CN114835821A - 一种高效特异实现碱基颠换的编辑系统、方法及用途 - Google Patents

Abstract

本发明属于生物技术领域，涉及一种高效特异实现碱基颠换的编辑系统、方法及用途。本发明所述融合蛋白自N端至C端依次包括第一Cas9切口酶片段、嵌合插入片段、第二Cas9切口酶片段，所述嵌合插入片段包含脱氨酶片段和尿嘧啶DNA结合蛋白片段。本发明提供的融合蛋白结合相应的引导RNA，可以对靶位点中的C颠换成G。本发明编辑系统或方法可以在多种细胞中高效实现C‑G碱基的颠换，为修复致病突变，研究基因功能以及改善细胞功能等提供了有效的工具，具备良好的应用前景。

Description

一种高效特异实现碱基颠换的编辑系统、方法及用途

技术领域

本发明属于生物技术领域，特别是涉及一种高效特异实现碱基颠换的编辑系统、方法及用途。

背景技术

随着基因组测序技术的普及，改造基因组是后基因组时代的重要内容。以类转录激活因子效应物核酸酶(TALEN)、锌指核酸酶(ZFN)为代表的基因编辑技术使得改造基因序列变得简单，CRISPR/Cas技术的出现更是极大地加速了这一过程。以CRISPR/Cas9为代表的基因编辑技术，由于具备简单、易操作、高效、普适性等特点迅速成为分子生物学领域的常用技术。基因编辑技术的出现加速了基因治疗以及细胞治疗的进程。

CRISPR/Cas9改造DNA的过程是通过引入双链断裂，利用细胞内的修复机制，进行非同源末端链接(NHEJ)的修复和以同源模板的同源重组修复(HDR)。而目前多种研究表明双链断裂会富集p53突变的细胞，进而有癌变的可能。以双链断裂为基础的DNA改造在临床的应用方面面临着巨大的挑战。

在CRISPR/Cas基础上通过添加不同的功能元件，目前可以实现了包括基因表达激活、抑制、表观调控、碱基编辑等操作，极大地丰富了基因编辑工具库。此种类型的基因编辑或者基因调控方法不需要利用双链断裂，即可实现改变细胞功能的目的。特别是碱基编辑技术，其不同于以双链断裂为基础的同源重组技术，其诱导点突变的效率得到显著提高。研究表明在胚胎中碱基编辑技术甚至可以实现100％的编辑。目前的碱基编辑技术主要是C转换为T的胞嘧啶碱基编辑技术(CBE)和A转换为G的腺嘌呤碱基编辑技术(ABE)。碱基编辑技术因为引入了脱氨酶，存在脱靶的风险。研究表明胞嘧啶脱氨酶可以在DNA和RNA水平均引起脱靶，这种脱靶是不依赖sgRNA的。脱氨酶随机的结合到单链DNA或RNA上是这种脱靶发生的原因。有效控制脱氨酶与单链DNA或者RNA的结合是降低脱靶的有效手段。已报道的手段有对脱氨酶进行突变，调控脱氨酶的表达等。

目前两种碱基编辑方式都是在嘧啶与嘧啶或者嘌呤与嘌呤直接的转换，而嘌呤与嘧啶直接的颠换目前还没有一种高效、特异的方法。胞嘧啶脱氨以后形成尿嘧啶，以尿嘧啶为模板进行修复的时候，尿嘧啶将变成胸腺嘧啶。如果胞嘧啶颠换成鸟嘌呤，需要将尿嘧啶切除，形成一个无碱基的位点(AP位点)。最后DNA修复时在这个AP位点插入鸟嘌呤，也可能插入腺嘌呤或者胸腺嘧啶。对于尿嘧啶的切除效果将显著影响修复的结果。

胞嘧啶碱基编辑器是实现C-T的转换，其加入了UGI抑制尿嘧啶糖基化酶(UNG)的作用。目前报道的碱基颠换工具主要是在胞嘧啶碱基编辑器的基础上去掉了UGI。已有报道指出添加DNA糖基化酶有助于提高C-G比例。来自UNG的同源物UdgX蛋白展示出了较好的编辑效果。目前报道的UdgX来自耻垢分枝杆菌(Mycobacterium smegmatis)，但其作用效果仍然存在位点选择性等问题。

基于此，开发更加高效、特异的碱基颠换的方法有助于实现更多种类的碱基替换，进而对基因与细胞治疗的进展起到促进作用。

发明内容

鉴于上述现有技术的缺点，本发明的目的在于提供一种高效、特异进行碱基颠换的碱基编辑蛋白、编辑系统、方法及其用途。

本发明的目的之一是提供一种融合蛋白，所述融合蛋白自N端至C端依次包括第一Cas9切口酶片段、嵌合插入片段、第二Cas9切口酶片段，所述嵌合插入片段包含脱氨酶片段和尿嘧啶DNA结合蛋白片段。

