Arg Arg Ile Ser Glu Leu Thr Gly Lys
385 390 395 400
Ile Thr Lys Ser Ala Lys Glu Lys Val Gln Arg Ser Leu Lys His Glu
405 410 415
Asp Ile Asn Leu Gln Glu Ile Ile Ser Ala Ala Gly Lys Glu Leu Ser
420 425 430
Glu Ala Phe Lys Gln Lys Thr Ser Glu Ile Leu Ser His Ala His Ala
435 440 445
Ala Leu Asp Gln Pro Leu Pro Thr Thr Leu Lys Lys Gln Glu Glu Lys
450 455 460
Glu Ile Leu Lys Ser Gln Leu Asp Ser Leu Leu Gly Leu Tyr His Leu
465 470 475 480
Leu Asp Trp Phe Ala Val Asp Glu Ser Asn Glu Val Asp Pro Glu Phe
485 490 495
Ser Ala Arg Leu Thr Gly Ile Lys Leu Glu Met Glu Pro Ser Leu Ser
500 505 510
Phe Tyr Asn Lys Ala Arg Asn Tyr Ala Thr Lys Lys Pro Tyr Ser Val
515 520 525
Glu Lys Phe Lys Leu Asn Phe Gln Met Pro Thr Leu Ala Arg Gly Trp
530 535 540
Asp Val Asn Arg Glu Lys Asn Asn Gly Ala Ile Leu Phe Val Lys Asn
545 550 555 560
Gly Leu Tyr Tyr Leu Gly Ile Met Pro Lys Gln Lys Gly Arg Tyr Lys
565 570 575
Ala Leu Ser Phe Glu Pro Thr Glu Lys Thr Ser Glu Gly Phe Asp Lys
580 585 590
Met Tyr Tyr Asp Tyr Phe Pro Asp Ala Ala Lys Met Ile Pro Lys Cys
595 600 605
Ser Thr Gln Leu Lys Ala Val Thr Ala His Phe Gln Thr His Thr Thr
610 615 620
Pro Ile Leu Leu Ser Asn Asn Phe Ile Glu Pro Leu Glu Ile Thr Lys
625 630 635 640
Glu Ile Tyr Asp Leu Asn Asn Pro Glu Lys Glu Pro Lys Lys Phe Gln
645 650 655
Thr Ala Tyr Ala Lys Lys Thr Gly Asp Gln Lys Gly Tyr Arg Glu Ala
660 665 670
Leu Cys Lys Trp Ile Asp Phe Thr Arg Asp Phe Leu Ser Lys Tyr Thr
675 680 685
Lys Thr Thr Ser Ile Asp Leu Ser Ser Leu Arg Pro Ser Ser Gln Tyr
690 695 700
Lys Asp Leu Gly Glu Tyr Tyr Ala Glu Leu Asn Pro Leu Leu Tyr His
705 710 715 720
Ile Ser Phe Gln Arg Ile Ala Glu Lys Glu Ile Met Asp Ala Val Glu
725 730 735
Thr Gly Lys Leu Tyr Leu Phe Gln Ile Tyr Asn Lys Asp Phe Ala Lys
740 745 750
Gly His His Gly Lys Pro Asn Leu His Thr Leu Tyr Trp Thr Gly Leu
755 760 765
Phe Ser Pro Glu Asn Leu Ala Lys Thr Ser Ile Lys Leu Asn Gly Gln
770 775 780
Ala Glu Leu Phe Tyr Arg Pro Lys Ser Arg Met Lys Arg Met Ala Ala
785 790 795 800
Arg Leu Gly Glu Lys Met Leu Asn Lys Lys Leu Lys Asp Gln Lys Thr
805 810 815
Pro Ile Pro Asp Thr Leu Tyr Gln Glu Leu Tyr Asp Tyr Val Asn His
820 825 830
Arg Leu Ser His Asp Leu Ser Asp Glu Ala Arg Ala Leu Leu Pro Asn
835 840 845
Val Ile Thr Lys Glu Val Ser His Glu Ile Ile Lys Asp Arg Arg Phe
850 855 860
Thr Ser Asp Lys Phe Phe Phe His Val Pro Ile Thr Leu Asn Tyr Gln
865 870 875 880
Ala Ala Asn Ser Pro Ser Lys Phe Asn Gln Arg Val Asn Ala Tyr Leu
885 890 895
Lys Glu His Pro Glu Thr Pro Ile Ile Gly Ile Asp Arg Gly Glu Arg
900 905 910
Asn Leu Ile Tyr Ile Thr Val Ile Asp Ser Thr Gly Lys Ile Leu Glu
915 920 925
Gln Arg Ser Leu Asn Thr Ile Gln Gln Phe Asp Tyr Gln Lys Lys Leu
930 935 940
Asp Asn Arg Glu Lys Glu Arg Val Ala Ala Arg Gln Ala Trp Ser Val
945 950 955 960
Val Gly Thr Ile Lys Asp Leu Lys Gln Gly Tyr Leu Ser Gln Val Ile
965 970 975
His Glu Ile Val Asp Leu Met Ile His Tyr Gln Ala Val Val Val Leu
980 985 990
Glu Asn Leu Asn Phe Gly Phe Lys Ser Lys Arg Thr Gly Ile Ala Glu
995 1000 1005
Lys Ala Val Tyr Gln Gln Phe Glu Lys Met Leu Ile Asp Lys Leu
1010 1015 1020
Asn Cys Leu Val Leu Lys Asp Tyr Pro Ala Glu Lys Val Gly Gly
1025 1030 1035
Val Leu Asn Pro Tyr Gln Leu Thr Asp Gln Phe Thr Ser Phe Ala
1040 1045 1050
Lys Met Gly Thr Gln Ser Gly Phe Leu Phe Tyr Val Pro Ala Pro
1055 1060 1065
Tyr Thr Ser Lys Ile Asp Pro Leu Thr Gly Phe Val Asp Pro Phe
1070 1075 1080
Val Trp Lys Thr Ile Lys Asn His Glu Ser Arg Lys His Phe Leu
1085 1090 1095
Glu Gly Phe Asp Phe Leu His Tyr Asp Val Lys Thr Gly Asp Phe
1100 1105 1110
Ile Leu His Phe Lys Met Asn Arg Asn Leu Ser Phe Gln Arg Gly
1115 1120 1125
Leu Pro Gly Phe Met Pro Ala Trp Asp Ile Val Phe Glu Lys Asn
1130 1135 1140
Glu Thr Gln Phe Asp Ala Lys Gly Thr Pro Phe Ile Ala Gly Lys
1145 1150 1155
Arg Ile Val Pro Val Ile Glu Asn His Arg Phe Thr Gly Arg Tyr
1160 1165 1170
Arg Asp Leu Tyr Pro Ala Asn Glu Leu Ile Ala Leu Leu Glu Glu
1175 1180 1185
Lys Gly Ile Val Phe Arg Asp Gly Ser Asn Ile Leu Pro Lys Leu
1190 1195 1200
Leu Glu Asn Asp Asp Ser His Ala Ile Asp Thr Met Val Ala Leu
1205 1210 1215
Ile Arg Ser Val Leu Gln Met Arg Asn Ser Asn Ala Ala Thr Gly
1220 1225 1230
Glu Asp Tyr Ile Asn Ser Pro Val Arg Asp Leu Asn Gly Val Cys
1235 1240 1245
Phe Asp Ser Arg Phe Gln Asn Pro Glu Trp Pro Met Asp Ala Asp
1250 1255 1260
Ala Asn Gly Ala Tyr His Ile Ala Leu Lys Gly Gln Leu Leu Leu
1265 1270 1275
Asn His Leu Lys Glu Ser Lys Asp Leu Lys Leu Gln Asn Gly Ile
1280 1285 1290
Ser Asn Gln Asp Trp Leu Ala Tyr Ile Gln Glu Leu Arg Asn
1295 1300 1305
<210> 150
<211> 1307
<212> PRT
<213> 人工序列
<220>
<223> 合成的EnAsCas12a
<400> 150
Met Thr Gln Phe Glu Gly Phe Thr Asn Leu Tyr Gln Val Ser Lys Thr
1 5 10 15
Leu Arg Phe Glu Leu Ile Pro Gln Gly Lys Thr Leu Lys His Ile Gln
20 25 30
Glu Gln Gly Phe Ile Glu Glu Asp Lys Ala Arg Asn Asp His Tyr Lys
35 40 45
Glu Leu Lys Pro Ile Ile Asp Arg Ile Tyr Lys Thr Tyr Ala Asp Gln
50 55 60
Cys Leu Gln Leu Val Gln Leu Asp Trp Glu Asn Leu Ser Ala Ala Ile
65 70 75 80
Asp Ser Tyr Arg Lys Glu Lys Thr Glu Glu Thr Arg Asn Ala Leu Ile
85 90 95
Glu Glu Gln Ala Thr Tyr Arg Asn Ala Ile His Asp Tyr Phe Ile Gly
100 105 110
Arg Thr Asp Asn Leu Thr Asp Ala Ile Asn Lys Arg His Ala Glu Ile
115 120 125
Tyr Lys Gly Leu Phe Lys Ala Glu Leu Phe Asn Gly Lys Val Leu Lys
130 135 140
Gln Leu Gly Thr Val Thr Thr Thr Glu His Glu Asn Ala Leu Leu Arg
145 150 155 160
Ser Phe Asp Lys Phe Thr Thr Tyr Phe Ser Gly Phe Tyr Arg Asn Arg
165 170 175
Lys Asn Val Phe Ser Ala Glu Asp Ile Ser Thr Ala Ile Pro His Arg
180 185 190
Ile Val Gln Asp Asn Phe Pro Lys Phe Lys Glu Asn Cys His Ile Phe
195 200 205
Thr Arg Leu Ile Thr Ala Val Pro Ser Leu Arg Glu His Phe Glu Asn
210 215 220
Val Lys Lys Ala Ile Gly Ile Phe Val Ser Thr Ser Ile Glu Glu Val
225 230 235 240
Phe Ser Phe Pro Phe Tyr Asn Gln Leu Leu Thr Gln Thr Gln Ile Asp
245 250 255
Leu Tyr Asn Gln Leu Leu Gly Gly Ile Ser Arg Glu Ala Gly Thr Glu
260 265 270
Lys Ile Lys Gly Leu Asn Glu Val Leu Asn Leu Ala Ile Gln Lys Asn
275 280 285
Asp Glu Thr Ala His Ile Ile Ala Ser Leu Pro His Arg Phe Ile Pro
290 295 300
Leu Phe Lys Gln Ile Leu Ser Asp Arg Asn Thr Leu Ser Phe Ile Leu
305 310 315 320
Glu Glu Phe Lys Ser Asp Glu Glu Val Ile Gln Ser Phe Cys Lys Tyr
325 330 335
Lys Thr Leu Leu Arg Asn Glu Asn Val Leu Glu Thr Ala Glu Ala Leu
340 345 350
Phe Asn Glu Leu Asn Ser Ile Asp Leu Thr His Ile Phe Ile Ser His
355 360 365
Lys Lys Leu Glu Thr Ile Ser Ser Ala Leu Cys Asp His Trp Asp Thr
370 375 380
Leu Arg Asn Ala Leu Tyr Glu Arg Arg Ile Ser Glu Leu Thr Gly Lys
385 390 395 400
Ile Thr Lys Ser Ala Lys Glu Lys Val Gln Arg Ser Leu Lys His Glu
405 410 415
Asp Ile Asn Leu Gln Glu Ile Ile Ser Ala Ala Gly Lys Glu Leu Ser
420 425 430
Glu Ala Phe Lys Gln Lys Thr Ser Glu Ile Leu Ser His Ala His Ala
435 440 445
Ala Leu Asp Gln Pro Leu Pro Thr Thr Leu Lys Lys Gln Glu Glu Lys
450 455 460
Glu Ile Leu Lys Ser Gln Leu Asp Ser Leu Leu Gly Leu Tyr His Leu
465 470 475 480
Leu Asp Trp Phe Ala Val Asp Glu Ser Asn Glu Val Asp Pro Glu Phe
485 490 495
Ser Ala Arg Leu Thr Gly Ile Lys Leu Glu Met Glu Pro Ser Leu Ser
500 505 510
Phe Tyr Asn Lys Ala Arg Asn Tyr Ala Thr Lys Lys Pro Tyr Ser Val
515 520 525
Glu Lys Phe Lys Leu Asn Phe Gln Met Pro Thr Leu Ala Arg Gly Trp
530 535 540
Asp Val Asn Arg Glu Lys Asn Asn Gly Ala Ile Leu Phe Val Lys Asn
545 550 555 560
Gly Leu Tyr Tyr Leu Gly Ile Met Pro Lys Gln Lys Gly Arg Tyr Lys
565 570 575
Ala Leu Ser Phe Glu Pro Thr Glu Lys Thr Ser Glu Gly Phe Asp Lys
580 585 590
Met Tyr Tyr Asp Tyr Phe Pro Asp Ala Ala Lys Met Ile Pro Lys Cys
595 600 605
Ser Thr Gln Leu Lys Ala Val Thr Ala His Phe Gln Thr His Thr Thr
610 615 620
Pro Ile Leu Leu Ser Asn Asn Phe Ile Glu Pro Leu Glu Ile Thr Lys
625 630 635 640
Glu Ile Tyr Asp Leu Asn Asn Pro Glu Lys Glu Pro Lys Lys Phe Gln
645 650 655
Thr Ala Tyr Ala Lys Lys Thr Gly Asp Gln Lys Gly Tyr Arg Glu Ala
660 665 670
Leu Cys Lys Trp Ile Asp Phe Thr Arg Asp Phe Leu Ser Lys Tyr Thr
675 680 685
Lys Thr Thr Ser Ile Asp Leu Ser Ser Leu Arg Pro Ser Ser Gln Tyr
690 695 700
Lys Asp Leu Gly Glu Tyr Tyr Ala Glu Leu Asn Pro Leu Leu Tyr His
705 710 715 720
Ile Ser Phe Gln Arg Ile Ala Glu Lys Glu Ile Met Asp Ala Val Glu
725 730 735
Thr Gly Lys Leu Tyr Leu Phe Gln Ile Tyr Asn Lys Asp Phe Ala Lys
740 745 750
Gly His His Gly Lys Pro Asn Leu His Thr Leu Tyr Trp Thr Gly Leu
755 760 765
Phe Ser Pro Glu Asn Leu Ala Lys Thr Ser Ile Lys Leu Asn Gly Gln
770 775 780
Ala Glu Leu Phe Tyr Arg Pro Lys Ser Arg Met Lys Arg Met Ala His
785 790 795 800
Arg Leu Gly Glu Lys Met Leu Asn Lys Lys Leu Lys Asp Gln Lys Thr
805 810 815
Pro Ile Pro Asp Thr Leu Tyr Gln Glu Leu Tyr Asp Tyr Val Asn His
820 825 830
Arg Leu Ser His Asp Leu Ser Asp Glu Ala Arg Ala Leu Leu Pro Asn
835 840 845
Val Ile Thr Lys Glu Val Ser His Glu Ile Ile Lys Asp Arg Arg Phe
850 855 860
Thr Ser Asp Lys Phe Phe Phe His Val Pro Ile Thr Leu Asn Tyr Gln
865 870 875 880
Ala Ala Asn Ser Pro Ser Lys Phe Asn Gln Arg Val Asn Ala Tyr Leu
885 890 895
Lys Glu His Pro Glu Thr Pro Ile Ile Gly Ile Asp Arg Gly Glu Arg
900 905 910
Asn Leu Ile Tyr Ile Thr Val Ile Asp Ser Thr Gly Lys Ile Leu Glu
915 920 925
Gln Arg Ser Leu Asn Thr Ile Gln Gln Phe Asp Tyr Gln Lys Lys Leu
930 935 940
Asp Asn Arg Glu Lys Glu Arg Val Ala Ala Arg Gln Ala Trp Ser Val
945 950 955 960
Val Gly Thr Ile Lys Asp Leu Lys Gln Gly Tyr Leu Ser Gln Val Ile
965 970 975
His Glu Ile Val Asp Leu Met Ile His Tyr Gln Ala Val Val Val Leu
980 985 990
Glu Asn Leu Asn Phe Gly Phe Lys Ser Lys Arg Thr Gly Ile Ala Glu
995 1000 1005
Lys Ala Val Tyr Gln Gln Phe Glu Lys Met Leu Ile Asp Lys Leu
1010 1015 1020
Asn Cys Leu Val Leu Lys Asp Tyr Pro Ala Glu Lys Val Gly Gly
1025 1030 1035
Val Leu Asn Pro Tyr Gln Leu Thr Asp Gln Phe Thr Ser Phe Ala
1040 1045 1050
Lys Met Gly Thr Gln Ser Gly Phe Leu Phe Tyr Val Pro Ala Pro
1055 1060 1065
Tyr Thr Ser Lys Ile Asp Pro Leu Thr Gly Phe Val Asp Pro Phe
1070 1075 1080
Val Trp Lys Thr Ile Lys Asn His Glu Ser Arg Lys His Phe Leu
1085 1090 1095
Glu Gly Phe Asp Phe Leu His Tyr Asp Val Lys Thr Gly Asp Phe
1100 1105 1110
Ile Leu His Phe Lys Met Asn Arg Asn Leu Ser Phe Gln Arg Gly
1115 1120 1125
Leu Pro Gly Phe Met Pro Ala Trp Asp Ile Val Phe Glu Lys Asn
1130 1135 1140
Glu Thr Gln Phe Asp Ala Lys Gly Thr Pro Phe Ile Ala Gly Lys
1145 1150 1155
Arg Ile Val Pro Val Ile Glu Asn His Arg Phe Thr Gly Arg Tyr
1160 1165 1170
Arg Asp Leu Tyr Pro Ala Asn Glu Leu Ile Ala Leu Leu Glu Glu
1175 1180 1185
Lys Gly Ile Val Phe Arg Asp Gly Ser Asn Ile Leu Pro Lys Leu
1190 1195 1200
Leu Glu Asn Asp Asp Ser His Ala Ile Asp Thr Met Val Ala Leu
1205 1210 1215
Ile Arg Ser Val Leu Gln Met Arg Asn Ser Asn Ala Ala Thr Gly
1220 1225 1230
Glu Asp Tyr Ile Asn Ser Pro Val Arg Asp Leu Asn Gly Val Cys
1235 1240 1245
Phe Asp Ser Arg Phe Gln Asn Pro Glu Trp Pro Met Asp Ala Asp
1250 1255 1260
Ala Asn Gly Ala Tyr His Ile Ala Leu Lys Gly Gln Leu Leu Leu
1265 1270 1275
Asn His Leu Lys Glu Ser Lys Asp Leu Lys Leu Gln Asn Gly Ile
1280 1285 1290
Ser Asn Gln Asp Trp Leu Ala Tyr Ile Gln Glu Leu Arg Asn