本发明的另一目的是提供一种分离的多核苷酸，其编码如上所述的融合蛋白。

本发明的另一目的是提供一种构建体，所述构建体含有如上所述的分离的多核苷酸。

本发明的另一目的是提供一种表达系统，所述表达系统含有如上所述的构建体或基因组中整合有外源的如上所述的多核苷酸。

本发明的另一目的是提供一种碱基编辑系统，包括如上所述的融合蛋白；优选地，所述碱基编辑系统还包括sgRNA。

本发明的另一目的是提供如上所述的融合蛋白、分离的多核苷酸、构建体、表达系统、碱基编辑系统在基因编辑中的用途。

本发明的另一目的是提供一种基因编辑方法，包括：通过如上所述的融合蛋白、分离的多核苷酸、构建体、表达系统或碱基编辑系统对靶标序列进行基因编辑。

本发明的另一目的是提供一种报告系统，所述报告系统包含如SEQ ID NO.31所示的核苷酸序列。当SEQ ID NO.31所示的核苷酸序列突变为SEQ ID NO.30时，所述报告系统呈现蓝色荧光；当SEQ ID NO.31所示的核苷酸序列突变为SEQ ID NO.32时，所述报告系统呈现绿色荧光。当SEQ ID NO.31所示的核苷酸序列突变为SEQ ID NO.33时，所述报告系统无荧光变化。在一优选实施方式中，所述报告系统包含质粒，所述质粒包含如SEQ ID NO.31所示的核苷酸序列，所述质粒的核苷酸序列如SEQ ID NO.2所示。

本发明的另一目的是提供一种新型的尿嘧啶结合蛋白，其作用是结合尿嘧啶，促进DNA的修复；所述尿嘧啶DNA结合蛋白选自来自微生物的Udgx家族的酶或其变体。

本发明的另一目的是提供如上所述的尿嘧啶DNA结合蛋白在用于构建碱基编辑蛋白或碱基编辑系统中的用途；所述碱基编辑蛋白或碱基编辑系统用于实现C至G或G至C的颠换。

附图说明

图1示出了荧光报告系统模式图(下方图中，横坐标从左至右依次为10⁰、10¹、10²、10³、10⁴、10⁵，纵坐标从下至上依次为10⁰、10¹、10²、10³、10⁴、10⁵)。

图2示出了三种碱基编辑蛋白结构示意图。

图3示出了三种碱基编辑蛋白对荧光比例的影响。

图4示出了不同来源的UdgX蛋白进化树分析。

图5示出了不同来源UdgX编辑蛋白对荧光比例的影响。

图6示出了利用转座酶将APOBEC-UdgX随机插入到Cas9蛋白中间的模式图。

图7示出了不同插入位点对报告系统编辑效率的影响。

图8示出了CE-CGBE-ancApobec1与CE-CGBE-A3A编辑蛋白模式图。

图9示出了C-G编辑蛋白对内源基因的编辑一代测序峰图。(A)展示了CE-CGBE-A3A的编辑效果，(B)展示了CE-CGBE-ancApobec1的编辑效果。

图10示出了CE-CGBE-ancApobec1与CE-CGBE-A3A对内源基因编辑效率分析。

图11示出了CE-CGBE-A3A在RNA与DNA水平脱靶分析；A为DNA水平脱靶情况，B为RNA水平脱靶情况。

具体实施方式

下面将结合实施例对本发明的实施方案进行清楚、完整的描述，显然，所描述的实施例仅用于说明本发明的一部分实施例，而不应视为限制本发明的范围。实施例中未注明具体条件者，按照常规条件或制造商建议的条件进行。所用试剂或仪器未注明生产厂商者，视为可以通过市售购买获得的常规产品。

本发明发明人经过大量研究探索，发现在nCas9蛋白内部的合适位置嵌合脱氨酶和尿嘧啶结合蛋白，能够构建适宜CG碱基颠换的碱基编辑器或碱基编辑系统，并且极大地降低在RNA和DNA上的脱靶情况，在此基础上完成了本发明。

本发明一方面提供了一种融合蛋白，所述融合蛋白自N端至C端依次包括第一Cas9切口酶片段、嵌合插入片段、第二Cas9切口酶片段，所述嵌合插入片段包含脱氨酶片段和尿嘧啶DNA结合蛋白片段。

本发明所述融合蛋白中，所述第一Cas9切口酶片段的氨基酸序列包括以下片段或其变体：如SEQ ID NO.34所示的片段(对应Cas9切口酶nCas9的第1位至第1003位氨基酸片段)；或如SEQ ID NO.35所示的片段(对应Cas9切口酶nCas9的第1位至第1027位氨基酸片段)；或，如SEQ ID NO.36所示的片段(对应Cas9切口酶nCas9的第1位至第1039位氨基酸片段)。

本发明所述融合蛋白中，所述第二Cas9切口酶片段的氨基酸序列包括以下片段或其变体：如SEQ ID NO.37所示的片段(对应Cas9切口酶nCas9的第1004位至C端氨基酸片段)；或如SEQ ID NO.38所示的片段(对应Cas9切口酶nCas9的第1028位至C端氨基酸片段)；或如SEQ ID NO.39所示的片段(对应Cas9切口酶nCas9的第1040位至C端氨基酸片段)。