1295 1300 1305
<210> 151
<211> 1107
<212> PRT
<213> 久志氏芽孢杆菌（Bacillus hisashii）
<400> 151
Ala Thr Arg Ser Phe Ile Leu Lys Ile Glu Pro Asn Glu Glu Val Lys
1 5 10 15
Lys Gly Leu Trp Lys Thr His Glu Val Leu Asn His Gly Ile Ala Tyr
20 25 30
Tyr Met Asn Ile Leu Lys Leu Ile Arg Gln Glu Ala Ile Tyr Glu His
35 40 45
His Glu Gln Asp Pro Lys Asn Pro Lys Lys Val Ser Lys Ala Glu Ile
50 55 60
Gln Ala Glu Leu Trp Asp Phe Val Leu Lys Met Gln Lys Cys Asn Ser
65 70 75 80
Phe Thr His Glu Val Asp Lys Asp Glu Val Phe Asn Ile Leu Arg Glu
85 90 95
Leu Tyr Glu Glu Leu Val Pro Ser Ser Val Glu Lys Lys Gly Glu Ala
100 105 110
Asn Gln Leu Ser Asn Lys Phe Leu Tyr Pro Leu Val Asp Pro Asn Ser
115 120 125
Gln Ser Gly Lys Gly Thr Ala Ser Ser Gly Arg Lys Pro Arg Trp Tyr
130 135 140
Asn Leu Lys Ile Ala Gly Asp Pro Ser Trp Glu Glu Glu Lys Lys Lys
145 150 155 160
Trp Glu Glu Asp Lys Lys Lys Asp Pro Leu Ala Lys Ile Leu Gly Lys
165 170 175
Leu Ala Glu Tyr Gly Leu Ile Pro Leu Phe Ile Pro Tyr Thr Asp Ser
180 185 190
Asn Glu Pro Ile Val Lys Glu Ile Lys Trp Met Glu Lys Ser Arg Asn
195 200 205
Gln Ser Val Arg Arg Leu Asp Lys Asp Met Phe Ile Gln Ala Leu Glu
210 215 220
Arg Phe Leu Ser Trp Glu Ser Trp Asn Leu Lys Val Lys Glu Glu Tyr
225 230 235 240
Glu Lys Val Glu Lys Glu Tyr Lys Thr Leu Glu Glu Arg Ile Lys Glu
245 250 255
Asp Ile Gln Ala Leu Lys Ala Leu Glu Gln Tyr Glu Lys Glu Arg Gln
260 265 270
Glu Gln Leu Leu Arg Asp Thr Leu Asn Thr Asn Glu Tyr Arg Leu Ser
275 280 285
Lys Arg Gly Leu Arg Gly Trp Arg Glu Ile Ile Gln Lys Trp Leu Lys
290 295 300
Met Asp Glu Asn Glu Pro Ser Glu Lys Tyr Leu Glu Val Phe Lys Asp
305 310 315 320
Tyr Gln Arg Lys His Pro Arg Glu Ala Gly Asp Tyr Ser Val Tyr Glu
325 330 335
Phe Leu Ser Lys Lys Glu Asn His Phe Ile Trp Arg Asn His Pro Glu
340 345 350
Tyr Pro Tyr Leu Tyr Ala Thr Phe Cys Glu Ile Asp Lys Lys Lys Lys
355 360 365
Asp Ala Lys Gln Gln Ala Thr Phe Thr Leu Ala Asp Pro Ile Asn His
370 375 380
Pro Leu Trp Val Arg Phe Glu Glu Arg Ser Gly Ser Asn Leu Asn Lys
385 390 395 400
Tyr Arg Ile Leu Thr Glu Gln Leu His Thr Glu Lys Leu Lys Lys Lys
405 410 415
Leu Thr Val Gln Leu Asp Arg Leu Ile Tyr Pro Thr Glu Ser Gly Gly
420 425 430
Trp Glu Glu Lys Gly Lys Val Asp Ile Val Leu Leu Pro Ser Arg Gln
435 440 445
Phe Tyr Asn Gln Ile Phe Leu Asp Ile Glu Glu Lys Gly Lys His Ala
450 455 460
Phe Thr Tyr Lys Asp Glu Ser Ile Lys Phe Pro Leu Lys Gly Thr Leu
465 470 475 480
Gly Gly Ala Arg Val Gln Phe Asp Arg Asp His Leu Arg Arg Tyr Pro
485 490 495
His Lys Val Glu Ser Gly Asn Val Gly Arg Ile Tyr Phe Asn Met Thr
500 505 510
Val Asn Ile Glu Pro Thr Glu Ser Pro Val Ser Lys Ser Leu Lys Ile
515 520 525
His Arg Asp Asp Phe Pro Lys Val Val Asn Phe Lys Pro Lys Glu Leu
530 535 540
Thr Glu Trp Ile Lys Asp Ser Lys Gly Lys Lys Leu Lys Ser Gly Ile
545 550 555 560
Glu Ser Leu Glu Ile Gly Leu Arg Val Met Ser Ile Asp Leu Gly Gln
565 570 575
Arg Gln Ala Ala Ala Ala Ser Ile Phe Glu Val Val Asp Gln Lys Pro
580 585 590
Asp Ile Glu Gly Lys Leu Phe Phe Pro Ile Lys Gly Thr Glu Leu Tyr
595 600 605
Ala Val His Arg Ala Ser Phe Asn Ile Lys Leu Pro Gly Glu Thr Leu
610 615 620
Val Lys Ser Arg Glu Val Leu Arg Lys Ala Arg Glu Asp Asn Leu Lys
625 630 635 640
Leu Met Asn Gln Lys Leu Asn Phe Leu Arg Asn Val Leu His Phe Gln
645 650 655
Gln Phe Glu Asp Ile Thr Glu Arg Glu Lys Arg Val Thr Lys Trp Ile
660 665 670
Ser Arg Gln Glu Asn Ser Asp Val Pro Leu Val Tyr Gln Asp Glu Leu
675 680 685
Ile Gln Ile Arg Glu Leu Met Tyr Lys Pro Tyr Lys Asp Trp Val Ala
690 695 700
Phe Leu Lys Gln Leu His Lys Arg Leu Glu Val Glu Ile Gly Lys Glu
705 710 715 720
Val Lys His Trp Arg Lys Ser Leu Ser Asp Gly Arg Lys Gly Leu Tyr
725 730 735
Gly Ile Ser Leu Lys Asn Ile Asp Glu Ile Asp Arg Thr Arg Lys Phe
740 745 750
Leu Leu Arg Trp Ser Leu Arg Pro Thr Glu Pro Gly Glu Val Arg Arg
755 760 765
Leu Glu Pro Gly Gln Arg Phe Ala Ile Asp Gln Leu Asn His Leu Asn
770 775 780
Ala Leu Lys Glu Asp Arg Leu Lys Lys Met Ala Asn Thr Ile Ile Met
785 790 795 800
His Ala Leu Gly Tyr Cys Tyr Asp Val Arg Lys Lys Lys Trp Gln Ala
805 810 815
Lys Asn Pro Ala Cys Gln Ile Ile Leu Phe Glu Asp Leu Ser Asn Tyr
820 825 830
Asn Pro Tyr Gly Glu Arg Ser Arg Phe Glu Asn Ser Arg Leu Met Lys
835 840 845
Trp Ser Arg Arg Glu Ile Pro Arg Gln Val Ala Leu Gln Gly Glu Ile
850 855 860
Tyr Gly Leu Gln Val Gly Glu Val Gly Ala Gln Phe Ser Ser Arg Phe
865 870 875 880
His Ala Lys Thr Gly Ser Pro Gly Ile Arg Cys Arg Val Val Thr Lys
885 890 895
Glu Lys Leu Gln Asp Asn Arg Phe Phe Lys Asn Leu Gln Arg Glu Gly
900 905 910
Arg Leu Thr Leu Asp Lys Ile Ala Val Leu Lys Glu Gly Asp Leu Tyr
915 920 925
Pro Asp Lys Gly Gly Glu Lys Phe Ile Ser Leu Ser Lys Asp Arg Lys
930 935 940
Cys Val Thr Thr His Ala Asp Ile Asn Ala Ala Gln Asn Leu Gln Lys
945 950 955 960
Arg Phe Trp Thr Arg Thr His Gly Phe Tyr Lys Val Tyr Cys Lys Ala
965 970 975
Tyr Gln Val Asp Gly Gln Thr Val Tyr Ile Pro Glu Ser Lys Asp Gln
980 985 990
Lys Gln Lys Ile Ile Glu Glu Phe Gly Glu Gly Tyr Phe Ile Leu Lys
995 1000 1005
Asp Gly Val Tyr Glu Trp Val Asn Ala Gly Lys Leu Lys Ile Lys
1010 1015 1020
Lys Gly Ser Ser Lys Gln Ser Ser Ser Glu Leu Val Asp Ser Asp
1025 1030 1035
Ile Leu Lys Asp Ser Phe Asp Leu Ala Ser Glu Leu Lys Gly Glu
1040 1045 1050
Lys Leu Met Leu Tyr Arg Asp Pro Ser Gly Asn Val Phe Pro Ser
1055 1060 1065
Asp Lys Trp Met Ala Ala Gly Val Phe Phe Gly Lys Leu Glu Arg
1070 1075 1080
Ile Leu Ile Ser Lys Leu Thr Asn Gln Tyr Ser Ile Ser Thr Ile
1085 1090 1095
Glu Asp Asp Ser Ser Lys Gln Ser Met
1100 1105
<210> 152
<211> 46
<212> PRT
<213> 智人
<400> 152
Met Gly Ile Ile Asn Thr Leu Gln Lys Tyr Tyr Cys Arg Val Arg Gly
1 5 10 15
Gly Arg Cys Ala Val Leu Ser Cys Leu Pro Lys Glu Glu Gln Ile Gly
20 25 30
Lys Cys Ser Thr Arg Gly Arg Lys Cys Cys Arg Arg Lys Lys
35 40 45
<210> 153
<211> 90
<212> PRT
<213> 葡萄藤病毒A
<400> 153
Met Asp Asp Pro Ser Phe Leu Thr Gly Arg Ser Thr Tyr Ala Lys Arg
1 5 10 15
Arg Arg Ala Arg Arg Met Asn Val Cys Lys Cys Gly Ala Ile Leu His
20 25 30
Asn Asn Lys Asp Cys Arg Ser Ser Thr Ile Ser Gly His Lys Leu Asp
35 40 45
Arg Leu Arg Phe Val Lys Glu Gly Arg Val Ala Leu Glu Gly Glu Thr
50 55 60
Pro Val Tyr Arg Thr Trp Val Lys Trp Val Glu Thr Glu Tyr His Ile
65 70 75 80
Asn Ile Leu Glu Thr Ser Asp Asp Glu Glu
85 90
<210> 154
<211> 6048
<212> DNA
<213> 人工序列
<220>
<223> 合成的在N-末端处融合的REDRAW编辑器
<400> 154
atggcaactt caagaaagag gcctagtgac ttggttcacg tattctctcc tccacgaaag 60
aaggctggtt cggctacatc aagaaagcgt cccagcgatt tagtgcatgt cttctctccg 120
ccaagaaaga aggccggatc cactctcaat atcgaagacg aacatcgttt acatgaaaca 180
tctaaagaac cggacgtttc tttaggttcc acatggttaa gcgacttccc tcaggcctgg 240
gcagaaacag gtggtatggg acttgccgtt cgacaagcgc ctttgatcat acccctaaag 300
gcgacaagta ccccagtatc cattaaacag tatcctatgt cccaggaggc acgcttgggg 360
ataaagcccc atatccaaag actattagat caggggatac ttgtcccatg ccaaagccct 420
tggaacactc cgttactgcc cgtgaaaaaa ccaggtacaa acgattatag acctgtgcaa 480
gacctgcgcg aggtgaacaa gagggtagag gatatacatc caacggttcc aaatccttat 540
aatttactat ctggtcttcc acctagtcac cagtggtata ccgtcttgga cttgaaggat 600
gcgtttttct gcttgcgttt acatccgacg tctcaacctt tattcgcatt cgagtggcga 660
gatcctgaga tgggtatctc tggtcagtta acttggacaa gattgcctca aggttttaag 720
aatagtccaa ctttatttaa tgaagccttg catagagatt tggctgattt taggattcaa 780
catcctgacc taattttgct acaatacgtt gatgatctat tgttggctgc cacctcggaa 840
cttgattgtc aacagggtac gagggctctg cttcaaacct taggaaactt gggataccga 900
gcttctgcta aaaaggcgca aatttgtcaa aagcaggtaa aatacttggg ttacctattg 960
aaagaagggc aaaggtggct aactgaggcc cgtaaagaaa ccgtgatggg tcaaccaacc 1020
cccaaaacac caagacaact cagggaattt ttaggcaaag cgggtttctg tagattgttt 1080
attcctggct ttgcagaaat ggccgcacca ctgtatccac tgacaaaacc aggcaccttg 1140
ttcaactggg gcccagatca gcagaaggct tatcaagaaa ttaaacaggc tttgctaaca 1200
gcaccagcac taggtctacc agatttaact aaacccttcg aattgtttgt tgatgaaaaa 1260
caaggatatg ctaaaggtgt cctgactcaa aaattgggac cctggagaag accggttgca 1320
tatttaagca aaaagttgga cccagtagcc gctgggtggc caccgtgttt aaggatggtt 1380
gctgctatcg ctgtcttaac gaaggatgcc ggcaaattaa cgatgggaca gccactggtt 1440
atactagctc cccatgctgt tgaagccttg gtgaaacaac ctcccgaccg ttggttatca 1500
aatgcaagaa tgacacacta tcaagccttg ttgctcgaca cagacagagt tcaatttgga 1560
cctgtagtcg cactaaatcc ggcgactctt ttaccactgc cagaagaggg tcttcaacat 1620
aactgcctcg atattttggc cgaagctcat gggaccagac cggacttaac agatcaacct 1680
ttaccagatg cagatcacac ttggtacact gatggtagtt cactccttca agagggtcaa 1740
cgtaaagctg gcgcagcagt taccactgaa acggaagtaa tttgggcaaa agcattacct 1800
gctggtactt cagcccaaag agctgagttg attgccttga ctcaagctct taaaatggct 1860
gaaggcaaga agctgaatgt ctatactgac tcaagatacg cttttgctac tgctcatatt 1920
cacggtgaaa tctacagaag aagaggatgg cttacgagtg aaggaaaaga aattaagaat 1980
aaggatgaga tattagcact tctcaaagca ttgtttctgc caaagagact ttcaattatt 2040
cactgtccag gacatcaaaa gggccactcc gccgaagcta ggggtaatag aatggcagat 2100
caggcagcga ggaaggctgc catcacagag actcctgata catctacctt attaatagaa 2160
aactcgtcgc caaattcacg gttaataaac tcaggtggat cgtccggtgg cagttctggt 2220
tcagaaactc ctggtacatc tgagagcgct accccagaat ccagtggagg gagctcaggg 2280
ggttctagca aactcgaaaa gttcactaat tgttactctc tatcaaagac cttaagattt 2340
aaggctatac ctgtagggaa gactcaagaa aacatcgata ataagaggtt gttagttgaa 2400
gatgaaaaga gggctgaaga ctacaagggt gtaaagaaat tgttggaccg atactacctt 2460
agcttcatca acgatgtctt gcattcaata aagttgaaaa atttaaataa ctacatctcg 2520
ctattccgta agaaaacgag gaccgaaaag gagaacaaag aactagaaaa cttggaaatc 2580
aacctgagaa aggagatagc caaagccttc aagggtaatg agggttacaa gtctcttttt 2640
aagaaagata ttatcgaaac tatactacca gaattcttag atgacaaaga cgagatcgcc 2700
ctggtaaact cttttaatgg tttcacaact gcgttcacag gtttctttga caatcgcgag 2760
aatatgttct ctgaagaggc taagagtaca tcaatcgcat tcaggtgcat caacgaaaac 2820
ttgacgaggt acatttcgaa catggacata tttgaaaagg ttgacgcaat cttcgacaag 2880
catgaagtac aagaaatcaa ggaaaagatc ttaaattcgg attacgatgt ggaggatttc 2940
ttcgaaggtg aattctttaa cttcgtcttg acccaagagg gtatcgatgt ctacaatgca 3000
atcataggag gtttcgttac cgaatccggc gagaagatca aaggtctaaa cgaatatatc 3060
aatctgtata atcaaaagac caagcagaaa ctacccaaat tcaagcctct ctacaagcaa 3120
gtcttgtcag acagggaatc cctatccttc tacggcgaag gatacacctc tgatgaagag 3180
gtgttggagg tcttccgtaa tactttgaat aagaactcag aaatcttttc ctctataaag 3240
aaactagaga aattattcaa gaatttcgac gaatactcta gtgcaggcat cttcgtgaag 3300
aatggtcctg ctataagcac aatctctaaa gatatcttcg gggaatggaa tgtcatccga 3360
gataagtgga atgcggaata cgatgatatc cacttaaaga aaaaggcagt tgtaaccgaa 3420
aaatacgaag atgacaggcg taagtcattc aagaagatcg gatcattcag tctcgaacag 3480
ttacaagagt acgccgacgc tgacctttct gtcgtggaaa agctaaaaga gattatcata 3540
cagaaagttg atgaaatata taaggtctat ggttcatcag aaaagttgtt cgatgctgat 3600
ttcgtattag aaaagagttt aaagaaaaac gacgctgtag tggcaatcat gaaggattta 3660
ttggacagtg tcaagtcatt tgaaaattac atcaaggcat tcttcggtga aggtaaggaa 3720
acaaaccgcg atgagtcctt ctacggagat ttcgtgttgg cctatgacat cctattaaag 3780
gtggatcaca tctatgatgc tatccgtaat tatgtcacac agaaacccta ctctaaggac 3840
aagttcaagt tgtacttcca gaacccacaa tttatgggtg gttgggataa ggataaggaa 3900