本发明所述融合蛋白中，当所述Cas9切口酶的N端是从起始密码子至第1003位氨基酸的片段或其变体，相应的Cas9切口酶的C端是从第1004位氨基酸至Cas9切口酶的C末端的片段或其变体；当所述Cas9切口酶的N端是从起始密码子至第1027位氨基酸的片段或其变体，相应的Cas9切口酶的C端是从第1028位氨基酸至Cas9切口酶的C末端的片段或其变体；当所述Cas9切口酶的N端是从起始密码子至第1039位氨基酸的片段或其变体，相应的Cas9切口酶的C端是从第1040位氨基酸至Cas9切口酶的C末端的片段或其变体。所述变体是指与原始序列相比具有80％以上序列相似性、且具有原始序列的功能的氨基酸片段，

本发明所述融合蛋白中，所述胞嘧啶脱氨酶的作用是对靶位点上的C脱氨形成U；所述脱氨酶选自胞嘧啶脱氨酶或其变体，所述胞嘧啶脱氨酶选自：1)APOBEC1、APOBEC2、APOBEC3A、APOBEC3B、APOBEC3C、APOBEC3D、APOBEC3F、APOBEC3G、APOBEC3H、APOBEC4、激活诱导的脱氨酶(AID)和pmCDA1；APOBEC家族；优选地，所述胞嘧啶脱氨酶选自ancAPOBEC1、Apobec3A。

本发明所述融合蛋白中，所述尿嘧啶DNA结合蛋白选自来自微生物的Udgx家族的酶或其变体；优选地，所述微生物选自类胞内分枝杆菌(Mycobacteriumparaintracellulare,NCBI Reference Sequence:WP_014385600.1)、少动鞘氨醇单胞菌(Sphingomonas paucimobilis,NCBI Reference Sequence:WP_007405629.1)、球形红假单胞菌(Cereibacter sphaeroides,NCBI Reference Sequence:WP_012643796.1)、柑桔溃疡病菌(Xanthomonas citri,NCBI Reference Sequence:WP_011052458.1)、苜蓿中华根瘤菌(Sinorhizobium meliloti,NCBI Reference Sequence:WP_010976097.1)、皮诺卡氏菌(Nocardia farcinica,NCBI Reference Sequence:WP_011209261.1)、盖尔森基兴奴卡氏菌(Nocardia cyriacigeorgica,NCBI Reference Sequence:WP_014350460.1)、哥伦布分枝杆菌(Mycobacterium colombiense,NCBI Reference Sequence:WP_007772952.1)、贪铜菌(Cupriavidus taiwanensis,NCBI Reference Sequence:WP_012351902.1)、阿氟曼链霉菌(Streptomyces scabiei,NCBI Reference Sequence:WP_012998863.1)、欺诈伯克霍尔德菌(Burkholderia dolosa,NCBI Reference Sequence:WP_006767162.1)、梅洛尼鞘氨醇单胞菌(Sphingomonas melonis,NCBI Reference Sequence:WP_017980260.1)、沼泽红假单胞菌(Rhodopseudomonas palustris,NCBI Reference Sequence:WP_011157305.1)、耻垢分枝杆菌(Mycolicibacterium smegmatis,NCBI Reference Sequence:WP_011726794.1)、胞内分枝杆菌(Mycobacterium intracellulare,NCBI ReferenceSequence:WP_009956825.1)、多噬伯克霍尔德菌(Burkholderia multivorans,NCBIReference Sequence:WP_006411759.1)、慢生型大豆根瘤菌(Bradyrhizobium japonicum,NCBI Reference Sequence:WP_014496800.1)、高效固氮慢生根瘤菌(Bradyrhizobiumdiazoefficiens,NCBI Reference Sequence:WP_011085807.1)、草莓角斑病菌(Xanthomonas fragariae,NCBI Reference Sequence:WP_011085807.1)；进一步优选地，所述微生物为皮诺卡氏菌。

本发明所述融合蛋白中，所述融合蛋白还包括核定位信号片段，所述核定位信号片段可以位于融合蛋白的N端，也可以位于融合蛋白的C端，或是同时位于融合蛋白的N端和C端；当位于N端时，所述核定位信号片段的N端与第一Cas9切口酶的C端连接；当位于C端时，所述核定位信号片段的C端与第一Cas9切口酶的N端连接。

其中，所述核定位信号片段优选为BPNLS或其变体，所述变体与BPNLS具有80％以上序列相似性且具有BPNLS的功能。

本发明所述融合蛋白中，所述融合蛋白还包括柔性连接肽片段。即，在第一Cas9切口酶片段与嵌合插入片段之间，嵌合插入片段与第二Cas9切口酶片段，嵌合插入片段中脱氨酶片段与尿嘧啶DNA结合蛋白片段之间，核定位信号与第一Cas9切口酶片段或第二Cas9切口酶片段之间均可以通过柔性连接肽连接。当所述融合蛋白还有多个柔性连接肽片段时，所述多个柔性连接肽片段可以是相同或不同的。优选的，所述柔性连接肽片段的氨基酸序列包括SEQ ID NO.40：SGSETPGTSESATPESGS、SEQ ID NO.41：SGSGSETPGTSESATPES；