actgactaca gagcaactat cttaaggtat ggttcaaagt actatctcgc tatcatggac 3960
aagaaatacg ctaagtgtct acaaaagatc gacaaagatg atgtcaatgg taattacgag 4020
aagatcaatt acaaattact accaggacca aataagatgc ttccgaaggt gttcttttca 4080
aagaagtgga tggcgtatta caaccctagt gaagacatcc agaagatata taagaatggg 4140
accttcaaaa agggagatat gttcaatctg aatgattgtc acaaattaat cgatttcttt 4200
aaggacagta tctccagata cccgaagtgg tcaaatgctt acgattttaa cttctcggaa 4260
acagagaagt acaaagacat cgctggtttc tacagagaag tggaggagca gggttacaag 4320
gtgtcattcg aaagtgcaag caagaaggag gtagataagc tggtagagga gggcaagttg 4380
tatatgtttc aaatctataa caaggatttc tcggataagt ctcacggaac accaaactta 4440
catactatgt actttaagct cctgttcgat gaaaacaatc atgggcaaat cagattatct 4500
ggtggtgctg aattgttcat gagacgtgca agtttaaaga aggaggaatt ggttgtcgct 4560
ccagcaaata gcccaatcgc gaataagaat ccagacaacc caaagaaaac gacgacgttg 4620
tcgtacgatg tttacaagga taagagattc tcagaagatc aatatgagct gcatatccct 4680
atcgcaatca ataagtgtcc taagaacatt ttcaagatta acactgaagt tagagtgcta 4740
ctcaagcatg atgataaccc ctacgttatc ggcatagata gaggcgagag gaatttgtta 4800
tacatcgtcg tggtggatgg caagggaaac atcgttgagc agtattctct taatgaaatt 4860
atcaataatt tcaatggaat tagaatcaaa actgattatc actccctgtt ggacaagaag 4920
gaaaaggaaa gattcgaagc tagacagaac tggacttcca ttgaaaacat taaagagcta 4980
aaggctggtt atatctccca agttgttcat aagatctgcg agttagtgga aaagtatgat 5040
gctgtaatcg cccttgaaga tcttaactct ggtttcaaga attcaagagt caaggttgaa 5100
aagcaagtgt accagaaatt tgagaagatg ctgatagaca aattgaatta catggtagat 5160
aagaagtcca atccatgtgc caccggggga gccttaaagg ggtatcagat cacaaacaag 5220
ttcgaatctt tcaagtcgat gagcacacag aatgggttta tattctacat tccggcctgg 5280
ttaacttcta agattgatcc cagcaccggc ttcgttaatc tgttgaagac taagtacacg 5340
tctatcgccg acagcaagaa gttcatctct agctttgata gaattatgta cgttccagaa 5400
gaagatctgt tcgaattcgc tcttgactac aagaacttct ctcggacaga tgcagattac 5460
atcaagaaat ggaagttgta tagttacggc aatagaatcc gtatcttccg taaccctaag 5520
aagaacaacg tcttcgattg ggaagaagtt tgcttaacat ccgcatacaa ggaactgttt 5580
aataagtacg gaatcaatta ccaacaaggt gatattcgag cactgttgtg tgaacaatct 5640
gacaaggctt tctattcgag tttcatggca ttaatgtcac tgatgttgca aatgcggaat 5700
agtatcactg gtagaactga tgtggatttc cttatcagtc ctgttaagaa ctctgatgga 5760
attttctatg actccaggaa ttacgaagcc caagaaaatg ctatccttcc taagaatgcg 5820
gacgctaacg gcgcttacaa tattgcaaga aaggtgttgt gggccatagg tcagttcaag 5880
aaggccgaag atgaaaagtt agataaggta aagattgcga tcagtaataa ggagtggttg 5940
gaatacgccc aaacatccgt taagcatggt agttcaggag gtagcaagag gactgctgat 6000
gggtctgaat tcgagccaaa gaagaagaga aaggttggct ccggttaa 6048
<210> 155
<211> 2015
<212> PRT
<213> 人工序列
<220>
<223> 合成的在N-末端处融合的REDRAW编辑器
<400> 155
Met Ala Thr Ser Arg Lys Arg Pro Ser Asp Leu Val His Val Phe Ser
1 5 10 15
Pro Pro Arg Lys Lys Ala Gly Ser Ala Thr Ser Arg Lys Arg Pro Ser
20 25 30
Asp Leu Val His Val Phe Ser Pro Pro Arg Lys Lys Ala Gly Ser Thr
35 40 45
Leu Asn Ile Glu Asp Glu His Arg Leu His Glu Thr Ser Lys Glu Pro
50 55 60
Asp Val Ser Leu Gly Ser Thr Trp Leu Ser Asp Phe Pro Gln Ala Trp
65 70 75 80
Ala Glu Thr Gly Gly Met Gly Leu Ala Val Arg Gln Ala Pro Leu Ile
85 90 95
Ile Pro Leu Lys Ala Thr Ser Thr Pro Val Ser Ile Lys Gln Tyr Pro
100 105 110
Met Ser Gln Glu Ala Arg Leu Gly Ile Lys Pro His Ile Gln Arg Leu
115 120 125
Leu Asp Gln Gly Ile Leu Val Pro Cys Gln Ser Pro Trp Asn Thr Pro
130 135 140
Leu Leu Pro Val Lys Lys Pro Gly Thr Asn Asp Tyr Arg Pro Val Gln
145 150 155 160
Asp Leu Arg Glu Val Asn Lys Arg Val Glu Asp Ile His Pro Thr Val
165 170 175
Pro Asn Pro Tyr Asn Leu Leu Ser Gly Leu Pro Pro Ser His Gln Trp
180 185 190
Tyr Thr Val Leu Asp Leu Lys Asp Ala Phe Phe Cys Leu Arg Leu His
195 200 205
Pro Thr Ser Gln Pro Leu Phe Ala Phe Glu Trp Arg Asp Pro Glu Met
210 215 220
Gly Ile Ser Gly Gln Leu Thr Trp Thr Arg Leu Pro Gln Gly Phe Lys
225 230 235 240
Asn Ser Pro Thr Leu Phe Asn Glu Ala Leu His Arg Asp Leu Ala Asp
245 250 255
Phe Arg Ile Gln His Pro Asp Leu Ile Leu Leu Gln Tyr Val Asp Asp
260 265 270
Leu Leu Leu Ala Ala Thr Ser Glu Leu Asp Cys Gln Gln Gly Thr Arg
275 280 285
Ala Leu Leu Gln Thr Leu Gly Asn Leu Gly Tyr Arg Ala Ser Ala Lys
290 295 300
Lys Ala Gln Ile Cys Gln Lys Gln Val Lys Tyr Leu Gly Tyr Leu Leu
305 310 315 320
Lys Glu Gly Gln Arg Trp Leu Thr Glu Ala Arg Lys Glu Thr Val Met
325 330 335
Gly Gln Pro Thr Pro Lys Thr Pro Arg Gln Leu Arg Glu Phe Leu Gly
340 345 350
Lys Ala Gly Phe Cys Arg Leu Phe Ile Pro Gly Phe Ala Glu Met Ala
355 360 365
Ala Pro Leu Tyr Pro Leu Thr Lys Pro Gly Thr Leu Phe Asn Trp Gly
370 375 380
Pro Asp Gln Gln Lys Ala Tyr Gln Glu Ile Lys Gln Ala Leu Leu Thr
385 390 395 400
Ala Pro Ala Leu Gly Leu Pro Asp Leu Thr Lys Pro Phe Glu Leu Phe
405 410 415
Val Asp Glu Lys Gln Gly Tyr Ala Lys Gly Val Leu Thr Gln Lys Leu
420 425 430
Gly Pro Trp Arg Arg Pro Val Ala Tyr Leu Ser Lys Lys Leu Asp Pro
435 440 445
Val Ala Ala Gly Trp Pro Pro Cys Leu Arg Met Val Ala Ala Ile Ala
450 455 460
Val Leu Thr Lys Asp Ala Gly Lys Leu Thr Met Gly Gln Pro Leu Val
465 470 475 480
Ile Leu Ala Pro His Ala Val Glu Ala Leu Val Lys Gln Pro Pro Asp
485 490 495
Arg Trp Leu Ser Asn Ala Arg Met Thr His Tyr Gln Ala Leu Leu Leu
500 505 510
Asp Thr Asp Arg Val Gln Phe Gly Pro Val Val Ala Leu Asn Pro Ala
515 520 525
Thr Leu Leu Pro Leu Pro Glu Glu Gly Leu Gln His Asn Cys Leu Asp
530 535 540
Ile Leu Ala Glu Ala His Gly Thr Arg Pro Asp Leu Thr Asp Gln Pro
545 550 555 560
Leu Pro Asp Ala Asp His Thr Trp Tyr Thr Asp Gly Ser Ser Leu Leu
565 570 575
Gln Glu Gly Gln Arg Lys Ala Gly Ala Ala Val Thr Thr Glu Thr Glu
580 585 590
Val Ile Trp Ala Lys Ala Leu Pro Ala Gly Thr Ser Ala Gln Arg Ala
595 600 605
Glu Leu Ile Ala Leu Thr Gln Ala Leu Lys Met Ala Glu Gly Lys Lys
610 615 620
Leu Asn Val Tyr Thr Asp Ser Arg Tyr Ala Phe Ala Thr Ala His Ile
625 630 635 640
His Gly Glu Ile Tyr Arg Arg Arg Gly Trp Leu Thr Ser Glu Gly Lys
645 650 655
Glu Ile Lys Asn Lys Asp Glu Ile Leu Ala Leu Leu Lys Ala Leu Phe
660 665 670
Leu Pro Lys Arg Leu Ser Ile Ile His Cys Pro Gly His Gln Lys Gly
675 680 685
His Ser Ala Glu Ala Arg Gly Asn Arg Met Ala Asp Gln Ala Ala Arg
690 695 700
Lys Ala Ala Ile Thr Glu Thr Pro Asp Thr Ser Thr Leu Leu Ile Glu
705 710 715 720
Asn Ser Ser Pro Asn Ser Arg Leu Ile Asn Ser Gly Gly Ser Ser Gly
725 730 735
Gly Ser Ser Gly Ser Glu Thr Pro Gly Thr Ser Glu Ser Ala Thr Pro
740 745 750
Glu Ser Ser Gly Gly Ser Ser Gly Gly Ser Ser Lys Leu Glu Lys Phe
755 760 765
Thr Asn Cys Tyr Ser Leu Ser Lys Thr Leu Arg Phe Lys Ala Ile Pro
770 775 780
Val Gly Lys Thr Gln Glu Asn Ile Asp Asn Lys Arg Leu Leu Val Glu
785 790 795 800
Asp Glu Lys Arg Ala Glu Asp Tyr Lys Gly Val Lys Lys Leu Leu Asp
805 810 815
Arg Tyr Tyr Leu Ser Phe Ile Asn Asp Val Leu His Ser Ile Lys Leu
820 825 830
Lys Asn Leu Asn Asn Tyr Ile Ser Leu Phe Arg Lys Lys Thr Arg Thr
835 840 845
Glu Lys Glu Asn Lys Glu Leu Glu Asn Leu Glu Ile Asn Leu Arg Lys
850 855 860
Glu Ile Ala Lys Ala Phe Lys Gly Asn Glu Gly Tyr Lys Ser Leu Phe
865 870 875 880
Lys Lys Asp Ile Ile Glu Thr Ile Leu Pro Glu Phe Leu Asp Asp Lys
885 890 895
Asp Glu Ile Ala Leu Val Asn Ser Phe Asn Gly Phe Thr Thr Ala Phe
900 905 910
Thr Gly Phe Phe Asp Asn Arg Glu Asn Met Phe Ser Glu Glu Ala Lys
915 920 925
Ser Thr Ser Ile Ala Phe Arg Cys Ile Asn Glu Asn Leu Thr Arg Tyr
930 935 940
Ile Ser Asn Met Asp Ile Phe Glu Lys Val Asp Ala Ile Phe Asp Lys
945 950 955 960
His Glu Val Gln Glu Ile Lys Glu Lys Ile Leu Asn Ser Asp Tyr Asp
965 970 975
Val Glu Asp Phe Phe Glu Gly Glu Phe Phe Asn Phe Val Leu Thr Gln
980 985 990
Glu Gly Ile Asp Val Tyr Asn Ala Ile Ile Gly Gly Phe Val Thr Glu
995 1000 1005
Ser Gly Glu Lys Ile Lys Gly Leu Asn Glu Tyr Ile Asn Leu Tyr
1010 1015 1020
Asn Gln Lys Thr Lys Gln Lys Leu Pro Lys Phe Lys Pro Leu Tyr
1025 1030 1035
Lys Gln Val Leu Ser Asp Arg Glu Ser Leu Ser Phe Tyr Gly Glu
1040 1045 1050
Gly Tyr Thr Ser Asp Glu Glu Val Leu Glu Val Phe Arg Asn Thr
1055 1060 1065
Leu Asn Lys Asn Ser Glu Ile Phe Ser Ser Ile Lys Lys Leu Glu
1070 1075 1080
Lys Leu Phe Lys Asn Phe Asp Glu Tyr Ser Ser Ala Gly Ile Phe
1085 1090 1095
Val Lys Asn Gly Pro Ala Ile Ser Thr Ile Ser Lys Asp Ile Phe
1100 1105 1110
Gly Glu Trp Asn Val Ile Arg Asp Lys Trp Asn Ala Glu Tyr Asp
1115 1120 1125
Asp Ile His Leu Lys Lys Lys Ala Val Val Thr Glu Lys Tyr Glu
1130 1135 1140
Asp Asp Arg Arg Lys Ser Phe Lys Lys Ile Gly Ser Phe Ser Leu
1145 1150 1155
Glu Gln Leu Gln Glu Tyr Ala Asp Ala Asp Leu Ser Val Val Glu
1160 1165 1170
Lys Leu Lys Glu Ile Ile Ile Gln Lys Val Asp Glu Ile Tyr Lys
1175 1180 1185
Val Tyr Gly Ser Ser Glu Lys Leu Phe Asp Ala Asp Phe Val Leu
1190 1195 1200
Glu Lys Ser Leu Lys Lys Asn Asp Ala Val Val Ala Ile Met Lys
1205 1210 1215
Asp Leu Leu Asp Ser Val Lys Ser Phe Glu Asn Tyr Ile Lys Ala
1220 1225 1230
Phe Phe Gly Glu Gly Lys Glu Thr Asn Arg Asp Glu Ser Phe Tyr
1235 1240 1245
Gly Asp Phe Val Leu Ala Tyr Asp Ile Leu Leu Lys Val Asp His
1250 1255 1260
Ile Tyr Asp Ala Ile Arg Asn Tyr Val Thr Gln Lys Pro Tyr Ser
1265 1270 1275
Lys Asp Lys Phe Lys Leu Tyr Phe Gln Asn Pro Gln Phe Met Gly
1280 1285 1290
Gly Trp Asp Lys Asp Lys Glu Thr Asp Tyr Arg Ala Thr Ile Leu
1295 1300 1305
Arg Tyr Gly Ser Lys Tyr Tyr Leu Ala Ile Met Asp Lys Lys Tyr
1310 1315 1320
Ala Lys Cys Leu Gln Lys Ile Asp Lys Asp Asp Val Asn Gly Asn
1325 1330 1335
Tyr Glu Lys Ile Asn Tyr Lys Leu Leu Pro Gly Pro Asn Lys Met
1340 1345 1350
Leu Pro Lys Val Phe Phe Ser Lys Lys Trp Met Ala Tyr Tyr Asn
1355 1360 1365
Pro Ser Glu Asp Ile Gln Lys Ile Tyr Lys Asn Gly Thr Phe Lys
1370 1375 1380
Lys Gly Asp Met Phe Asn Leu Asn Asp Cys His Lys Leu Ile Asp
1385 1390 1395
Phe Phe Lys Asp Ser Ile Ser Arg Tyr Pro Lys Trp Ser Asn Ala
1400 1405 1410
Tyr Asp Phe Asn Phe Ser Glu Thr Glu Lys Tyr Lys Asp Ile Ala
1415 1420 1425
Gly Phe Tyr Arg Glu Val Glu Glu Gln Gly Tyr Lys Val Ser Phe
1430 1435 1440
Glu Ser Ala Ser Lys Lys Glu Val Asp Lys Leu Val Glu Glu Gly
1445 1450 1455
Lys Leu Tyr Met Phe Gln Ile Tyr Asn Lys Asp Phe Ser Asp Lys
1460 1465 1470
Ser His Gly Thr Pro Asn Leu His Thr Met Tyr Phe Lys Leu Leu
1475 1480 1485
Phe Asp Glu Asn Asn His Gly Gln Ile Arg Leu Ser Gly Gly Ala
1490 1495 1500
Glu Leu Phe Met Arg Arg Ala Ser Leu Lys Lys Glu Glu Leu Val
1505 1510 1515
Val Ala Pro Ala Asn Ser Pro Ile Ala Asn Lys Asn Pro Asp Asn
1520 1525 1530
Pro Lys Lys Thr Thr Thr Leu Ser Tyr Asp Val Tyr Lys Asp Lys
1535 1540 1545
Arg Phe Ser Glu Asp Gln Tyr Glu Leu His Ile Pro Ile Ala Ile
1550 1555 1560
Asn Lys Cys Pro Lys Asn Ile Phe Lys Ile Asn Thr Glu Val Arg
1565 1570 1575
Val Leu Leu Lys His Asp Asp Asn Pro Tyr Val Ile Gly Ile Asp
1580 1585 1590
Arg Gly Glu Arg Asn Leu Leu Tyr Ile Val Val Val Asp Gly Lys
1595 1600 1605
Gly Asn Ile Val Glu Gln Tyr Ser Leu Asn Glu Ile Ile Asn Asn
1610 1615 1620
Phe Asn Gly Ile Arg Ile Lys Thr Asp Tyr His Ser Leu Leu Asp
1625 1630 1635
Lys Lys Glu Lys Glu Arg Phe Glu Ala Arg Gln Asn Trp Thr Ser
1640 1645 1650
Ile Glu Asn Ile Lys Glu Leu Lys Ala Gly Tyr Ile Ser Gln Val
1655 1660 1665
Val His Lys Ile Cys Glu Leu Val Glu Lys Tyr Asp Ala Val Ile
1670 1675 1680
Ala Leu Glu Asp Leu Asn Ser Gly Phe Lys Asn Ser Arg Val Lys
1685 1690 1695
Val Glu Lys Gln Val Tyr Gln Lys Phe Glu Lys Met Leu Ile Asp
1700 1705 1710
Lys Leu Asn Tyr Met Val Asp Lys Lys Ser Asn Pro Cys Ala Thr
1715 1720 1725
Gly Gly Ala Leu Lys Gly Tyr Gln Ile Thr Asn Lys Phe Glu Ser
1730 1735 1740
Phe Lys Ser Met Ser Thr Gln Asn Gly Phe Ile Phe Tyr Ile Pro
1745 1750 1755
Ala Trp Leu Thr Ser Lys Ile Asp Pro Ser Thr Gly Phe Val Asn
1760 1765 1770
Leu Leu Lys Thr Lys Tyr Thr Ser Ile Ala Asp Ser Lys Lys Phe
1775 1780 1785
Ile Ser Ser Phe Asp Arg Ile Met Tyr Val Pro Glu Glu Asp Leu
1790 1795 1800
Phe Glu Phe Ala Leu Asp Tyr Lys Asn Phe Ser Arg Thr Asp Ala
1805 1810 1815
Asp Tyr Ile Lys Lys Trp Lys Leu Tyr Ser Tyr Gly Asn Arg Ile
1820 1825 1830
Arg Ile Phe Arg Asn Pro Lys Lys Asn Asn Val Phe Asp Trp Glu
1835 1840 1845
Glu Val Cys Leu Thr Ser Ala Tyr Lys Glu Leu Phe Asn Lys Tyr
1850 1855 1860
Gly Ile Asn Tyr Gln Gln Gly Asp Ile Arg Ala Leu Leu Cys Glu
1865 1870 1875
Gln Ser Asp Lys Ala Phe Tyr Ser Ser Phe Met Ala Leu Met Ser
1880 1885 1890
Leu Met Leu Gln Met Arg Asn Ser Ile Thr Gly Arg Thr Asp Val
1895 1900 1905
Asp Phe Leu Ile Ser Pro Val Lys Asn Ser Asp Gly Ile Phe Tyr
1910 1915 1920
Asp Ser Arg Asn Tyr Glu Ala Gln Glu Asn Ala Ile Leu Pro Lys
1925 1930 1935
Asn Ala Asp Ala Asn Gly Ala Tyr Asn Ile Ala Arg Lys Val Leu
1940 1945 1950
Trp Ala Ile Gly Gln Phe Lys Lys Ala Glu Asp Glu Lys Leu Asp
1955 1960 1965
Lys Val Lys Ile Ala Ile Ser Asn Lys Glu Trp Leu Glu Tyr Ala
1970 1975 1980
Gln Thr Ser Val Lys His Gly Ser Ser Gly Gly Ser Lys Arg Thr
1985 1990 1995
Ala Asp Gly Ser Glu Phe Glu Pro Lys Lys Lys Arg Lys Val Gly
2000 2005 2010
Ser Gly
2015
<210> 156
<211> 6048
<212> DNA
<213> 人工序列
<220>
<223> 合成的在C-末端处融合的REDRAW编辑器
<400> 156
atggcaactt caagaaagag gcctagtgac ttggttcacg tattctctcc tccacgaaag 60
aaggctggtt cggctacatc aagaaagcgt cccagcgatt tagtgcatgt cttctctccg 120
ccaagaaaga aggccggatc cagcaaactc gaaaagttca ctaattgtta ctctctatca 180
aagaccttaa gatttaaggc tatacctgta gggaagactc aagaaaacat cgataataag 240
aggttgttag ttgaagatga aaagagggct gaagactaca agggtgtaaa gaaattgttg 300
gaccgatact accttagctt catcaacgat gtcttgcatt caataaagtt gaaaaattta 360
aataactaca tctcgctatt ccgtaagaaa acgaggaccg aaaaggagaa caaagaacta 420
gaaaacttgg aaatcaacct gagaaaggag atagccaaag ccttcaaggg taatgagggt 480
tacaagtctc tttttaagaa agatattatc gaaactatac taccagaatt cttagatgac 540
aaagacgaga tcgccctggt aaactctttt aatggtttca caactgcgtt cacaggtttc 600
tttgacaatc gcgagaatat gttctctgaa gaggctaaga gtacatcaat cgcattcagg 660
tgcatcaacg aaaacttgac gaggtacatt tcgaacatgg acatatttga aaaggttgac 720
gcaatcttcg acaagcatga agtacaagaa atcaaggaaa agatcttaaa ttcggattac 780
gatgtggagg atttcttcga aggtgaattc tttaacttcg tcttgaccca agagggtatc 840
gatgtctaca atgcaatcat aggaggtttc gttaccgaat ccggcgagaa gatcaaaggt 900
ctaaacgaat atatcaatct gtataatcaa aagaccaagc agaaactacc caaattcaag 960
cctctctaca agcaagtctt gtcagacagg gaatccctat ccttctacgg cgaaggatac 1020
acctctgatg aagaggtgtt ggaggtcttc cgtaatactt tgaataagaa ctcagaaatc 1080
ttttcctcta taaagaaact agagaaatta ttcaagaatt tcgacgaata ctctagtgca 1140
ggcatcttcg tgaagaatgg tcctgctata agcacaatct ctaaagatat cttcggggaa 1200
tggaatgtca tccgagataa gtggaatgcg gaatacgatg atatccactt aaagaaaaag 1260
gcagttgtaa ccgaaaaata cgaagatgac aggcgtaagt cattcaagaa gatcggatca 1320
ttcagtctcg aacagttaca agagtacgcc gacgctgacc tttctgtcgt ggaaaagcta 1380
aaagagatta tcatacagaa agttgatgaa atatataagg tctatggttc atcagaaaag 1440
ttgttcgatg ctgatttcgt attagaaaag agtttaaaga aaaacgacgc tgtagtggca 1500
atcatgaagg atttattgga cagtgtcaag tcatttgaaa attacatcaa ggcattcttc 1560
ggtgaaggta aggaaacaaa ccgcgatgag tccttctacg gagatttcgt gttggcctat 1620
gacatcctat taaaggtgga tcacatctat gatgctatcc gtaattatgt cacacagaaa 1680
ccctactcta aggacaagtt caagttgtac ttccagaacc cacaatttat gggtggttgg 1740
gataaggata aggaaactga ctacagagca actatcttaa ggtatggttc aaagtactat 1800
ctcgctatca tggacaagaa atacgctaag tgtctacaaa agatcgacaa agatgatgtc 1860
aatggtaatt acgagaagat caattacaaa ttactaccag gaccaaataa gatgcttccg 1920
aaggtgttct tttcaaagaa gtggatggcg tattacaacc ctagtgaaga catccagaag 1980
atatataaga atgggacctt caaaaaggga gatatgttca atctgaatga ttgtcacaaa 2040
ttaatcgatt tctttaagga cagtatctcc agatacccga agtggtcaaa tgcttacgat 2100
tttaacttct cggaaacaga gaagtacaaa gacatcgctg gtttctacag agaagtggag 2160
gagcagggtt acaaggtgtc attcgaaagt gcaagcaaga aggaggtaga taagctggta 2220
gaggagggca agttgtatat gtttcaaatc tataacaagg atttctcgga taagtctcac 2280
ggaacaccaa acttacatac tatgtacttt aagctcctgt tcgatgaaaa caatcatggg 2340
caaatcagat tatctggtgg tgctgaattg ttcatgagac gtgcaagttt aaagaaggag 2400
gaattggttg tcgctccagc aaatagccca atcgcgaata agaatccaga caacccaaag 2460
aaaacgacga cgttgtcgta cgatgtttac aaggataaga gattctcaga agatcaatat 2520
gagctgcata tccctatcgc aatcaataag tgtcctaaga acattttcaa gattaacact 2580
gaagttagag tgctactcaa gcatgatgat aacccctacg ttatcggcat agatagaggc 2640
gagaggaatt tgttatacat cgtcgtggtg gatggcaagg gaaacatcgt tgagcagtat 2700
tctcttaatg aaattatcaa taatttcaat ggaattagaa tcaaaactga ttatcactcc 2760
ctgttggaca agaaggaaaa ggaaagattc gaagctagac agaactggac ttccattgaa 2820
aacattaaag agctaaaggc tggttatatc tcccaagttg ttcataagat ctgcgagtta 2880
gtggaaaagt atgatgctgt aatcgccctt gaagatctta actctggttt caagaattca 2940
agagtcaagg ttgaaaagca agtgtaccag aaatttgaga agatgctgat agacaaattg 3000
aattacatgg tagataagaa gtccaatcca tgtgccaccg ggggagcctt aaaggggtat 3060
cagatcacaa acaagttcga atctttcaag tcgatgagca cacagaatgg gtttatattc 3120
tacattccgg cctggttaac ttctaagatt gatcccagca ccggcttcgt taatctgttg 3180
aagactaagt acacgtctat cgccgacagc aagaagttca tctctagctt tgatagaatt 3240
atgtacgttc cagaagaaga tctgttcgaa ttcgctcttg actacaagaa cttctctcgg 3300
acagatgcag attacatcaa gaaatggaag ttgtatagtt acggcaatag aatccgtatc 3360
ttccgtaacc ctaagaagaa caacgtcttc gattgggaag aagtttgctt aacatccgca 3420
tacaaggaac tgtttaataa gtacggaatc aattaccaac aaggtgatat tcgagcactg 3480
ttgtgtgaac aatctgacaa ggctttctat tcgagtttca tggcattaat gtcactgatg 3540
ttgcaaatgc ggaatagtat cactggtaga actgatgtgg atttccttat cagtcctgtt 3600
aagaactctg atggaatttt ctatgactcc aggaattacg aagcccaaga aaatgctatc 3660
cttcctaaga atgcggacgc taacggcgct tacaatattg caagaaaggt gttgtgggcc 3720
ataggtcagt tcaagaaggc cgaagatgaa aagttagata aggtaaagat tgcgatcagt 3780
aataaggagt ggttggaata cgcccaaaca tccgttaagc attcaggtgg atcgtccggt 3840
ggcagttctg gttcagaaac tcctggtaca tctgagagcg ctaccccaga atccagtgga 3900
gggagctcag ggggttctac tctcaatatc gaagacgaac atcgtttaca tgaaacatct 3960
aaagaaccgg acgtttcttt aggttccaca tggttaagcg acttccctca ggcctgggca 4020
gaaacaggtg gtatgggact tgccgttcga caagcgcctt tgatcatacc cctaaaggcg 4080
acaagtaccc cagtatccat taaacagtat cctatgtccc aggaggcacg cttggggata 4140
aagccccata tccaaagact attagatcag gggatacttg tcccatgcca aagcccttgg 4200
aacactccgt tactgcccgt gaaaaaacca ggtacaaacg attatagacc tgtgcaagac 4260
ctgcgcgagg tgaacaagag ggtagaggat atacatccaa cggttccaaa tccttataat 4320
ttactatctg gtcttccacc tagtcaccag tggtataccg tcttggactt gaaggatgcg 4380
tttttctgct tgcgtttaca tccgacgtct caacctttat tcgcattcga gtggcgagat 4440
cctgagatgg gtatctctgg tcagttaact tggacaagat tgcctcaagg ttttaagaat 4500
agtccaactt tatttaatga agccttgcat agagatttgg ctgattttag gattcaacat 4560
cctgacctaa ttttgctaca atacgttgat gatctattgt tggctgccac ctcggaactt 4620
gattgtcaac agggtacgag ggctctgctt caaaccttag gaaacttggg ataccgagct 4680
tctgctaaaa aggcgcaaat ttgtcaaaag caggtaaaat acttgggtta cctattgaaa 4740
gaagggcaaa ggtggctaac tgaggcccgt aaagaaaccg tgatgggtca accaaccccc 4800
aaaacaccaa gacaactcag ggaattttta ggcaaagcgg gtttctgtag attgtttatt 4860
cctggctttg cagaaatggc cgcaccactg tatccactga caaaaccagg caccttgttc 4920
aactggggcc cagatcagca gaaggcttat caagaaatta aacaggcttt gctaacagca 4980
ccagcactag gtctaccaga tttaactaaa cccttcgaat tgtttgttga tgaaaaacaa 5040
ggatatgcta aaggtgtcct gactcaaaaa ttgggaccct ggagaagacc ggttgcatat 5100
ttaagcaaaa agttggaccc agtagccgct gggtggccac cgtgtttaag gatggttgct 5160
gctatcgctg tcttaacgaa ggatgccggc aaattaacga tgggacagcc actggttata 5220
ctagctcccc atgctgttga agccttggtg aaacaacctc ccgaccgttg gttatcaaat 5280
gcaagaatga cacactatca agccttgttg ctcgacacag acagagttca atttggacct 5340
gtagtcgcac taaatccggc gactctttta ccactgccag aagagggtct tcaacataac 5400
tgcctcgata ttttggccga agctcatggg accagaccgg acttaacaga tcaaccttta 5460
ccagatgcag atcacacttg gtacactgat ggtagttcac tccttcaaga gggtcaacgt 5520
aaagctggcg cagcagttac cactgaaacg gaagtaattt gggcaaaagc attacctgct 5580
ggtacttcag cccaaagagc tgagttgatt gccttgactc aagctcttaa aatggctgaa 5640
ggcaagaagc tgaatgtcta tactgactca agatacgctt ttgctactgc tcatattcac 5700
ggtgaaatct acagaagaag aggatggctt acgagtgaag gaaaagaaat taagaataag 5760
gatgagatat tagcacttct caaagcattg tttctgccaa agagactttc aattattcac 5820
tgtccaggac atcaaaaggg ccactccgcc gaagctaggg gtaatagaat ggcagatcag 5880
gcagcgagga aggctgccat cacagagact cctgatacat ctaccttatt aatagaaaac 5940
tcgtcgccaa attcacggtt aataaacggt agttcaggag gtagcaagag gactgctgat 6000
gggtctgaat tcgagccaaa gaagaagaga aaggttggct ccggttaa 6048
<210> 157
<211> 2015
<212> PRT
<213> 人工序列
<220>
<223> 合成的在C-末端处融合的REDRAW编辑器
<400> 157
Met Ala Thr Ser Arg Lys Arg Pro Ser Asp Leu Val His Val Phe Ser
1 5 10 15
Pro Pro Arg Lys Lys Ala Gly Ser Ala Thr Ser Arg Lys Arg Pro Ser
20 25 30
Asp Leu Val His Val Phe Ser Pro Pro Arg Lys Lys Ala Gly Ser Ser
35 40 45
Lys Leu Glu Lys Phe Thr Asn Cys Tyr Ser Leu Ser Lys Thr Leu Arg
50 55 60
Phe Lys Ala Ile Pro Val Gly Lys Thr Gln Glu Asn Ile Asp Asn Lys
65 70 75 80
Arg Leu Leu Val Glu Asp Glu Lys Arg Ala Glu Asp Tyr Lys Gly Val
85 90 95
Lys Lys Leu Leu Asp Arg Tyr Tyr Leu Ser Phe Ile Asn Asp Val Leu
100 105 110
His Ser Ile Lys Leu Lys Asn Leu Asn Asn Tyr Ile Ser Leu Phe Arg
115 120 125
Lys Lys Thr Arg Thr Glu Lys Glu Asn Lys Glu Leu Glu Asn Leu Glu
130 135 140
Ile Asn Leu Arg Lys Glu Ile Ala Lys Ala Phe Lys Gly Asn Glu Gly
145 150 155 160
Tyr Lys Ser Leu Phe Lys Lys Asp Ile Ile Glu Thr Ile Leu Pro Glu
165 170 175
Phe Leu Asp Asp Lys Asp Glu Ile Ala Leu Val Asn Ser Phe Asn Gly
180 185 190
Phe Thr Thr Ala Phe Thr Gly Phe Phe Asp Asn Arg Glu Asn Met Phe
195 200 205
Ser Glu Glu Ala Lys Ser Thr Ser Ile Ala Phe Arg Cys Ile Asn Glu
210 215 220
Asn Leu Thr Arg Tyr Ile Ser Asn Met Asp Ile Phe Glu Lys Val Asp
225 230 235 240
Ala Ile Phe Asp Lys His Glu Val Gln Glu Ile Lys Glu Lys Ile Leu
245 250 255
Asn Ser Asp Tyr Asp Val Glu Asp Phe Phe Glu Gly Glu Phe Phe Asn
260 265 270
Phe Val Leu Thr Gln Glu Gly Ile Asp Val Tyr Asn Ala Ile Ile Gly
275 280 285
Gly Phe Val Thr Glu Ser Gly Glu Lys Ile Lys Gly Leu Asn Glu Tyr
290 295 300
Ile Asn Leu Tyr Asn Gln Lys Thr Lys Gln Lys Leu Pro Lys Phe Lys
305 310 315 320
Pro Leu Tyr Lys Gln Val Leu Ser Asp Arg Glu Ser Leu Ser Phe Tyr
325 330 335
Gly Glu Gly Tyr Thr Ser Asp Glu Glu Val Leu Glu Val Phe Arg Asn
340 345 350
Thr Leu Asn Lys Asn Ser Glu Ile Phe Ser Ser Ile Lys Lys Leu Glu
355 360 365
Lys Leu Phe Lys Asn Phe Asp Glu Tyr Ser Ser Ala Gly Ile Phe Val
370 375 380
Lys Asn Gly Pro Ala Ile Ser Thr Ile Ser Lys Asp Ile Phe Gly Glu
385 390 395 400