本发明所述融合蛋白中，所述胞嘧啶脱氨酶片段、尿嘧啶DNA结合蛋白嵌合入Cas9切口酶中，所述胞嘧啶脱氨酶片段和尿嘧啶DNA结合蛋白在融合蛋白中的顺序不作限制，可以是胞嘧啶脱氨酶片段与第一Cas9切口酶片段连接，也可以是尿嘧啶DNA结合蛋白与第一Cas9切口酶片段连接。优选地，所述融合蛋白依次包括第一Cas9切口酶片段、脱氨酶片段、尿嘧啶DNA结合蛋白片段、第二Cas9切口酶片段。

在一些实施方式中，本发明所述融合蛋白自N端到C端依次是核定位信号片段、第一Cas9切口酶片段、尿嘧啶DNA结合蛋白、胞嘧啶脱氨酶片段、第二Cas9切口酶片段和核定位信号片段。

在一些实施方式中，本发明所述融合蛋白自N端到C端依次是核定位信号片段、第一Cas9切口酶片段、胞嘧啶脱氨酶片段、尿嘧啶DNA结合蛋白、第二Cas9切口酶片段和核定位信号片段。

在另一些优选实施方式中，所述融合蛋白的氨基酸序列如SEQ ID No.11、SEQ IDNo.12、SEQ ID No.13至少任一所示；其编码多核苷酸分别如SEQ ID NO.42、SEQ ID NO.43、SEQ ID NO.44所示。

本发明的第二方面是提供一种分离的多核苷酸，其编码本发明第一方面所提供的融合蛋白。

本发明的第三方面是提供一种构建体，所述构建体含有本发明第二方面所提供的分离的多核苷酸。所述构建体通常可以通过将所述分离的多核苷酸插入合适的表达载体中构建获得，本领域技术人员可选择合适的表达载体。

本发明的第四方面是提供一种表达系统，所述表达系统含有本发明第三方面所述的构建体或基因组中整合有外源的本发明第二方面所述的分离的多核苷酸。所述表达系统可以是宿主细胞，所述宿主细胞可以表达如上所述的融合蛋白，所述融合蛋白可以与sgRNA相配合，从而可以将所述融合蛋白定位到目标区域，实现目标区域的碱基编辑。所述宿主细胞选自真核细胞或原核细胞；优选地，所述宿主细胞选自小鼠细胞、人细胞，例如是NK细胞、T细胞、脑神经瘤细胞、胚胎肾细胞、宫颈癌细胞、结肠癌细胞、骨肉瘤细胞等。

本发明的第五方面是提供一种碱基编辑系统，其包括如上所述的融合蛋白。优选地，所述碱基编辑系统还包括引导RNA，所述融合蛋白在引导RNA(sgRNA)作用下对靶位点实现C至G或G至C的颠换。本领域技术人员可以根据基因的目标编辑区域，选择合适的靶向特异性位点的sgRNA。例如，所述sgRNA的序列通常可以与目标区域至少部分互补，从而可以与所述融合蛋白相配合，将所述融合蛋白定位到目标区域，实现靶点区域内的碱基编辑，例如，可以是胞嘧啶脱氨基反应，即将胞嘧啶(cytosin，C)脱氨产生胸腺嘧啶(thymine，T)，进而通过Udgx将尿嘧啶切除产生AP位点，并通过基因修复插入鸟嘌呤，实现C至G的编辑(相对应的互补链上实现G至C的编辑)。

本发明所述碱基编辑系统中，所述碱基编辑系统包括：a)所述融合蛋白或其编码多核苷酸，b)引导RNA核苷酸序列或其编码多核苷酸。所述引导RNA将所述融合蛋白靶向靶标序列中的靶C碱基。本发明所述碱基编辑系统用于碱基编辑时，所述靶标序列可以与a)、b)分别接触，或与a)和b)形成的RNP复合物接触。

在一些实施方式中，所述碱基编辑系统包含一个或多个载体；所述一个或多个载体包含(i)第一调控元件，所述第一调控元件可操作地连接至所述融合蛋白的编码多核苷酸；以及(ii)第二调控元件，所述第二调控元件可操作地连接至所述引导RNA核苷酸序列的编码多核苷酸；所述(i)和(ii)位于相同或不同载体上。

在一些实施方式中，所述碱基编辑系统包含(i)融合蛋白，以及(ii)包含所述引导核苷酸序列的编码多核苷酸的载体。所述碱基编辑系统用于碱基编辑时，所述靶标序列可以与i)、ii)分别接触，或与i)和ii)形成的RNP复合物接触。