Trp Asn Val Ile Arg Asp Lys Trp Asn Ala Glu Tyr Asp Asp Ile His
405 410 415
Leu Lys Lys Lys Ala Val Val Thr Glu Lys Tyr Glu Asp Asp Arg Arg
420 425 430
Lys Ser Phe Lys Lys Ile Gly Ser Phe Ser Leu Glu Gln Leu Gln Glu
435 440 445
Tyr Ala Asp Ala Asp Leu Ser Val Val Glu Lys Leu Lys Glu Ile Ile
450 455 460
Ile Gln Lys Val Asp Glu Ile Tyr Lys Val Tyr Gly Ser Ser Glu Lys
465 470 475 480
Leu Phe Asp Ala Asp Phe Val Leu Glu Lys Ser Leu Lys Lys Asn Asp
485 490 495
Ala Val Val Ala Ile Met Lys Asp Leu Leu Asp Ser Val Lys Ser Phe
500 505 510
Glu Asn Tyr Ile Lys Ala Phe Phe Gly Glu Gly Lys Glu Thr Asn Arg
515 520 525
Asp Glu Ser Phe Tyr Gly Asp Phe Val Leu Ala Tyr Asp Ile Leu Leu
530 535 540
Lys Val Asp His Ile Tyr Asp Ala Ile Arg Asn Tyr Val Thr Gln Lys
545 550 555 560
Pro Tyr Ser Lys Asp Lys Phe Lys Leu Tyr Phe Gln Asn Pro Gln Phe
565 570 575
Met Gly Gly Trp Asp Lys Asp Lys Glu Thr Asp Tyr Arg Ala Thr Ile
580 585 590
Leu Arg Tyr Gly Ser Lys Tyr Tyr Leu Ala Ile Met Asp Lys Lys Tyr
595 600 605
Ala Lys Cys Leu Gln Lys Ile Asp Lys Asp Asp Val Asn Gly Asn Tyr
610 615 620
Glu Lys Ile Asn Tyr Lys Leu Leu Pro Gly Pro Asn Lys Met Leu Pro
625 630 635 640
Lys Val Phe Phe Ser Lys Lys Trp Met Ala Tyr Tyr Asn Pro Ser Glu
645 650 655
Asp Ile Gln Lys Ile Tyr Lys Asn Gly Thr Phe Lys Lys Gly Asp Met
660 665 670
Phe Asn Leu Asn Asp Cys His Lys Leu Ile Asp Phe Phe Lys Asp Ser
675 680 685
Ile Ser Arg Tyr Pro Lys Trp Ser Asn Ala Tyr Asp Phe Asn Phe Ser
690 695 700
Glu Thr Glu Lys Tyr Lys Asp Ile Ala Gly Phe Tyr Arg Glu Val Glu
705 710 715 720
Glu Gln Gly Tyr Lys Val Ser Phe Glu Ser Ala Ser Lys Lys Glu Val
725 730 735
Asp Lys Leu Val Glu Glu Gly Lys Leu Tyr Met Phe Gln Ile Tyr Asn
740 745 750
Lys Asp Phe Ser Asp Lys Ser His Gly Thr Pro Asn Leu His Thr Met
755 760 765
Tyr Phe Lys Leu Leu Phe Asp Glu Asn Asn His Gly Gln Ile Arg Leu
770 775 780
Ser Gly Gly Ala Glu Leu Phe Met Arg Arg Ala Ser Leu Lys Lys Glu
785 790 795 800
Glu Leu Val Val Ala Pro Ala Asn Ser Pro Ile Ala Asn Lys Asn Pro
805 810 815
Asp Asn Pro Lys Lys Thr Thr Thr Leu Ser Tyr Asp Val Tyr Lys Asp
820 825 830
Lys Arg Phe Ser Glu Asp Gln Tyr Glu Leu His Ile Pro Ile Ala Ile
835 840 845
Asn Lys Cys Pro Lys Asn Ile Phe Lys Ile Asn Thr Glu Val Arg Val
850 855 860
Leu Leu Lys His Asp Asp Asn Pro Tyr Val Ile Gly Ile Asp Arg Gly
865 870 875 880
Glu Arg Asn Leu Leu Tyr Ile Val Val Val Asp Gly Lys Gly Asn Ile
885 890 895
Val Glu Gln Tyr Ser Leu Asn Glu Ile Ile Asn Asn Phe Asn Gly Ile
900 905 910
Arg Ile Lys Thr Asp Tyr His Ser Leu Leu Asp Lys Lys Glu Lys Glu
915 920 925
Arg Phe Glu Ala Arg Gln Asn Trp Thr Ser Ile Glu Asn Ile Lys Glu
930 935 940
Leu Lys Ala Gly Tyr Ile Ser Gln Val Val His Lys Ile Cys Glu Leu
945 950 955 960
Val Glu Lys Tyr Asp Ala Val Ile Ala Leu Glu Asp Leu Asn Ser Gly
965 970 975
Phe Lys Asn Ser Arg Val Lys Val Glu Lys Gln Val Tyr Gln Lys Phe
980 985 990
Glu Lys Met Leu Ile Asp Lys Leu Asn Tyr Met Val Asp Lys Lys Ser
995 1000 1005
Asn Pro Cys Ala Thr Gly Gly Ala Leu Lys Gly Tyr Gln Ile Thr
1010 1015 1020
Asn Lys Phe Glu Ser Phe Lys Ser Met Ser Thr Gln Asn Gly Phe
1025 1030 1035
Ile Phe Tyr Ile Pro Ala Trp Leu Thr Ser Lys Ile Asp Pro Ser
1040 1045 1050
Thr Gly Phe Val Asn Leu Leu Lys Thr Lys Tyr Thr Ser Ile Ala
1055 1060 1065
Asp Ser Lys Lys Phe Ile Ser Ser Phe Asp Arg Ile Met Tyr Val
1070 1075 1080
Pro Glu Glu Asp Leu Phe Glu Phe Ala Leu Asp Tyr Lys Asn Phe
1085 1090 1095
Ser Arg Thr Asp Ala Asp Tyr Ile Lys Lys Trp Lys Leu Tyr Ser
1100 1105 1110
Tyr Gly Asn Arg Ile Arg Ile Phe Arg Asn Pro Lys Lys Asn Asn
1115 1120 1125
Val Phe Asp Trp Glu Glu Val Cys Leu Thr Ser Ala Tyr Lys Glu
1130 1135 1140
Leu Phe Asn Lys Tyr Gly Ile Asn Tyr Gln Gln Gly Asp Ile Arg
1145 1150 1155
Ala Leu Leu Cys Glu Gln Ser Asp Lys Ala Phe Tyr Ser Ser Phe
1160 1165 1170
Met Ala Leu Met Ser Leu Met Leu Gln Met Arg Asn Ser Ile Thr
1175 1180 1185
Gly Arg Thr Asp Val Asp Phe Leu Ile Ser Pro Val Lys Asn Ser
1190 1195 1200
Asp Gly Ile Phe Tyr Asp Ser Arg Asn Tyr Glu Ala Gln Glu Asn
1205 1210 1215
Ala Ile Leu Pro Lys Asn Ala Asp Ala Asn Gly Ala Tyr Asn Ile
1220 1225 1230
Ala Arg Lys Val Leu Trp Ala Ile Gly Gln Phe Lys Lys Ala Glu
1235 1240 1245
Asp Glu Lys Leu Asp Lys Val Lys Ile Ala Ile Ser Asn Lys Glu
1250 1255 1260
Trp Leu Glu Tyr Ala Gln Thr Ser Val Lys His Ser Gly Gly Ser
1265 1270 1275
Ser Gly Gly Ser Ser Gly Ser Glu Thr Pro Gly Thr Ser Glu Ser
1280 1285 1290
Ala Thr Pro Glu Ser Ser Gly Gly Ser Ser Gly Gly Ser Thr Leu
1295 1300 1305
Asn Ile Glu Asp Glu His Arg Leu His Glu Thr Ser Lys Glu Pro
1310 1315 1320
Asp Val Ser Leu Gly Ser Thr Trp Leu Ser Asp Phe Pro Gln Ala
1325 1330 1335
Trp Ala Glu Thr Gly Gly Met Gly Leu Ala Val Arg Gln Ala Pro
1340 1345 1350
Leu Ile Ile Pro Leu Lys Ala Thr Ser Thr Pro Val Ser Ile Lys
1355 1360 1365
Gln Tyr Pro Met Ser Gln Glu Ala Arg Leu Gly Ile Lys Pro His
1370 1375 1380
Ile Gln Arg Leu Leu Asp Gln Gly Ile Leu Val Pro Cys Gln Ser
1385 1390 1395
Pro Trp Asn Thr Pro Leu Leu Pro Val Lys Lys Pro Gly Thr Asn
1400 1405 1410
Asp Tyr Arg Pro Val Gln Asp Leu Arg Glu Val Asn Lys Arg Val
1415 1420 1425
Glu Asp Ile His Pro Thr Val Pro Asn Pro Tyr Asn Leu Leu Ser
1430 1435 1440
Gly Leu Pro Pro Ser His Gln Trp Tyr Thr Val Leu Asp Leu Lys
1445 1450 1455
Asp Ala Phe Phe Cys Leu Arg Leu His Pro Thr Ser Gln Pro Leu
1460 1465 1470
Phe Ala Phe Glu Trp Arg Asp Pro Glu Met Gly Ile Ser Gly Gln
1475 1480 1485
Leu Thr Trp Thr Arg Leu Pro Gln Gly Phe Lys Asn Ser Pro Thr
1490 1495 1500
Leu Phe Asn Glu Ala Leu His Arg Asp Leu Ala Asp Phe Arg Ile
1505 1510 1515
Gln His Pro Asp Leu Ile Leu Leu Gln Tyr Val Asp Asp Leu Leu
1520 1525 1530
Leu Ala Ala Thr Ser Glu Leu Asp Cys Gln Gln Gly Thr Arg Ala
1535 1540 1545
Leu Leu Gln Thr Leu Gly Asn Leu Gly Tyr Arg Ala Ser Ala Lys
1550 1555 1560
Lys Ala Gln Ile Cys Gln Lys Gln Val Lys Tyr Leu Gly Tyr Leu
1565 1570 1575
Leu Lys Glu Gly Gln Arg Trp Leu Thr Glu Ala Arg Lys Glu Thr
1580 1585 1590
Val Met Gly Gln Pro Thr Pro Lys Thr Pro Arg Gln Leu Arg Glu
1595 1600 1605
Phe Leu Gly Lys Ala Gly Phe Cys Arg Leu Phe Ile Pro Gly Phe
1610 1615 1620
Ala Glu Met Ala Ala Pro Leu Tyr Pro Leu Thr Lys Pro Gly Thr
1625 1630 1635
Leu Phe Asn Trp Gly Pro Asp Gln Gln Lys Ala Tyr Gln Glu Ile
1640 1645 1650
Lys Gln Ala Leu Leu Thr Ala Pro Ala Leu Gly Leu Pro Asp Leu
1655 1660 1665
Thr Lys Pro Phe Glu Leu Phe Val Asp Glu Lys Gln Gly Tyr Ala
1670 1675 1680
Lys Gly Val Leu Thr Gln Lys Leu Gly Pro Trp Arg Arg Pro Val
1685 1690 1695
Ala Tyr Leu Ser Lys Lys Leu Asp Pro Val Ala Ala Gly Trp Pro
1700 1705 1710
Pro Cys Leu Arg Met Val Ala Ala Ile Ala Val Leu Thr Lys Asp
1715 1720 1725
Ala Gly Lys Leu Thr Met Gly Gln Pro Leu Val Ile Leu Ala Pro
1730 1735 1740
His Ala Val Glu Ala Leu Val Lys Gln Pro Pro Asp Arg Trp Leu
1745 1750 1755
Ser Asn Ala Arg Met Thr His Tyr Gln Ala Leu Leu Leu Asp Thr
1760 1765 1770
Asp Arg Val Gln Phe Gly Pro Val Val Ala Leu Asn Pro Ala Thr
1775 1780 1785
Leu Leu Pro Leu Pro Glu Glu Gly Leu Gln His Asn Cys Leu Asp
1790 1795 1800
Ile Leu Ala Glu Ala His Gly Thr Arg Pro Asp Leu Thr Asp Gln
1805 1810 1815
Pro Leu Pro Asp Ala Asp His Thr Trp Tyr Thr Asp Gly Ser Ser
1820 1825 1830
Leu Leu Gln Glu Gly Gln Arg Lys Ala Gly Ala Ala Val Thr Thr
1835 1840 1845
Glu Thr Glu Val Ile Trp Ala Lys Ala Leu Pro Ala Gly Thr Ser
1850 1855 1860
Ala Gln Arg Ala Glu Leu Ile Ala Leu Thr Gln Ala Leu Lys Met
1865 1870 1875
Ala Glu Gly Lys Lys Leu Asn Val Tyr Thr Asp Ser Arg Tyr Ala
1880 1885 1890
Phe Ala Thr Ala His Ile His Gly Glu Ile Tyr Arg Arg Arg Gly
1895 1900 1905
Trp Leu Thr Ser Glu Gly Lys Glu Ile Lys Asn Lys Asp Glu Ile
1910 1915 1920
Leu Ala Leu Leu Lys Ala Leu Phe Leu Pro Lys Arg Leu Ser Ile
1925 1930 1935
Ile His Cys Pro Gly His Gln Lys Gly His Ser Ala Glu Ala Arg
1940 1945 1950
Gly Asn Arg Met Ala Asp Gln Ala Ala Arg Lys Ala Ala Ile Thr
1955 1960 1965
Glu Thr Pro Asp Thr Ser Thr Leu Leu Ile Glu Asn Ser Ser Pro
1970 1975 1980
Asn Ser Arg Leu Ile Asn Gly Ser Ser Gly Gly Ser Lys Arg Thr
1985 1990 1995
Ala Asp Gly Ser Glu Phe Glu Pro Lys Lys Lys Arg Lys Val Gly
2000 2005 2010
Ser Gly
2015
<210> 158
<211> 37
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 158
cgcggttcta tctagttacg cgttaaacca actagaa 37
<210> 159
<211> 23
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 159
gcatacgatg gaagaggtaa ctt 23
<210> 160
<211> 23
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 160
gcatacgatt aaagaggtaa ctt 23
<210> 161
<211> 23
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 161
caaatagtcc tctttcaaca ata 23
<210> 162
<211> 48
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 162
cgttgtaaag aataaggaaa tgattccgga agctttggaa gtactgaa 48
<210> 163
<211> 48
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 163
ataatgtcag atcctgtaga gaccacatca tccacggttc tatactgt 48
<210> 164
<211> 40
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 164
tgaagtcgag gactttggca tacgatggaa gaggtaactt 40
<210> 165
<211> 50
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 165
ccattcgtct tgaagtcgag gactttggca tacgatggaa gaggtaactt 50
<210> 166
<211> 72
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 166
tgttggaaga gatttgggtt ttccattcgt cttgaagtcg aggactttgg catacgatgg 60
aagaggtaac tt 72
<210> 167
<211> 47
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 167
ctaccttagc atcccttccc tttgcaaata gtcctctctc aacaata 47
<210> 168
<211> 55
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 168
ttcaccctct accttagcat cccttccctt tgcaaatagt cctctctcaa caata 55
<210> 169
<211> 63
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 169
ctgtaacgtt caccctctac cttagcatcc cttccctttg caaatagtcc tctctcaaca 60
ata 63
<210> 170
<211> 21
<212> DNA
<213> 人工序列
<220>
<223> 合成的LbCas12a crRNA支架
<400> 170
taatttctac taagtgtaga t 21
<210> 171
<211> 23
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 171
cctcactcct gctcggtgaa ttt 23
<210> 172
<211> 683
<212> PRT
<213> 莫洛尼鼠类白血病病毒
<400> 172
Thr Leu Asn Ile Glu Asp Glu His Arg Leu His Glu Thr Ser Lys Glu
1 5 10 15
Pro Asp Val Ser Leu Gly Ser Thr Trp Leu Ser Asp Phe Pro Gln Ala
20 25 30
Trp Ala Glu Thr Gly Gly Met Gly Leu Ala Val Arg Gln Ala Pro Leu
35 40 45
Ile Ile Pro Leu Lys Ala Thr Ser Thr Pro Val Ser Ile Lys Gln Tyr
50 55 60
Pro Met Ser Gln Glu Ala Arg Leu Gly Ile Lys Pro His Ile Gln Arg
65 70 75 80
Leu Leu Asp Gln Gly Ile Leu Val Pro Cys Gln Ser Pro Trp Asn Thr
85 90 95
Pro Leu Leu Pro Val Lys Lys Pro Gly Thr Asn Asp Tyr Arg Pro Val
100 105 110
Gln Asp Leu Arg Glu Val Asn Lys Arg Val Glu Asp Ile His Pro Thr
115 120 125
Val Pro Asn Pro Tyr Asn Leu Leu Ser Gly Leu Pro Pro Ser His Gln
130 135 140
Trp Tyr Thr Val Leu Asp Leu Lys Asp Ala Phe Phe Cys Leu Arg Leu
145 150 155 160
His Pro Thr Ser Gln Pro Leu Phe Ala Phe Glu Trp Arg Asp Pro Glu
165 170 175
Met Gly Ile Ser Gly Gln Leu Thr Trp Thr Arg Leu Pro Gln Gly Phe
180 185 190
Lys Asn Ser Pro Thr Leu Phe Asp Glu Ala Leu His Arg Asp Leu Ala
195 200 205
Asp Phe Arg Ile Gln His Pro Asp Leu Ile Leu Leu Gln Tyr Val Asp
210 215 220
Asp Leu Leu Leu Ala Ala Thr Ser Glu Leu Asp Cys Gln Gln Gly Thr
225 230 235 240
Arg Ala Leu Leu Gln Thr Leu Gly Asn Leu Gly Tyr Arg Ala Ser Ala
245 250 255
Lys Lys Ala Gln Ile Cys Gln Lys Gln Val Lys Tyr Leu Gly Tyr Leu
260 265 270
Leu Lys Glu Gly Gln Arg Trp Leu Thr Glu Ala Arg Lys Glu Thr Val
275 280 285
Met Gly Gln Pro Thr Pro Lys Thr Pro Arg Gln Leu Arg Glu Phe Leu
290 295 300
Gly Thr Ala Gly Phe Cys Arg Leu Trp Ile Pro Gly Phe Ala Glu Met
305 310 315 320
Ala Ala Pro Leu Tyr Pro Leu Thr Lys Thr Gly Thr Leu Phe Asn Trp
325 330 335
Gly Pro Asp Gln Gln Lys Ala Tyr Gln Glu Ile Lys Gln Ala Leu Leu
340 345 350
Thr Ala Pro Ala Leu Gly Leu Pro Asp Leu Thr Lys Pro Phe Glu Leu
355 360 365
Phe Val Asp Glu Lys Gln Gly Tyr Ala Lys Gly Val Leu Thr Gln Lys
370 375 380
Leu Gly Pro Trp Arg Arg Pro Val Ala Tyr Leu Ser Lys Lys Leu Asp
385 390 395 400
Pro Val Ala Ala Gly Trp Pro Pro Cys Leu Arg Met Val Ala Ala Ile
405 410 415
Ala Val Leu Thr Lys Asp Ala Gly Lys Leu Thr Met Gly Gln Pro Leu
420 425 430
Val Ile Leu Ala Pro His Ala Val Glu Ala Leu Val Lys Gln Pro Pro
435 440 445
Asp Arg Trp Leu Ser Asn Ala Arg Met Thr His Tyr Gln Ala Leu Leu
450 455 460
Leu Asp Thr Asp Arg Val Gln Phe Gly Pro Val Val Ala Leu Asn Pro
465 470 475 480
Ala Thr Leu Leu Pro Leu Pro Glu Glu Gly Leu Gln His Asn Cys Leu
485 490 495
Asp Ile Leu Ala Glu Ala His Gly Thr Arg Pro Asp Leu Thr Asp Gln
500 505 510
Pro Leu Pro Asp Ala Asp His Thr Trp Tyr Thr Asp Gly Ser Ser Leu
515 520 525
Leu Gln Glu Gly Gln Arg Lys Ala Gly Ala Ala Val Thr Thr Glu Thr
530 535 540
Glu Val Ile Trp Ala Lys Ala Leu Pro Ala Gly Thr Ser Ala Gln Arg
545 550 555 560
Ala Glu Leu Ile Ala Leu Thr Gln Ala Leu Lys Met Ala Glu Gly Lys
565 570 575
Lys Leu Asn Val Tyr Thr Asp Ser Arg Tyr Ala Phe Ala Thr Ala His
580 585 590
Ile His Gly Glu Ile Tyr Arg Arg Arg Gly Leu Leu Thr Ser Glu Gly
595 600 605
Lys Glu Ile Lys Asn Lys Asp Glu Ile Leu Ala Leu Leu Lys Ala Leu
610 615 620
Phe Leu Pro Lys Arg Leu Ser Ile Ile His Cys Pro Gly His Gln Lys
625 630 635 640
Gly His Ser Ala Glu Ala Arg Gly Asn Arg Met Ala Asp Gln Ala Ala
645 650 655
Arg Lys Ala Ala Ile Thr Glu Thr Pro Asp Thr Ser Thr Leu Leu Ile
660 665 670
Glu Asn Ser Ser Pro Asn Ser Arg Leu Ile Asn
675 680
<210> 173
<211> 122
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 173
acagcaggcc tttggtcagg ttggctgctg ggctggccct ggggccgtaa ccctcactcc 60
tgctcggtga atttggctca gcaggcacct gcctcagctg ctcacttgag cctctgggtc 120
ta 122
<210> 174
<211> 100
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 174
ggctgctggg ctggccctgg ggccgtaacc ctcactcctg ctcggtgaat ttggctcagc 60
aggcacctgc ctcagctgct cacttgagcc tctgggtcta 100
<210> 175
<211> 92
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 175
ggctggccct ggggccgtaa ccctcactcc tgctcggtga atttggctca gcaggcacct 60
gcctcagctg ctcacttgag cctctgggtc ta 92
<210> 176
<211> 84
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 176
ctggggccgt aaccctcact cctgctcggt gaatttggct cagcaggcac ctgcctcagc 60
tgctcacttg agcctctggg tcta 84
<210> 177
<211> 122
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 177
acagcaggcc tttggtcagg ttggctgctg ggctggccct ggggccgttt ccctcactcc 60
tggacggtga atttggctca gcaggcacct gcctcagctg ctcacttgag cctctgggtc 120
ta 122
<210> 178
<211> 122
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 178
acagcaggcc tttggtcagg ttggctgctg ggctggccct ggggccgttt ccctcactcc 60
tgctgcgtga atttggctca gcaggcacct gcctcagctg ctcacttgag cctctgggtc 120
ta 122
<210> 179
<211> 100
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 179
aatagcattg cagagaggcg tatcatttcg cggatgttgg aatcagtacg cagagagtcg 60
ccgtctccaa ggtgaaagcg gaagtagggc cttcgcgcac 100
<210> 180
<211> 100
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 180
aatagcattg cagagaggcg tatcatttcg cggatgttcc tttcagtacg cagagagtcg 60
ccgtctccaa ggtgaaagcg gaagtagggc cttcgcgcac 100
<210> 181
<211> 122
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 181
acagcaggcc tttggtcagg ttggctgctg ggctggccct ggggccgtaa ccctcactcc 60
tgctcggtga atttggctca gcaggcacct gcctcagctg ctcacttgag cctctgggtc 120
ta 122
<210> 182
<211> 100
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 182
ggctgctggg ctggccctgg ggccgtaacc ctcactcctg ctcggtgaat ttggctcagc 60
aggcacctgc ctcagctgct cacttgagcc tctgggtcta 100
<210> 183
<211> 92
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 183
ggctggccct ggggccgtaa ccctcactcc tgctcggtga atttggctca gcaggcacct 60
gcctcagctg ctcacttgag cctctgggtc ta 92
<210> 184
<211> 84
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 184
ctggggccgt aaccctcact cctgctcggt gaatttggct cagcaggcac ctgcctcagc 60
tgctcacttg agcctctggg tcta 84
<210> 185
<211> 122
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 185
gcactctgcc acttattggg tcagctgtta acatcagtac gttaatgtaa cctgatggtc 60
catgtctgtt actcgcctgt caagtggcgt gacaccgggc gtgttcccca gagtgacttt 120
tc 122
<210> 186
<211> 100
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 186
agctgttaac atcagtacgt taatgtaacc tgatggtcca tgtctgttac tcgcctgtca 60
agtggcgtga caccgggcgt gttccccaga gtgacttttc 100
<210> 187
<211> 27
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 187
tttggctcac tcctgctcgg tgaattt 27
<210> 188
<211> 27
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 188
tttcgcgctt gttccaatca gtacgca 27
<210> 189
<211> 20
<212> DNA
<213> 氨基酸球菌属物种
<400> 189
taatttctac tcttgtagat 20
<210> 190
<211> 19
<212> DNA
<213> 噬菌体lambda
<400> 190
gggccctgaa gaagggccc 19
<210> 191
<211> 42
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 191
ttgacgcggt tctatctagt tacgcgttaa accaacuaga aa 42
<210> 192
<211> 63
<212> DNA
<213> 莫洛尼鼠类白血病病毒
<400> 192
ggaggtcagg gtcaggagcc cccccctgaa cccaggataa ccctcaaagt cggggggcaa 60
ccc 63
<210> 193
<211> 55
<212> DNA
<213> 莫洛尼鼠类白血病病毒
<400> 193
gggtcaggag cccccccctg aacccaggat aaccctcaaa gtcggggggc aaccc 55
<210> 194
<211> 34
<212> DNA
<213> 噬菌体MS2
<400> 194
ggccaacatg aggatcaccc atgtctgcag ggcc 34
<210> 195
<211> 29
<212> DNA
<213> 假单胞菌（Pseudomonas）噬菌体PP7
<400> 195
ccggagcaga cgatatggcg tcgctccgg 29
<210> 196
<211> 26
<212> DNA
<213> 智人
<400> 196
ccaaaggctc ttctcagagc caccca 26
<210> 197
<211> 31
<212> DNA
<213> 人免疫缺陷病毒
<400> 197
ggccagatct gagcctggga gctctctggc c 31
<210> 198
<211> 36
<212> DNA
<213> 嗜热四膜虫（Tetrahymena thermophila）
<400> 198
atataacctt caccattagg ttcaaataat ggtaat 36
<210> 199
<211> 269
<212> PRT
<213> 酿酒糖酵母（Saccharomyces cerevisiae）
<400> 199
Asp Met Asp Ser Ile Gly Glu Ser Glu Val Arg Ala Leu Tyr Lys Ala
1 5 10 15
Ile Leu Lys Phe Gly Asn Leu Lys Glu Ile Leu Asp Glu Leu Ile Ala
20 25 30
Asp Gly Thr Leu Pro Val Lys Ser Phe Glu Lys Tyr Gly Glu Thr Tyr
35 40 45
Asp Glu Met Met Glu Ala Ala Lys Asp Cys Val His Glu Glu Glu Lys
50 55 60
Asn Arg Lys Glu Ile Leu Glu Lys Leu Glu Lys His Ala Thr Ala Tyr
65 70 75 80
Arg Ala Lys Leu Lys Ser Gly Glu Ile Lys Ala Glu Asn Gln Pro Lys
85 90 95
Asp Asn Pro Leu Thr Arg Leu Ser Leu Lys Lys Arg Glu Lys Lys Ala
100 105 110
Val Leu Phe Asn Phe Lys Gly Val Lys Ser Leu Asn Ala Glu Ser Leu
115 120 125
Leu Ser Arg Val Glu Asp Leu Lys Tyr Leu Lys Asn Leu Ile Asn Ser
130 135 140
Asn Tyr Lys Asp Asp Pro Leu Lys Phe Ser Leu Gly Asn Asn Thr Pro
145 150 155 160
Lys Pro Val Gln Asn Trp Ser Ser Asn Trp Thr Lys Glu Glu Asp Glu
165 170 175
Lys Leu Leu Ile Gly Val Phe Lys Tyr Gly Tyr Gly Ser Trp Thr Gln
180 185 190
Ile Arg Asp Asp Pro Phe Leu Gly Ile Thr Asp Lys Ile Phe Leu Asn
195 200 205
Glu Val His Asn Pro Val Ala Lys Lys Ser Ala Ser Ser Ser Asp Thr
210 215 220
Thr Pro Thr Pro Ser Lys Lys Gly Lys Gly Ile Thr Gly Ser Ser Lys
225 230 235 240
Lys Val Pro Gly Ala Ile His Leu Gly Arg Arg Val Asp Tyr Leu Leu
245 250 255
Ser Phe Leu Arg Gly Gly Leu Asn Thr Lys Ser Pro Ser
260 265
<210> 200
<211> 80
<212> PRT
<213> 智人
<400> 200
Ser Thr Asp His Pro Lys Tyr Ser Asp Met Ile Val Ala Ala Ile Gln
1 5 10 15
Ala Glu Lys Asn Arg Ala Gly Ser Ser Arg Gln Ser Ile Gln Lys Tyr
20 25 30
Ile Lys Ser His Tyr Lys Val Gly Glu Asn Ala Asp Ser Gln Ile Lys
35 40 45
Leu Ser Ile Lys Arg Leu Val Thr Thr Gly Val Leu Lys Gln Thr Lys
50 55 60
Gly Val Gly Ala Ser Gly Ser Phe Arg Leu Ala Lys Ser Asp Glu Pro
65 70 75 80
<210> 201
<211> 83
<212> PRT
<213> 智人
<400> 201
Gly Lys Gly Asp Pro Lys Lys Pro Arg Gly Lys Met Ser Ser Tyr Ala
1 5 10 15
Phe Phe Val Gln Thr Cys Arg Glu Glu His Lys Lys Lys His Pro Asp
20 25 30
Ala Ser Val Asn Phe Ser Glu Phe Ser Lys Lys Cys Ser Glu Arg Trp
35 40 45
Lys Thr Met Ser Ala Lys Glu Lys Gly Lys Phe Glu Asp Met Ala Lys
50 55 60
Ala Asp Lys Ala Arg Tyr Glu Arg Glu Met Lys Thr Tyr Ile Pro Pro
65 70 75 80
Lys Gly Glu
<210> 202
<211> 100
<212> PRT
<213> 猎神狒狒（Papio anubis）
<400> 202
Met Pro Lys Arg Lys Val Ser Ser Ala Glu Gly Ala Ala Lys Glu Glu
1 5 10 15
Pro Lys Arg Arg Ser Ala Arg Leu Ser Ala Lys Pro Pro Ala Lys Val
20 25 30
Glu Ala Lys Pro Lys Lys Ala Ala Ala Lys Asp Lys Ser Ser Asp Lys
35 40 45
Lys Val Gln Thr Lys Gly Lys Arg Gly Ala Lys Gly Lys Gln Ala Glu
50 55 60
Val Ala Asn Gln Glu Thr Lys Glu Asp Leu Pro Ala Glu Asn Gly Glu
65 70 75 80
Thr Lys Thr Glu Glu Ser Pro Ala Ser Asp Glu Ala Gly Glu Lys Glu
85 90 95
Ala Lys Ser Asp
100
<210> 203
<211> 21
<212> RNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 203
uaagucucca uagaauggag g 21
<210> 204
<211> 102
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 204
gaacgtcacc tggaaccgac cttgctggcc gtacatttat acggctccgc agtggatggc 60
ggctgaaagc cacacagtga tattgatttg ctggttacgg tg 102
<210> 205
<211> 21
<212> PRT
<213> 人工序列
<220>
<223> 合成的肽
<400> 205
Glu Arg His Leu Glu Pro Thr Leu Leu Ala Val His Leu Tyr Gly Ser
1 5 10 15
Ala Val Asp Gly Gly
20
<210> 206
<211> 12
<212> PRT
<213> 人工序列
<220>
<223> 合成的肽
<400> 206
Lys Pro His Ser Asp Ile Asp Leu Leu Val Thr Val
1 5 10
<210> 207
<211> 102
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 207
gaacgtcacc tggaaccgac cttgctggcc gtacatttat acggctccgc agtggatggc 60
ggcctgaagc cacacagtga tattgatttg ctggttacgg tg 102
<210> 208
<211> 34
<212> PRT
<213> 人工序列
<220>
<223> 合成的肽
<400> 208
Glu Arg His Leu Glu Pro Thr Leu Leu Ala Val His Leu Tyr Gly Ser
1 5 10 15
Ala Val Asp Gly Gly Leu Lys Pro His Ser Asp Ile Asp Leu Leu Val
20 25 30
Thr Val
<210> 209
<211> 98
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 209
catgcccgca aaaacgtggc gtatgtactc gtgtatatac taccactcct aaaaaaccga 60
actccgcgct gcgtaaagta tgccgtgttc gtctgact 98
<210> 210
<211> 33
<212> PRT
<213> 人工序列
<220>
<223> 合成的肽
<400> 210
Ala Cys Pro Gln Lys Arg Gly Val Cys Thr Arg Val Tyr Thr Thr Thr
1 5 10 15
Pro Lys Lys Pro Asn Ser Ala Leu Arg Lys Val Cys Arg Val Arg Leu
20 25 30
Thr
<210> 211
<211> 98
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 211
catgcccgca aaaacgtggc gtatgtactc gtgtatatac taccactcct cgtaaaccga 60
actccgcgct gcgtaaagta tgccgtgttc gtctgact 98
<210> 212
<211> 120
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 212
atttatacgg ctccgcagtg gatggcggcc tgaagccaca cagtgatatt gatttgctgg 60
ttacggtgac cgtacgcctt tgagaaacaa cgcgccgcgc tttgatcaac gaccttttgg 120
<210> 213
<211> 27
<212> PRT