本发明第六方面是提供如上所述的融合蛋白、分离的多核苷酸、构建体、表达系统或碱基编辑系统在基因编辑中的用途。具体地，所述基因编辑是指碱基颠换；优选地，所述基因编辑实现C至G或G至C的编辑。所述基因编辑用于实现模型构建(例如，疾病模型、细胞模型、动物模型等)、致病位点的修正、基因功能研究、增强细胞功能、细胞治疗等。例如，所述的融合蛋白、分离的多核苷酸、构建体、表达系统或碱基编辑系统用于以下疾病治病位点的修正：自身免疫性疾病、肿瘤、病毒感染性疾病、细菌感染性疾病，进而，实现自身免疫性疾病、肿瘤、病毒感染性疾病、细菌感染性疾病的预防和/或治疗。

本发明第七方面是提供一种碱基编辑方法，包括：通过本发明第一方面所提供的融合蛋白、或本发明第五方面所提供的碱基编辑系统进行基因编辑。例如，所述基因编辑方法可以包括：在适当条件下培养本发明第四方面所提供的表达系统，从而表达所述融合蛋白，所述融合蛋白可以在与其配合的靶向目标区域的sgRNA存在的条件下，对靶标区域进行碱基编辑。提供所述sgRNA存在的条件的方法对于本领域技术人员来说应该是已知的，例如，可以是在适当条件下培养能够表达所述sgRNA的表达系统，所述表达系统可以是包括含有编码所述sgRNA的多核苷酸的表达载体的宿主细胞、或染色体中整合有编码所述sgRNA的多核苷酸的宿主细胞。在本发明一具体实施例中，所述sgRNA与所述融合蛋白可以在同一宿主细胞中表达，所述宿主细胞可以为靶细胞。本发明所述方法适用的基因编辑的对象没有特别的限制，可以在体外或体内实施。在一些实施方式中，所述方法在体外实施；优选地，所述方法在培养的细胞中实施；可以在体细胞或生殖细胞，可以是动物细胞或人细胞；例如所述细胞为自然杀伤细胞即NK细胞；所述NK细胞选自来自外周血细胞的原代NK细胞(优选来自PBMC的原代NK细胞)、脐带血来源NK细胞、胚胎干细胞(ESC)、诱导多能干细胞(ips)或诱导多能干细胞(ips)诱导的NK细胞。在一些实施方式中，所述方法在体内实施；优选地，所述方法在哺乳动物中实施；进一步优选地，所述哺乳动物是啮齿动物；更进一步优选地，所述哺乳动物是人。例如所述方法靶向哺乳动物的NK细胞。

可以使用基因递送载剂递送本文所述的多核苷酸至细胞或组织。本文所用“基因递送”、“基因转移”、“转导”等，其指代将外源性多核苷酸导入宿主细胞，如载体介导的基因转移(通过，例如，病毒感染/转染，或各种其他基于蛋白质或基于脂质的基因递送复合物)以及协助“裸”多核苷酸递送的技术(如电穿孔，“基因枪”递送以及用于导入多核苷酸的各种其他技术)。导入的多核苷酸可以稳定地或瞬时地维持在宿主细胞中。稳定维持通常需要导入的多核苷酸包含与宿主细胞相容的复制起点或者纳入到宿主细胞的复制子，诸如染色体外复制子(例如，质粒)或核或线粒体染色体。许多“载体”已知能够介导基因向哺乳动物细胞的转移，如本领域已知和本文所述。

本发明所述方法中，在一些实施方式中，可以直接将所述融合蛋白或其变体递送至待编辑对象如体外体系或宿主细胞中。

本发明所述方法中，在一些实施方式中，也可以将所述融合蛋白或其变体的编码多核苷酸递送至待编辑对象如体外体系或宿主细胞中，然后翻译成所述融合蛋白或其变体。其中所述融合蛋白或其变体的编码多核苷酸可以是DNA形式或RNA形式；DNA形式包括cDNA、基因组DNA或人工合成的DNA，DNA可以是单链的或是双链的，DNA可以是编码链或非编码链；RNA形式例如是信使RNA(mRNA)。在一些优选实施方式中，所述融合蛋白或其变体的编码多核苷酸可以以表达载体的形式递送，所述表达载体上包含编码一个或多个拷贝的编码融合蛋白或其变体的多核苷酸。

本发明所述方法中，在一些实施方式中，可以直接将所述引导核苷酸序列递送至待编辑对象如体外体系或宿主细胞中。

本发明所述方法中，在一些实施方式中，也可以将所述引导核苷酸序列的编码多核苷酸递送至待编辑对象如体外体系或宿主细胞中，然后翻译成所述引导核苷酸序列。其中所述引导核苷酸序列的编码多核苷酸可以是DNA形式；DNA形式包括cDNA、基因组DNA或人工合成的DNA，DNA可以是单链的或是双链的，DNA可以是编码链或非编码链。在一些优选实施方式中，所述引导核苷酸序列的编码多核苷酸可以以表达载体的形式递送，所述表达载体上包含编码一个或多个拷贝的编码引导核苷酸序列的多核苷酸。

本发明所述方法中，编码所述融合蛋白或其变体的多核苷酸包括：只编码融合蛋白或其变体的编码序列；融合蛋白或其变体的编码序列和各种附加编码序列；融合蛋白或其变体的编码序列(和任选的附加编码序列)以及非编码序列。编码所述引导核苷酸序列的多核苷酸包括：只编码引导核苷酸序列的编码序列；引导核苷酸序列的编码序列和各种附加编码序列；引导核苷酸序列的编码序列(和任选的附加编码序列)以及非编码序列。在一些实施方式中，本发明所述方法中，所述碱基编辑系统可以是包含一个或多个载体；所述一个或多个载体包含(i)第一调控元件，所述第一调控元件可操作地连接至所述融合蛋白或其变体的编码序列；以及(ii)第二调控元件，所述第二调控元件可操作地连接至所述引导核苷酸序列的编码序列；所述(i)和(ii)位于相同或不同载体上。在一些实施方式中，所述碱基编辑系统包含(i)融合蛋白或其变体，以及(ii)包含所述引导核苷酸序列的编码序列的载体。

所述第一调控元件可以调控所述融合蛋白或其变体的编码多核苷酸的转录。所述融合蛋白或其变体的编码多核苷酸可以是一个或多个，所述第一调控元件可以是一个或多个。所述第二调控元件可以调控所述引导核苷酸序列的编码多核苷酸的转录。所述引导核苷酸序列的编码多核苷酸可以是一个或多个，所述第二调控元件可以是一个或多个。

本发明第八方面提供了一种预防和/或病症的方法，所述方法包括向有需要的受试者施用治疗有效量的如上所述方法中所述的融合蛋白、分离的多核苷酸、构建体、表达系统或碱基编辑系统；所述病症选自以下至少任一项：自身免疫性疾病、肿瘤、病毒感染性疾病、细菌感染性疾病等。

本发明中，所述自身免疫性疾病选自系统性红斑狼疮、类风湿关节炎、系统性硬化症、口眼干燥综合征、多发性肌炎中的一种或多种。

本发明中，所述肿瘤选自淋巴瘤、血液瘤或实体瘤；优选地，选自肾上腺皮质癌、膀胱尿路上皮癌、乳腺癌、宫颈鳞状细胞癌、宫颈内腺癌、胆管癌、结肠腺癌、淋巴样肿瘤、弥散性大B细胞淋巴瘤、食管癌、多形性成胶质细胞瘤、头颈部鳞状细胞癌、肾嫌色细胞癌、肾透明细胞癌、肾乳头状细胞癌、急性髓性白血病、脑低度胶质瘤、肝细胞癌、肺腺癌、肺鳞状细胞癌、间皮细胞癌、卵巢癌、胰腺癌、嗜铬细胞瘤和副神经节瘤、前列腺癌、直肠癌、恶性肉瘤、黑色素瘤、胃癌、睾丸生殖细胞肿瘤、甲状腺癌、胸腺癌、子宫内膜癌、子宫肉瘤、葡萄膜黑色素瘤、多发性骨髓瘤、急性淋系白血病、慢性淋系白血病、慢性髓性白血病、T细胞淋巴瘤、B细胞淋巴瘤肿瘤细胞中的一种或多种。

本发明中，所述病毒选自流感病毒、副流感病毒、麻疹病毒、腮腺炎病毒、疱疹病毒、腺病毒、呼吸道合胞病毒、脊髓灰质炎病毒、柯萨奇病毒、或埃可病毒中的一种或几种。

本发明中，所述细菌选自大肠杆菌、干酪乳杆菌、脆弱拟杆菌、鲁氏不动杆菌、具核梭杆菌、乔氏拟杆菌、拟南芥拟杆菌、鼠李糖乳杆菌、马赛拟杆菌、卵形拟杆菌、空肠弯曲菌、腐生葡萄球菌、粪肠球菌、多形拟杆菌、普通拟杆菌、单形拟杆菌、粪副拟杆菌、死亡梭杆菌以及短双歧杆菌中的一种或几种。

本发明中，所述融合蛋白、分离的多核苷酸、构建体、表达系统或碱基编辑系统可以与其他药物或试剂联用。即，所述融合蛋白、分离的多核苷酸、构建体、表达系统或碱基编辑系统可以是单一有效成分，也可以与其他活性组分进行组合，构成联合制剂。所述其他活性组分可以是其他各种可以用于治疗自身免疫性疾病、肿瘤、病毒感染性疾病、细菌感染性疾病的药物。组合物中活性组分的含量通常为安全有效量，所述安全有效量对于本领域技术人员来说应该是可以调整的，例如，所述活性成分的施用量通常依赖于患者的体重、应用的类型、疾病的病情和严重程度，例如，作为活性成分的所述碱基编辑系统或NK细胞的施用量通常可以为1～1000mg/kg/day、20～200mg/kg/day、1～3mg/kg/day、3～5mg/kg/day、5～10mg/kg/day、10～20mg/kg/day、20～30mg/kg/day、30～40mg/kg/day、40～60mg/kg/day、60～80mg/kg/day、80～100mg/kg/day、100～150mg/kg/day、150～200mg/kg/day、200～300mg/kg/day、300～500mg/kg/day、或500～1000mg/kg/day。