<213> 人工序列
<220>
<223> 合成的肽
<400> 213
His Leu Tyr Gly Ser Ala Val Asp Gly Gly Leu Lys Pro His Ser Asp
1 5 10 15
Ile Asp Leu Leu Val Thr Val Thr Val Arg Leu
20 25
<210> 214
<211> 12
<212> PRT
<213> 人工序列
<220>
<223> 合成的肽
<400> 214
Glu Thr Thr Arg Arg Ala Leu Ile Asn Asp Leu Leu
1 5 10
<210> 215
<211> 120
<212> DNA
<213> 人工序列
<220>
<223> 合成的寡核苷酸
<400> 215
atttatacgg ctccgcagtg gatggcggcc tgaagccaca cagtgatatt gatttgctgg 60
ttacggtgac cgtacgcctt gatgaaacaa cgcgccgcgc tttgatcaac gaccttttgg 120

Claims (173)

1.修饰靶核酸的方法，所述方法包括：
使所述靶核酸与下列组分相接触：
(a)V型CRISPR-Cas效应蛋白或II型CRISPR-Cas效应蛋白；
(b)逆转录酶，和
(c)延伸型指导核酸(例如，延伸型II型或V型CRISPR RNA、延伸型II型或V型CRISPRDNA、延伸型II型或V型crRNA、延伸型II型或V型crDNA)，
由此修饰所述靶核酸。
2.权利要求1的方法，其中所述V型CRISPR-Cas效应蛋白或所述II型CRISPR-Cas效应蛋白、所述逆转录酶和所述延伸型指导核酸形成复合物或被包含在复合物中。
3.权利要求1或权利要求2的方法，其中所述延伸型指导核酸包含：
(i)V型CRISPR核酸或II型CRISPR核酸(II型或V型CRISPR RNA、II型或V型CRISPR DNA、II型或V型crRNA、II型或V型crDNA)和/或V型CRISPR核酸或II型CRISPR核酸和tracr核酸(例如，II型或V型tracrRNA、II型或V型tracrDNA)；和
(ii)包含引物结合位点和逆转录酶模板(RT模板)(RTT)的延伸部分。
4.权利要求3的方法，其中所述延伸部分融合至所述CRISPR核酸的5'末端或3'末端(例如，5'至3'：重复序列-间隔子-延伸部分，或者延伸部分-重复序列-间隔子)，和/或融合至所述tracr核酸的5'或3'末端。
5.权利要求3或权利要求4的方法，其中所述延伸型指导核酸的延伸部分从5'至3'包含RT模板和引物结合位点。
6.权利要求1-5中任一项的方法，其中所述延伸型指导核酸进一步包含结构化RNA基序。
7.权利要求6的方法，其中所述结构化RNA基序位于所述延伸型指导核酸的3'末端处。
8.权利要求6或权利要求7的方法，其中所述结构化RNA基序为AsCpf1BB(SEQ ID NO:189)、BoxB(SEQ ID NO:190)、假结(诱饵)(SEQ ID NO:95、SEQ ID NO:203)、假结(tEvoPreQ1)(SEQ ID NO:191)、fmp结(SEQ ID NO:192)、mp结(SEQ ID NO:193)、MS2(SEQID NO:194)、PP7(SEQ ID NO:195)、SLBP(SEQ ID NO:196)、TAR(SEQ ID NO:197)和/或ThermoPh(SEQ ID NO:198)。
9.权利要求6-8中任一项的方法，其中所述结构化RNA基序为假结。
10.权利要求9的方法，其中所述假结为包含SEQ ID NO:158的核酸序列的tEvoPreQ1假结或者包含SEQ ID NO:191的核酸序列的EvoPreQ1假结。
11.权利要求9的方法，其中所述假结包含SEQ ID NO:95或SEQ ID NO:203的核酸序列。
12.权利要求5-11中任一项的方法，其中所述靶核酸是双链的并且包含第一链和第二链，并且所述引物结合位点与所述靶核酸的第二链(非靶标，上部链)相结合。
13.权利要求5-11中任一项的方法，其中所述靶核酸是双链的并且包含第一链和第二链，并且所述引物结合位点与所述靶核酸的第一链相结合(例如，结合至所述靶标链，所述CRISPR-Cas效应蛋白被募集至的同一链，下部链)。
14.权利要求5-11中任一项的方法，其中所述靶核酸是双链的并且包含第一链和第二链，并且所述引物结合位点与所述靶核酸的第二链(非靶标链，与所述CRISPR-Cas效应蛋白被募集至的链相反的链)相结合。
15.权利要求3-14中任一项的方法，其中所述引物结合位点(PBS)的长度为大约一个核苷酸至大约100个核苷酸，任选地其中所述PBS的长度为大约4个核苷酸至大约80个核苷酸，大约30个核苷酸至大约80个核苷酸，大约40个核苷酸至大约60个核苷酸，或者长度为大约8、16、24、32、40、48、56、64、72或80个核苷酸。
16.权利要求3-15中任一项的方法，其中所述RT模板(RTT)的长度为大约一个至大约100个核苷酸，任选地其中所述RTT的长度为大约35个核苷酸至大约100个核苷酸，大约35个核苷酸至大约80个核苷酸，任选地大约35个核苷酸至大约75个核苷酸，大约40个核苷酸至大约75个核苷酸，大约45个核苷酸至大约75个核苷酸，大约45个核苷酸至大约60个核苷酸，或者长度为大约36、40、44、47、50、52、55、63、72或74个核苷酸。
17.权利要求3至16中任一项的方法，其中所述RTT的长度为大约35个核苷酸至大约75个核苷酸并且所述PBS的长度为大约30个核苷酸至大约80个核苷酸，任选地其中所述PBS的长度为大约8、16、24、32、40、48、56、64、72或80个核苷酸并且所述RTT的长度为大约36、40、44、47、50、52、55、63、72或74个核苷酸。
18.权利要求3至17中任一项的方法，其中所述延伸型指导RNA的延伸部分经由连接体与所述CRISPR核酸和/或所述tracrRNA相连接。
19.权利要求18的方法，其中所述连接体的长度为1至100个核苷酸。
20.权利要求3至19中任一项的方法，其中当所述延伸部分位于所述crRNA的5'时，修饰所述V型CRISPR-Cas效应蛋白以降低(或消除)自加工RNA酶活性。
21.前述权利要求中任一项的方法，其中所述V型CRISPR-Cas效应蛋白或所述II型CRISPR-Cas效应蛋白为融合蛋白和/或所述逆转录酶为融合蛋白，其中所述V型CRISPR-Cas融合蛋白或II型CRISPR-Cas融合蛋白、所述逆转录酶融合蛋白和/或所述延伸型指导核酸融合至一个或多个将所述逆转录酶募集至所述V型CRISPR-Cas效应蛋白或II型CRISPR-Cas效应蛋白的组分，任选地所述一个或多个组分经由蛋白质-蛋白质相互作用、蛋白质-RNA相互作用和/或化学相互作用来进行募集。
22.前述权利要求中任一项的方法，其中所述V型CRISPR-Cas效应蛋白为包含融合(连接)至肽标签(例如，表位或多聚化表位)的V型CRISPR-Cas效应蛋白结构域的V型CRISPR-Cas效应融合蛋白，并且所述逆转录酶为包含融合(连接)至与所述肽标签相结合的亲和多肽的逆转录酶结构域的逆转录酶融合蛋白，任选地其中所述靶核酸与两个或更多个逆转录酶融合蛋白相接触，或者所述II型CRISPR-Cas效应蛋白为包含融合(连接)至肽标签(例如，表位或多聚化表位)的II型CRISPR-Cas效应蛋白结构域的II型CRISPR-Cas效应融合蛋白，并且所述逆转录酶为包含融合(连接)至与所述肽标签相结合的亲和多肽的逆转录酶结构域的逆转录酶融合蛋白，任选地其中所述靶核酸与两个或更多个逆转录酶融合蛋白相接触。
23.权利要求22的方法，其中所述肽标签包括GCN4肽标签(例如，Sun-Tag)、c-Myc亲和标签、HA亲和标签、His亲和标签、S亲和标签、甲硫氨酸-His亲和标签、RGD-His亲和标签、FLAG八肽、strep标签或strep标签II、V5标签和/或VSV-G表位。
24.权利要求22或权利要求23的方法，其中所述肽标签包括两个或更多个拷贝的所述肽标签。
25.权利要求22-24中任一项的方法，其中所述亲和多肽为抗体、affibody、anticalin、monobody和/或DARPin。
26.权利要求25的方法，其中所述抗体为scFv抗体。
27.前述权利要求中任一项的方法，其中所述延伸型指导核酸连接至RNA募集基序，并且所述逆转录酶为包含融合(连接)至与所述RNA募集基序相结合的亲和多肽的逆转录酶结构域的逆转录酶融合蛋白，任选地其中所述靶核酸连接至两个或更多个逆转录酶融合蛋白相接触，任选地其中所述延伸型指导RNA连接至两个或更多个RNA募集基序，任选地其中所述两个或更多个RNA募集基序为相同的RNA募集基序或不同的RNA募集基序。
28.权利要求27的方法，其中所述募集基序位于所述延伸型指导核酸的延伸部分的3'末端处或者包埋在所述延伸部分中。
29.权利要求27或权利要求28的方法，其中所述RNA募集基序和相应的亲和多肽为端粒酶Ku结合基序(例如，Ku结合发夹)和Ku的亲和多肽(例如，Ku异二聚体)；端粒酶Sm7结合基序和Sm7的亲和多肽；MS2噬菌体操纵基因茎-环和亲和多肽MS2外壳蛋白(MCP)；PP7噬菌体操纵基因茎-环和亲和多肽PP7外壳蛋白(PCP)；SfMu噬菌体Com茎-环和亲和多肽Com RNA结合蛋白；PUF结合位点(PBS)和亲和多肽Pumilio/fem-3mRNA结合因子(PUF)；和/或合成的RNA-适体和相应的适体配体。
30.权利要求27-29中任一项的方法，其中所述RNA募集基序和相应的亲和多肽为MS2噬菌体操纵基因茎-环和亲和多肽MS2外壳蛋白(MCP)，和/或PUF结合位点(PBS)和亲和多肽Pumilio/fem-3mRNA结合因子(PUF)。
31.权利要求21的方法，其中募集化学相互作用的所述一个或多个组分为雷帕霉素可诱导的FRB–FKBP的二聚化；生物素-链霉抗生物素蛋白；SNAP标签；Halo标签；CLIP标签；由化合物诱导的DmrA-DmrC异二聚体；双功能配体(例如，两个蛋白质结合化学品融合在一起，例如二氢叶酸还原酶(DHFR))。
32.前述权利要求中任一项的方法，其进一步包括使所述靶核酸与下列组分相接触：
(a)CRISPR-Cas效应蛋白；和
(b)指导核酸，
其中(i)所述CRISPR-Cas效应蛋白对于位于离已经由所述II型或V型CRISPR-Cas效应蛋白产生切口的在第二链上的位点大约10至大约125个碱基对处(5'或3')的在所述靶核酸的第一链上的位点产生切口或进行切割，或(ii)所述CRISPR-Cas效应蛋白对于位于离已经由所述II型或V型CRISPR-Cas效应蛋白产生切口的在第一链上的位点大约10至大约125个碱基对处(5'或3')的在所述靶核酸的第二链上的位点产生切口或进行切割，由此改善错配修复，其中所述CRISPR-Cas效应蛋白为I型、II型、III型、IV型或V型CRISPR-Cas效应蛋白。
33.权利要求21-32中任一项的方法，其中所述V型CRISPR-Cas融合蛋白或II型CRISPR-Cas融合蛋白融合至染色质调节肽。
34.权利要求21-33中任一项的方法，其中所述染色质调节肽融合至所述V型CRISPR-Cas融合蛋白或II型CRISPR-Cas融合蛋白的C-末端和/或N-末端。
35.权利要求21-34中任一项的方法，其中所述V型CRISPR-Cas融合蛋白或II型CRISPR-Cas融合蛋白在其N-末端处融合至逆转录酶，并且所述染色质调节肽融合至所述逆转录酶的N-末端。
36.权利要求33-35中任一项的方法，其中所述染色质调节肽为CHD1(例如，SEQ ID NO:199)、H1G(例如，SEQ ID NO:200)、HB1(例如，SEQ ID NO:201)和HN1(例如，SEQ ID NO:202)。
37.前述权利要求中任一项的方法，其进一步包括使所述靶核酸与Dna2多肽和/或5'翼片核酸内切酶(FEN)，任选地FEN1多肽相接触。
38.权利要求37的方法，其中所述FEN和/或Dna2多肽被过表达(在所述靶核酸存在下)。
39.权利要求37或权利要求38的方法，其中所述FEN为包含融合至所述II型或V型CRISPR-Cas效应蛋白或结构域的FEN结构域的融合蛋白，和/或其中所述Dna2多肽为包含融合至所述II型或V型CRISPR-Cas效应蛋白或结构域的Dna2结构域的融合蛋白。
40.前述权利要求中任一项的方法，其中所述V型CRISPR-Cas效应蛋白为包含融合(连接)至肽标签(例如，表位或多聚化表位)的V型CRISPR-Cas效应蛋白结构域的V型CRISPR-Cas融合蛋白，并且所述FEN为包含融合至与所述肽标签相结合的亲和多肽的FEN结构域的FEN融合蛋白，和/或其中所述V型CRISPR-Cas效应蛋白为包含融合至肽标签的V型CRISPR-Cas效应蛋白结构域的V型CRISPR-Cas融合蛋白，并且所述Dna2多肽为包含融合至与所述肽标签相结合的亲和多肽的Dna2结构域的Dna2融合蛋白，任选地其中所述靶核酸与两个或更多个FEN融合蛋白和/或两个或更多个Dna2融合蛋白相接触，由此将所述FEN和/或Dna2募集至所述V型CRISPR-Cas效应蛋白结构域和所述靶核酸。
41.权利要求1-39中任一项的方法，其中所述II型CRISPR-Cas效应蛋白为包含融合(连接)至肽标签(例如，表位或多聚化表位)的II型CRISPR-Cas效应蛋白结构域的II型CRISPR-Cas融合蛋白，并且所述FEN为包含融合至与所述肽标签相结合的亲和多肽的FEN结构域的FEN融合蛋白，和/或其中所述II型CRISPR-Cas效应蛋白为包含融合至肽标签的II型CRISPR-Cas效应蛋白结构域的II型CRISPR-Cas融合蛋白，并且所述Dna2多肽为包含融合至与所述肽标签相结合的亲和多肽的Dna2结构域的Dna2融合蛋白，任选地其中所述靶核酸与两个或更多个FEN融合蛋白和/或两个或更多个Dna2融合蛋白相接触，由此将所述FEN和/或Dna2募集至所述II型CRISPR-Cas效应蛋白结构域和所述靶核酸。
42.前述权利要求中任一项的方法，其中所述V型CRISPR-Cas效应蛋白为Cas12a(Cpf1)结构域、Cas12b(C2c1)结构域、Cas12c(C2c3)结构域、Cas12d(CasY)结构域、Cas12e(CasX)结构域、Cas12g结构域、Cas12h结构域、Cas12i结构域、C2c4结构域、C2c5结构域、C2c8结构域、C2c9结构域、C2c10结构域、Cas14a结构域、Cas14b结构域和/或Cas14c结构域，或者所述II型CRISPR-Cas效应蛋白为Cas9结构域，任选地其中所述V型CRISPR-Cas效应蛋白和/或所述II型CRISPR-Cas效应蛋白包含一个或多个在本文中所描述的突变。
43.权利要求42的方法，其中所述V型CRISPR-Cas效应蛋白为包含通过参考SEQ ID NO:9的氨基酸位置编号而言的R1138A突变的LbCas12a切口酶，包含通过参考SEQ ID NO:2的氨基酸位置编号而言的R1226A突变的AsCas12a切口酶，包含通过参考SEQ ID NO:6的氨基酸位置编号而言的R1228A的FnCas12a，或包含通过参考SEQ ID NO:14的氨基酸位置编号而言的R1241A突变的PdCas12a切口酶。
44.权利要求42的方法，其中所述V型CRISPR-Cas效应蛋白为来自氨基酸球菌属物种(Acidaminococcus sp.)的V型CRISPR-Cas效应蛋白(AsCas12a)或来自毛螺菌科(Lachnospiraceae)细菌的V型CRISPR-Cas效应蛋白(例如，LbCas12a)，或者从其而来的经修饰的V型CRISPR-Cas效应蛋白。
45.权利要求42的方法，其中所述V型CRISPR-Cas效应蛋白为来自亨盖特氏丁酸弧菌(Butyrivibrio hungatei)的V型CRISPR-Cas效应蛋白(BhCas12)，或者从其而来的经修饰的V型CRISPR-Cas效应蛋白。
46.权利要求42的方法，其中所述II型CRISPR-Cas效应蛋白为Cas9效应蛋白/结构域，任选地来自链球菌属(Streptococcus)。
47.权利要求1-40或42-46中任一项的方法，其中所述V型CRISPR-Cas效应蛋白包含降低的单链DNA切割活性(ss DNA酶活性)，或者被修饰(突变)以降低(或消除)ss DNA酶活性。
48.权利要求1-40或42-47中任一项的方法，其中所述V型CRISPR-Cas效应蛋白被修饰(突变)以降低(或消除)自加工RNA酶活性。
49.权利要求48的方法，其中所述V型CRISPR-Cas效应蛋白为具有通过参考SEQ ID NO:1的氨基酸位置编号而言的H759A突变的Cas12a CRISPR-Cas效应蛋白。
50.权利要求49的方法，其中所述具有H759A突变的Cas12aCRISPR-Cas效应蛋白包含与SEQ ID NO:148的氨基酸序列具有至少80％序列同一性的序列。
51.权利要求50的方法，其中所述具有H759A突变的Cas12aCRISPR-Cas效应蛋白为与SEQ ID NO:148的氨基酸序列具有至少90％序列同一性的LbCas12a CRISPR-Cas效应蛋白。
52.前述权利要求中任一项的方法，其中所述V型CRISPR-Cas效应蛋白被修饰以降低或消除核酸酶活性(例如，在核酸酶活性位点中(例如，在RuvC结构域中)的突变)，或者所述II型CRISPR-Cas效应蛋白被修饰以降低或消除核酸酶活性(例如，核酸酶活性位点中(例如，在RuvC或HNH结构域中)的突变)，以产生失活的V型CRISPR-Cas效应蛋白或失活的II型CRISPR-Cas效应蛋白(例如，deadCas(dCas、dCas12a、dCas9))。
53.权利要求52的方法，其中所述失活的V型CRISPR-Cas效应蛋白或所述失活的II型CRISPR-Cas效应蛋白包含切口酶活性。
54.权利要求52或权利要求53的方法，其中所述失活的V型CRISPR-Cas效应蛋白或所述失活的II型CRISPR-Cas效应蛋白融合至产生切口的酶(例如，Fok1、BFi1，例如经改造的Fok1或BFiI)。
55.前述权利要求中任一项的方法，其中所述V型CRISPR-Cas效应蛋白包含氨基酸修饰以降低所述V型CRISPR-Cas效应蛋白的DNA结合亲和力。
56.权利要求55的方法，其中所述修饰为氨基酸置换。
57.权利要求55或权利要求56的方法，其中所述修饰为至丙氨酸的氨基酸置换，任选地其中所述丙氨酸替换带正电荷的氨基酸残基。
58.权利要求57的方法，其中所述氨基酸置换为通过参考SEQ ID NO:148的氨基酸位置编号而言的K167A、K272A和/或K349A。
59.权利要求58的方法，其中所述V型CRISPR-Cas效应蛋白为包含通过参考SEQ ID NO:148的氨基酸位置编号而言的K167A、K272A、K349A、K167A+K272A、K167A+K349A、K272A+K349A或K167A+K272A+K349A的氨基酸置换的Cas12a CRISPR-Cas效应蛋白，任选地其中所述V型CRISPR-Cas效应蛋白为LbCas12a。
60.前述权利要求中任一项的方法，其中所述逆转录酶包含氨基酸修饰。
61.权利要求60的方法，其中所述氨基酸修饰为氨基酸置换。
62.权利要求61的方法，其中所述氨基酸置换为通过参考SEQ ID NO:172的氨基酸位置编号而言的L139P、D200N、W388R、E607K、T306K、W313F、F155Y、H638G、Q221R、V223M和/或D524N。
63.权利要求61或权利要求62的方法，其中所述逆转录酶包含通过参考SEQ ID NO:172的氨基酸位置编号而言的L139P、D200N、W388R和E607K；L139P、D200N、T306K、W313F、W388R和E607K；5M(T355A/Q357M/K358R/A359G/S360A)、F155Y和H638G；5M(T355A/Q357M/K358R/A359G/S360A)、Q221R和V223M；或5M(T355A/Q357M/K358R/A359G/S360A)和D524N的氨基酸置换。
64.前述权利要求中任一项的方法，其中所述V型CRISPR-Cas效应蛋白为包含融合至所述逆转录酶的V型CRISPR-Cas效应蛋白结构域的V型CRISPR-Cas融合蛋白，或者所述II型CRISPR-Cas效应蛋白为包含融合至所述逆转录酶的II型CRISPR-Cas效应蛋白结构域的II型CRISPR-Cas融合蛋白。
65.权利要求64的方法，其中所述逆转录酶融合至所述V型CRISPR-Cas效应多肽或所述II型CRISPR-Cas效应多肽的C-末端。
66.权利要求65的方法，其中所述逆转录酶融合至所述V型CRISPR-Cas效应多肽或所述II型CRISPR-Cas效应多肽的N-末端。
67.前述权利要求中任一项的方法，其中所述逆转录酶融合至一个或多个单链RNA结合结构域(RBD)，由此改善所述逆转录酶的热稳定性、持续能力和模板亲和力。
68.权利要求67的方法，其中所述一个或多个单链RNA结合结构域融合至所述逆转录酶的N-末端，任选地其中所述逆转录酶进一步在其C-末端处融合至所述II型CRISPR-Cas效应蛋白和/或V型CRISPR-Cas效应蛋白的N-末端。
69.前述权利要求中任一项的方法，其进一步包括使所述靶核酸与5'-3'核酸外切酶相接触。
70.权利要求69的方法，其中所述5'-3'核酸外切酶融合至V型CRISPR-Cas效应蛋白，任选地融合至V型CRISPR-Cas融合蛋白，或者所述5'-3'核酸外切酶融合至II型CRISPR-Cas效应蛋白，任选地融合至II型CRISPR-Cas融合蛋白。
71.权利要求70的方法，其中所述5'-3'核酸外切酶融合至所述V型CRISPR-Cas效应蛋白的C-末端，任选地融合至所述V型CRISPR-Cas融合蛋白的C-末端，或者融合至所述II型CRISPR-Cas效应蛋白的C-末端，任选地融合至所述II型CRISPR-Cas融合蛋白的C-末端。
72.权利要求69的方法，其中所述5'-3'核酸外切酶为包含融合至肽标签的5'-3'核酸外切酶的融合蛋白，并且所述II型或V型CRISPR-Cas效应蛋白为包含融合至能够与所述肽标签相结合的亲和多肽的II型或V型CRISPR-Cas效应蛋白结构域的融合蛋白。
73.权利要求69的方法，其中所述5'-3'核酸外切酶为包含融合至能够与所述肽标签相结合的亲和多肽的5'-3'核酸外切酶的融合蛋白，并且所述II型或V型CRISPR-Cas效应蛋白为包含融合至肽标签的II型或V型CRISPR-Cas效应蛋白结构域的融合蛋白。
74.权利要求69的方法，其中所述5'-3'核酸外切酶为包含融合至能够与RNA募集基序相结合的亲和多肽的5'-3'核酸外切酶的融合蛋白，并且所述延伸型指导核酸连接至RNA募集基序。
75.权利要求69-74中任一项的方法，其中所述5'-3'核酸外切酶为RecE核酸外切酶(RecE)、RecJ核酸外切酶(RecJ)、T5核酸外切酶(T5_Exo)、T7核酸外切酶(T7_Exo)、λ核酸外切酶(λ_exo)、sbcB(脱氧核糖核酸外切酶)或hExo1(人核酸外切酶1)。
76.前述权利要求中任一项的方法，其进一步包括使所述靶核酸与单链DNA结合蛋白(ssDNA结合蛋白)相接触。
77.权利要求76的方法，其中所述ssDNA结合蛋白融合至所述IIV型CRISPR-Cas效应蛋白或V型CRISPR-Cas效应蛋白。
78.权利要求77的方法，其中所述ssDNA结合蛋白融合至所述II型或V型CRISPR-Cas效应蛋白的C-末端。
79.权利要求77的方法，其中所述ssDNA结合蛋白融合至所述II型或V型CRISPR-Cas效应蛋白的N-末端。
80.权利要求76至79中任一项的方法，其中所述ssDNA结合蛋白来自人、细菌或噬菌体。
81.权利要求76至80中任一项的方法，其中所述ssDNA结合蛋白为hRad51(任选地，hRad51_S208E_A209D)、hRad52、BsRecA、EcRecA、T4ssB和/或Brex27。
82.前述权利要求中任一项的方法，其进一步包括通过引入非同源末端连接(NHEJ)的化学抑制剂，通过引入靶向NHEJ蛋白的CRISPR指导核酸或siRNA以瞬时敲低所述NHEJ蛋白的表达，或者通过引入阻止NHEJ的多肽来减少双链断裂。
83.权利要求82的方法，其中所述阻止NHEJ的多肽融合至所述逆转录酶和/或所述CRISPR-Cas效应蛋白，任选地所述阻止NHEJ的多肽融合至所述逆转录酶和/或所述CRISPR-Cas效应蛋白的N-末端。
84.权利要求82或权利要求83的方法，其中所述阻止NHEJ的多肽为Gam蛋白。
85.权利要求84的方法，其中所述Gam蛋白为埃希氏菌(Escherichia)噬菌体Mu Gam蛋白，任选地所述Gam蛋白包含SEQ ID NO:147的氨基酸序列。
86.前述权利要求中任一项的方法，其中所述II型或V型CRISPR-Cas效应蛋白由多核苷酸编码，和/或所述逆转录酶由多核苷酸编码。