本发明所述方法和应用中，当所述活性成分与其他治疗剂联用时，所述活性成分与其他治疗剂共同给予。“共同给予”表示在同一制剂中或在两种不同制剂中经由相同或不同途径同时给予，或通过相同或不同途径顺次给予。“顺次”给予表示在两种或多种不同化合物的给药之间具有以秒、分钟、小时或天计的时间差异。

本发明第九方面是提供一种碱基编辑的细胞，所述细胞包含C至G的突变。在一些优选实施方式中，所述细胞通过如上所述的碱基方法制备得到。

本发明第十方面是提供一种报告系统，用于检测C-G突变效率，所述报告系统包含如SEQ ID NO.31所示的核苷酸序列。如SEQ ID NO.31所示的核苷酸序列编码的蛋白不发荧光，且编码的报告蛋白的第67位氨基酸对应的密码子为GAT。当GAT突变为CAT，即当SEQ IDNO.31所示的核苷酸序列突变为SEQ ID NO.30时，所述报告系统呈现蓝色荧光(SEQ IDNO.1)；当GAT突变为TAT，即当SEQ ID NO.31所示的核苷酸序列突变为SEQ ID NO.32时，所述报告系统呈现绿色荧光。当SEQ ID NO.31所示的核苷酸序列突变为SEQ ID NO.33时，所述报告系统无荧光变化。在一优选实施方式中，包含所述报告系统的质粒的核苷酸序列如SEQ ID NO.2所示。

本发明第十一方面是提供如上所述的报告系统，所述报告系统用于检测如上所述的融合蛋白、分离的多核苷酸、构建体、表达系统或碱基编辑系统的C-G编辑效率。

本发明第十二方面是提供一种尿嘧啶DNA结合蛋白，所述尿嘧啶DNA结合蛋白选自来自微生物的Udgx家族的酶或其变体；优选地，所述微生物选自类胞内分枝杆菌、少动鞘氨醇单胞菌、球形红假单胞菌、柑桔溃疡病菌、苜蓿中华根瘤菌、皮诺卡氏菌、盖尔森基兴奴卡氏菌、哥伦布分枝杆菌、贪铜菌、阿氟曼链霉菌、欺诈伯克霍尔德菌、梅洛尼鞘氨醇单胞菌、沼泽红假单胞菌、耻垢分枝杆菌、胞内分枝杆菌、多噬伯克霍尔德菌、慢生型大豆根瘤菌、高效固氮慢生根瘤菌、草莓角斑病菌；进一步优选地，所述微生物为皮诺卡氏菌。

本发明第十三方面是提供如上所述的尿嘧啶DNA结合蛋白在用于构建碱基编辑蛋白或碱基编辑系统中的用途；所述碱基编辑蛋白或碱基编辑系统用于实现C至G或G至C的颠换。

本发明第十四方面还提供了一种组合物，其包含如上任一项所述的融合蛋白、分离的多核苷酸、构建体、表达系统或碱基编辑系统。在一些优选实施方式，其进一步包含药学上可接受的载体。所述可接受的载体例如无菌水或生理盐水、稳定剂、赋形剂、抗氧化剂(抗坏血酸等)、缓冲剂(磷酸、枸橼酸、其它的有机酸等)、防腐剂、表面活性剂(PEG、Tween等)、螯合剂(EDTA等)、粘合剂等。而且，也可含有其它低分子量的多肽；血清白蛋白、明胶或免疫球蛋白等蛋白质；甘氨酸、谷酰胺、天冬酰胺、精氨酸和赖氨酸等氨基酸；多糖和单糖等糖类或碳水化物；甘露糖醇或山梨糖醇等糖醇。当制备用于注射的水溶液时，例如生理盐水、含有葡萄糖或其它的辅助药物的等渗溶液，如D-山梨糖醇、D-甘露糖、D-甘露糖醇、氯化钠，可并用适当的增溶剂例如醇(乙醇等)、多元醇(丙二醇，PEG等)、非离子表面活性剂(吐温80，HCO-50)等。

本发明还提供了一种试剂盒，其包含如上所述的组合物。

本发明中，所述组合物或药物组合物或联合制剂的剂型选自：注射剂、注射用无菌粉末、片剂、丸剂、胶囊、锭剂、醑剂、散剂、颗粒剂、糖浆剂、溶液剂、酊剂、气雾剂、粉雾剂、或栓剂。本领域技术人员可根据给药方式，选择合适的制剂形式，例如，适合于口服给药的制剂形式可以是包括但不限于丸剂、片剂、咀嚼剂、胶囊剂、颗粒剂、溶液剂、滴剂、糖浆、气雾剂或粉雾剂等。