87.权利要求32至86中任一项的方法，其中所述CRISPR-Cas效应蛋白由多核苷酸编码。
88.权利要求37-87中任一项的方法，其中所述5'翼片核酸内切酶(FEN)由多核苷酸编码。
89.权利要求69-88中任一项的方法，其中所述5'-3'核酸外切酶由多核苷酸编码。
90.权利要求1-89中任一项的方法，其中所述靶核酸在细胞中、在无细胞系统中或在病毒中。
91.权利要求90的方法，其中所述细胞为真核细胞或原核细胞。
92.权利要求90或权利要求91的方法，其中所述细胞来自动物、植物、真菌、古细菌、细菌或病毒。
93.权利要求86至92中任一项的方法，其中编码所述II型或V型CRISPR-Cas效应蛋白的多核苷酸和/或编码所述逆转录酶的多核苷酸被包含在相同或分开的表达盒中，任选地当编码所述II型或V型CRISPR-Cas效应蛋白的多核苷酸和编码所述逆转录酶的多核苷酸被包含在相同的表达盒中时，编码所述II型或V型CRISPR-Cas效应蛋白的多核苷酸和编码所述逆转录酶的多核苷酸可操作地连接至单一启动子或者以任何组合可操作地连接至两个或更多个分开的启动子。
94.权利要求87至93中任一项的方法，其中编码所述CRISPR-Cas效应蛋白的多核苷酸被包含在表达盒中，任选地其中所述CRISPR-Cas效应蛋白可操作地连接至启动子。
95.权利要求94的方法，其中所述相同或分开的表达盒被包含在相同或分开的载体中。
96.权利要求94的方法，其中所述表达盒被包含在载体中。
97.权利要求88至96中任一项的方法，其中编码所述5'翼片核酸内切酶的多核苷酸被包含在表达盒中，任选地其中所述表达盒被包含在载体中。
98.权利要求83至91中任一项的方法，其中编码所述5'-3'核酸外切酶的多核苷酸被包含在表达盒中，任选地其中所述表达盒被包含在载体中。
99.前述权利要求中任一项的方法，其中所述延伸型指导核酸被包含在表达盒中，任选地其中所述表达盒被包含在载体中。
100.权利要求32至99中任一项的方法，其中所述指导核酸被包含在表达盒中，任选地其中所述表达盒被包含在载体中。
101.权利要求85至100中任一项的方法，其中对编码所述II型或V型CRISPR-Cas效应蛋白、所述CRISPR-Cas效应蛋白、所述逆转录酶、所述5'翼片核酸内切酶、所述5'-3'核酸外切酶的多核苷酸以及包含其的表达盒和/或载体进行密码子优化以用于在生物中，任选地在原核生物或真核生物中，任选地在动物、植物、真菌、古细菌、细菌或病毒中进行表达。
102.权利要求101的方法，其中所述密码子优化是用于在双子叶植物中进行表达。
103.权利要求101的方法，其中所述密码子优化是用于在单子叶植物中进行表达。
104.修饰靶核酸的方法，所述方法包括：
使所述靶核酸在第一位点处与下列组分相接触：
(a)(i)第一CRISPR-Cas效应蛋白；和
(ii)第一延伸型指导核酸(例如，延伸型CRISPR RNA、延伸型CRISPR DNA、延伸型crRNA、延伸型crDNA)；和
(b)(i)第二CRISPR-Cas效应蛋白；
(ii)第一逆转录酶；和
(ii)第一指导核酸，
由此修饰所述靶核酸。
105.权利要求104的方法，其中所述第一延伸型指导核酸包含：
(a)CRISPR核酸(CRISPR RNA、CRISPR DNA、crRNA、crDNA)；和
(b)包含引物结合位点和逆转录酶模板(RT模板)的延伸部分。
106.权利要求104或权利要求105的方法，其中所述第一延伸型指导核酸的CRISPR核酸包含能够与在所述靶核酸的第一链上的第一位点相结合(与之具有实质上的同源性)的间隔子序列。
107.权利要求104-106中任一项的方法，其中所述第一指导核酸包含CRISPR核酸(CRISPR RNA、CRISPR DNA、crRNA、crDNA)。
108.权利要求107的方法，其中所述第一指导核酸的CRISPR核酸包含与处于在所述靶核酸的第一链上的第一位点上游的在所述靶核酸的第一链上的第二位点相结合的间隔子序列。
109.权利要求104-108中任一项的方法，其中所述第二CRISPR-Cas效应蛋白为包含融合至所述逆转录酶的CRISPR-Cas效应蛋白结构域的CRISPR-Cas融合蛋白。
110.权利要求104-108中任一项的方法，其中所述第二CRISPR-Cas效应蛋白为包含融合至肽标签的CRISPR-Cas效应蛋白结构域的CRISPR-Cas融合蛋白，并且所述第一逆转录酶为包含融合至能够结合所述肽标签的亲和多肽的逆转录酶结构域的逆转录酶融合蛋白。
111.权利要求104-108中任一项的方法，其中所述第一指导核酸连接至RNA募集基序，并且所述第一逆转录酶为包含融合至能够结合所述RNA募集基序的亲和多肽的逆转录酶结构域的逆转录酶融合蛋白。
112.权利要求104-111中任一项的方法，其进一步包括使所述靶核酸与下列组分相接触：
(a)第三CRISPR-Cas效应蛋白；和
(b)第二指导核酸，
其中所述第三CRISPR-Cas效应蛋白对于位于离已经由所述第二CRISPR-Cas效应蛋白产生切口的在第二链上的第二位点大约10至大约125个碱基对处(5'或3')的在所述靶核酸的第一链上的位点产生切口，由此改善错配修复。
113.权利要求104-112中任一项的方法，其进一步包括使所述靶核酸与5'-3'核酸外切酶相接触。
114.权利要求113的方法，其中所述5'-3'核酸外切酶融合至所述第一CRISPR-Cas效应蛋白。
115.权利要求114的方法，其中所述5'-3'核酸外切酶融合至所述第一CRISPR-Cas效应蛋白的C-末端。
116.权利要求113的方法，其中所述5'-3'核酸外切酶为包含融合至肽标签的5'-3'核酸外切酶的融合蛋白，并且所述第一CRISPR-Cas效应蛋白为包含融合至能够与所述肽标签相结合的亲和多肽的CRISPR-Cas效应蛋白结构域的融合蛋白。
117.权利要求113的方法，其中所述5'-3'核酸外切酶为包含融合至能够与所述肽标签相结合的亲和多肽的5'-3'核酸外切酶的融合蛋白，并且所述第一CRISPR-Cas效应蛋白为包含融合至肽标签的CRISPR-Cas效应蛋白结构域的融合蛋白。
118.权利要求113的方法，其中所述5'-3'核酸外切酶为包含融合至能够结合RNA募集基序的亲和多肽的5'-3'核酸外切酶的融合蛋白，并且所述延伸型指导核酸连接至RNA募集基序。
119.权利要求113-118中任一项的方法，其中所述5'-3'核酸外切酶为RecE核酸外切酶(RecE)、RecJ核酸外切酶(RecJ)、T5核酸外切酶(T5_Exo)、T7核酸外切酶(T7_Exo)、λ核酸外切酶(λ_exo)、sbcB(脱氧核糖核酸外切酶)或hExo1(人核酸外切酶1)。
120.权利要求104-119中任一项的方法，其进一步包括使所述靶核酸与下列组分相接触：
(a)第四CRISPR-Cas效应蛋白；
(b)第二逆转录酶；和
(c)第二延伸型指导核酸(例如，延伸型CRISPR RNA、延伸型CRISPR DNA、延伸型crRNA、延伸型crDNA)，
其中所述第二延伸型指导核酸靶向在所述靶核酸的第一链上的位点(间隔子与之实质上互补/与之相结合)，由此修饰所述靶核酸。
121.权利要求104-120中任一项的方法，其中所述第一CRISPR-Cas效应蛋白、所述第二CRISPR-Cas效应蛋白、所述第三CRISPR-Cas效应蛋白和/或所述第四CRISPR-Cas效应蛋白以任何组合地为I型、II型、III型、IV型或V型CRISPR-Cas效应蛋白。
122.权利要求1至103中任一项的方法，其进一步包括使所述靶核酸与下列组分相接触：
(a)第二CRISPR-Cas效应蛋白，任选地II型或V型CRISPR-Cas效应蛋白；
(b)第二逆转录酶；和
(c)第二延伸型指导核酸(例如，延伸型CRISPR RNA、延伸型CRISPR DNA、延伸型crRNA、延伸型crDNA)，
其中所述第二延伸型指导核酸靶向在所述靶核酸的第一链(靶标链)或第二链(非靶标链)上的位点(间隔子与之实质上互补/与之相结合)，由此修饰所述靶核酸。
123.权利要求104-122的方法，其中所述第一和/或第二延伸型指导核酸进一步包含假结。
124.权利要求123的方法，其中所述假结位于所述延伸型指导核酸的3'末端处。
125.权利要求123或权利要求124的方法，其中所述假结为包含SEQ ID NO:158的核酸序列的tEvoPreQ1假结或包含SEQ ID NO:191的核酸序列的EvoPreQ1假结。
126.权利要求123或权利要求124的方法，其中所述假结包含SEQ ID NO:95或SEQ IDNO:203的核酸序列。
127.权利要求104-126中任一项的方法，其进一步包括使所述靶核酸与单链DNA结合蛋白(ssDNA结合蛋白)相接触。
128.权利要求127的方法，其中所述ssDNA结合蛋白融合至所述第一、第二、第三和/或第四CRISPR-Cas效应蛋白。
129.权利要求127或权利要求128的方法，其中所述ssDNA结合蛋白融合至所述第一、第二、第三和/或第四CRISPR-Cas效应蛋白的C-末端。
130.权利要求127或权利要求128的方法，其中所述ssDNA结合蛋白融合至所述第一、第二、第三和/或第四CRISPR-Cas效应蛋白的N-末端。
131.权利要求127-130中任一项的方法，其中所述ssDNA结合蛋白来自人、细菌或噬菌体。
132.权利要求127-131中任一项的方法，其中所述ssDNA结合蛋白为hRad51(任选地，hRad51_S208E_A209D)、hRad52、BsRecA、EcRecA、T4ssB和/或Brex27。
133.权利要求104-132中任一项的方法，其进一步包括通过引入非同源末端连接(NHEJ)的化学抑制剂，通过引入靶向NHEJ蛋白的CRISPR指导核酸或siRNA以瞬时敲低所述NHEJ蛋白的表达，或者通过引入阻止NHEJ的多肽来减少双链断裂。
134.权利要求133的方法，其中所述阻止NHEJ的多肽融合至所述第一和/或第二逆转录酶和/或所述第一、第二、第三和/或第四CRISPR-Cas效应蛋白，任选地所述阻止NHEJ的多肽融合至所述第一和/或第二逆转录酶和/或所述第一、第二、第三和/或第四CRISPR-Cas效应蛋白的N-末端。
135.权利要求133或权利要求134的方法，其中所述阻止NHEJ的多肽为Gam蛋白。
136.权利要求135的方法，其中所述Gam蛋白为埃希氏菌噬菌体Mu Gam蛋白，任选地所述Gam蛋白包含SEQ ID NO:147的氨基酸序列。
137.延伸型指导核酸，其包含(i)V型CRISPR核酸或II型CRISPR核酸(II型或V型CRISPRRNA、II型或V型CRISPR DNA、II型或V型crRNA、II型或V型crDNA)和/或V型CRISPR核酸或II型CRISPR核酸和tracr核酸(例如，II型或V型tracrRNA、II型或V型tracrDNA)；和(ii)包含引物结合位点和逆转录酶模板(RT模板)(RTT)的延伸部分。
138.权利要求137的延伸型指导核酸，其中所述延伸型指导核酸进一步包含假结。
139.权利要求138的延伸型指导核酸，其中所述假结位于所述延伸型指导核酸的3'末端处。
140.权利要求138或权利要求139的延伸型指导核酸，其中所述假结为包含SEQ ID NO:158的核酸序列的tEvoPreQ1假结或包含SEQ ID NO:191的核酸序列的EvoPreQ1假结。
141.权利要求138或权利要求139的延伸型指导核酸，其中所述假结包含SEQ ID NO:95或SEQ ID NO:203的核酸序列。
142.权利要求137-141中任一项的延伸型指导核酸，其被包含在表达盒中，任选地其中所述表达盒被包含在载体中。
143.复合物，其包含：
(a)V型CRISPR-Cas效应蛋白或II型CRISPR-Cas效应蛋白；
(b)逆转录酶；和
(c)延伸型指导核酸(例如，延伸型CRISPR RNA、延伸型CRISPR DNA、延伸型crRNA、延伸型crDNA)。
144.权利要求143的复合物，其中所述延伸型指导核酸进一步包含假结。
145.权利要求144的复合物，其中所述假结位于所述延伸型指导核酸的3'末端处。
146.权利要求144或权利要求145的复合物，其中所述假结为包含SEQ ID NO:158的核酸序列的tEvoPreQ1假结或包含SEQ ID NO:191的核酸序列的EvoPreQ1假结。
147.权利要求144或权利要求145的复合物，其中所述假结包含SEQ ID NO:95或SEQ IDNO:203的核酸序列。
148.权利要求143-147的复合物，其中所述II型或V型CRISPR-Cas效应蛋白为包含融合至肽标签的II型或V型CRISPR-Cas效应蛋白结构域的融合蛋白。
149.权利要求143-147的复合物，其中所述II型或V型CRISPR-Cas效应蛋白为包含融合至能够结合肽标签的亲和多肽的V型CRISPR-Cas效应蛋白结构域的融合蛋白。
150.权利要求143-147的复合物，其中所述II型或V型CRISPR-Cas效应蛋白为包含融合至能够结合RNA募集基序的亲和多肽的II型或V型CRISPR-Cas效应蛋白结构域的融合蛋白。
151.权利要求143-150中任一项的复合物，其中所述逆转录酶为包含融合至肽标签的逆转录酶结构域的融合蛋白。
152.权利要求143-150中任一项的复合物，其中所述逆转录酶为包含融合至能够结合肽标签的亲和多肽的逆转录酶结构域的融合蛋白。
153.权利要求143-150中任一项的复合物，其中所述逆转录酶为包含融合至能够结合RNA募集多肽的亲和多肽的逆转录酶结构域的融合蛋白。
154.权利要求143-153中任一项的复合物，其被包含在表达盒中，任选地其中所述表达盒被包含在载体中。
155.经密码子优化以用于在生物中进行表达的表达盒，其以5'至3'包含：
(a)编码启动子序列的多核苷酸；
(b)编码V型CRISPR-Cas核酸酶(例如，Cpf1(Cas12a)、dCas12a等)或II型CRISPR-Cas核酸酶(例如，Cas9、dCas9等)的多核苷酸，其进行了密码子优化以用于在所述生物中进行表达；
(c)连接体序列；和
(d)编码逆转录酶的多核苷酸，其进行了密码子优化以用于在所述生物中进行表达。
156.权利要求155的表达盒，其中所述生物为动物、植物、真菌、古细菌、细菌或病毒。
157.权利要求156的表达盒，其中所述生物为植物，所述编码V型CRISPR-Cas核酸酶的多核苷酸进行了密码子优化以用于在植物中进行表达，并且所述启动子序列为植物特异性启动子序列(例如，ZmUbi1、MtUb2、RNA聚合酶II(PolII))。
158.权利要求155-157中任一项的表达盒，其中所述逆转录酶融合至一个或多个ssRNA结合结构域(RBD)。
159.权利要求158的方法，其中所述一个或多个单链RNA结合结构域融合至所述逆转录酶的N-末端，任选地其中所述逆转录酶进一步在其C-末端处融合至所述II型CRISPR-Cas效应蛋白和/或V型CRISPR-Cas效应蛋白的N-末端。
160.权利要求155至159中任一项的表达盒，其中所述连接体为氨基酸或肽连接体。
161.权利要求160的表达盒，其中所述肽连接体的长度为大约2至大约100个氨基酸(残基)。
162.权利要求160或权利要求161的表达盒，其中所述肽连接体为GS连接体。
163.经密码子优化以用于在植物中进行表达的表达盒，其包含：
(a)编码植物特异性启动子序列(例如，ZmUbi1、MtUb2)的多核苷酸，和
(b)延伸型指导核酸，其中所述延伸型指导核酸包含延伸部分，所述延伸部分在其3'末端处包含引物结合位点和待被掺入到所述靶核酸中的编辑(例如，逆转录酶模板)，任选地其中所述延伸型指导核酸被包含在表达盒中，任选地其中所述延伸型指导核酸可操作地连接至PolII启动子。
164.权利要求163的表达盒，其中所述延伸型指导核酸进一步包含假结。
165.权利要求164的表达盒，其中所述假结位于所述延伸型指导核酸的3'末端处。
166.权利要求164或权利要求165的表达盒，其中所述假结为包含SEQ ID NO:158的核酸序列的tEvoPreQ1假结或包含SEQ ID NO:191的核酸序列的EvoPreQ1假结。
167.权利要求164或权利要求165的表达盒，其中所述假结包含SEQ ID NO:95或SEQ IDNO:203的核酸序列。
168.权利要求157-167中任一项的表达盒，其中所述植物特异性启动子与内含子相关联或者为包含内含子的启动子区(例如，包含内含子的ZmUbi1；包含内含子的MtUb2)。
169.权利要求156-168中任一项的表达盒，其中所述密码子优化是用于在双子叶植物中进行表达。
170.权利要求156-168中任一项的表达盒，其中所述密码子优化是用于在单子叶植物中进行表达。
171.在植物或植物细胞中修饰靶核酸的方法，其包括将权利要求155-170中任一项的表达盒引入到所述植物或植物细胞中，由此在所述植物或植物细胞中修饰所述靶核酸。
172.权利要求171的方法，其进一步包括使包含所述经修饰的靶核酸的植物细胞进行再生，以产生包含所述经修饰的靶核酸的植物。
173.权利要求1至132、171或172中任一项的方法，其中所述接触在大约20℃至42℃的温度下进行。
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* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2021092130A1 (en) 2019-11-05 2021-05-14 Pairwise Plants Services, Inc. Compositions and methods for rna-encoded dna-replacement of alleles
US20220403475A1 (en) 2021-06-14 2022-12-22 Pairwise Plants Services, Inc. Reporter constructs, compositions comprising the same, and methods of use thereof
US20230266293A1 (en) 2021-09-21 2023-08-24 Pairwise Plants Services, Inc. Color-based and/or visual methods for identifying the presence of a transgene and compositions and constructs relating to the same
WO2023114992A1 (en) * 2021-12-17 2023-06-22 Massachusetts Institute Of Technology Programmable insertion approaches via reverse transcriptase recruitment

Family Cites Families (20)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
NZ221259A (en) 1986-07-31 1990-05-28 Calgene Inc Seed specific transcriptional regulation
US6040504A (en) 1987-11-18 2000-03-21 Novartis Finance Corporation Cotton promoter
CA1339684C (en) 1988-05-17 1998-02-24 Peter H. Quail Plant ubquitin promoter system
US5641876A (en) 1990-01-05 1997-06-24 Cornell Research Foundation, Inc. Rice actin gene and promoter
ATE225853T1 (de) 1990-04-12 2002-10-15 Syngenta Participations Ag Gewebe-spezifische promotoren
US5459252A (en) 1991-01-31 1995-10-17 North Carolina State University Root specific gene promoter
CA2116449C (en) 1991-08-27 2005-04-05 Vaughan Alan Hilder Proteins with insecticidal properties against homopteran insects and their use in plant protection
UA48104C2 (uk) 1991-10-04 2002-08-15 Новартіс Аг Фрагмент днк, який містить послідовність,що кодує інсектицидний протеїн, оптимізовану для кукурудзи,фрагмент днк, який забезпечує направлену бажану для серцевини стебла експресію зв'язаного з нею структурного гена в рослині, фрагмент днк, який забезпечує специфічну для пилку експресію зв`язаного з нею структурного гена в рослині, рекомбінантна молекула днк, спосіб одержання оптимізованої для кукурудзи кодуючої послідовності інсектицидного протеїну, спосіб захисту рослин кукурудзи щонайменше від однієї комахи-шкідника
CA2319079A1 (en) 1998-02-20 1999-08-26 Zeneca Limited Pollen specific promoter
IL151886A (en) 2000-03-27 2010-11-30 Syngenta Participations Ag Promoters of a virus that causes curling of a yellow leaf of a cyst
AR044724A1 (es) 2000-03-27 2005-10-05 Syngenta Participations Ag Promotores del virus de la rizadura amarilla del cestrum
WO2005037993A2 (en) 2003-10-06 2005-04-28 Syngenta Participations Ag Promoters functional in plant plastids
KR100537955B1 (ko) 2003-10-29 2005-12-20 학교법인고려중앙학원 꽃가루 특이적 유전자 발현 프로모터
JP6349261B2 (ja) 2012-02-01 2018-06-27 ダウ アグロサイエンシィズ エルエルシー 合成アブラナ属由来葉緑体輸送ペプチド
PT3177644T (pt) 2014-08-05 2021-01-13 MabQuest SA Reagentes imunológicos que se ligam a pd-1
US9790490B2 (en) 2015-06-18 2017-10-17 The Broad Institute Inc. CRISPR enzymes and systems
WO2018022634A1 (en) * 2016-07-26 2018-02-01 The General Hospital Corporation Variants of crispr from prevotella and francisella 1 (cpf1)
US11268092B2 (en) * 2018-01-12 2022-03-08 GenEdit, Inc. Structure-engineered guide RNA
MX2021011426A (es) * 2019-03-19 2022-03-11 Broad Inst Inc Metodos y composiciones para editar secuencias de nucleótidos.
WO2021092130A1 (en) * 2019-11-05 2021-05-14 Pairwise Plants Services, Inc. Compositions and methods for rna-encoded dna-replacement of alleles

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