本发明中，蛋白或其片段(如第一Cas9切口酶片段、第二Cas9切口酶片段、脱氨酶片段、尿嘧啶DNA结合蛋白片段)的变体为原始蛋白或其片段的片段、衍生物和类似物，其可以是(i)有一个或多个保守或非保守性氨基酸残基(优选保守性氨基酸残基)被取代的蛋白，而这样的取代的氨基酸残基可以是也可以不是由遗传密码编码的，或(ii)在一个或多个氨基酸残基中具有取代基团的蛋白，或(iii)附加的氨基酸序列融合到此蛋白序列而形成的蛋白(如前导序列或分泌序列或用来纯化此蛋白的序列或蛋白原序列)。根据本发明的定义，这些片段、衍生物和类似物属于本领域熟练技术人员公知的范围。例如，在一些实施方式中，所述第一Cas9切口酶片段的变体是指与所述融合蛋白的氨基酸序列具有75％或更高，或85％或更高，或90％或更高，或95％或更高同一性，且具有所述第一Cas9切口酶片段相同或相似功能的蛋白。上述75％或75％以上同一性，可为75％、80％、85％、90％或95％以上的同一性；具体地可为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％。上述90％或90％以上同一性，可为91％、92％、93％、94％、95％、96％、97％、98％、99％同一性。所述相似功能是指保留原蛋白的75％或更高，或85％或更高，或90％或更高，或95％或更高的功能。

本发明中，所述碱基编辑系统能够实现C至G或G至C的编辑或突变。例如，当靶向编码链时，所述碱基编辑系统能够实现C至G的编辑，相应地，对于非编码链，实现G至C的编辑；当靶向非编码链时，所述碱基编辑系统能够实现C至G的编辑，相应地，对于编码链，实现G至C的编辑。

除非另行定义，文中所使用的所有专业与科学用语与本领域熟练人员所熟悉的意义相同。此外，任何与所记载内容相似或均等的方法及材料皆可应用于本发明方法中。文中所述的较佳实施方法与材料仅作示范之用。

如本文所用，序列相似性或同一性可以用肉眼或计算机软件进行评价。使用计算机软件，两个或多个序列之间的同一性可以用百分比(％)表示，其可以用来评价相关序列之间的同一性。

如本文所用，“包含”、“含有”等应理解为是包括性的意思，而没有排他性或穷尽的意思；即“包括但不限于”的意思。

如本文所用，“治疗有效量”通常指一用量在经过适当的给药期间后，能够达到治疗如上所列出的疾病的效果。

如本文所用，“治疗性”和“预防性”应理解为其最宽的意义。术语“治疗性”不一定暗示哺乳动物接受治疗直至完全恢复。类似地，“预防性”不一定表示对象最终不会感染疾病病症。因此，治疗和预防包括缓解具体病症的症状或防止或降低具体病症产生的风险。术语“预防”可理解为降低具体病症发作的严重程度。治疗也可降低已有病症的严重程度或急性发作的频率。

如本文所用，进行治疗性或预防性治疗的对象或个体优选哺乳动物，例如但不限于人、灵长类、牲畜(如绵羊、牛、马、驴、猪)、宠物(如狗、猫)、实验室试验动物(如小鼠、家兔、大鼠、豚鼠、仓鼠)或被捕获的野生动物(如狐狸、鹿)。所述对象优选灵长类。所述对象最优选人。

如本文所用，术语“核酸”和“核酸组分”可互换使用，其指具有核碱基和酸性部分的化合物，例如核苷、核苷酸或核苷酸的聚合物。在一些实施方式中，“核酸”是指单个核酸残基(例如，核苷酸和/或核苷)。在一些实施方式中，“核酸”是指包含三个或更多个单个核苷酸残基的寡核苷酸链。本文中所用术语“寡核苷酸”和“多核苷酸”可互换使用以指核苷酸的聚合物(例如，一串至少三个核苷酸)。在一些实施方式中，“核酸”包括RNA以及单链和/或双链DNA。核酸可以是天然产生的或是非天然产生的分子。

如本文所用，术语“表达”指多核苷酸转录成mRNA的过程和/或转录的mRNA随后被翻译成肽、多肽或蛋白质的过程。如果多核苷酸源自基因组DNA，则表达可包括真核细胞中mRNA的剪接。基因的表达水平可以通过测量细胞或组织样品内mRNA或蛋白质的量予以确定。

术语“调节元件”包括启动子、增强子、内部核糖体进入位点(IRES)、和其他表达控制元件(例如转录终止信号，如多聚腺苷酸化信号和多聚U序列)。调节元件包括指导一个核苷酸序列在许多类型的宿主细胞中的组成型表达的那些序列以及指导该核苷酸序列只在某些宿主细胞中表达的那些序列(例如，组织特异型调节序列)。组织特异型启动子可指导在感兴趣的组织表达，例如肌肉、神经元、骨、皮肤等。在一些实施例中，一个载体包含一个或多个pol III启动子、pol II启动子、pol I启动子、或其组合。pol III启动子包括但不限于U6和H1启动子。pol II启动子包括但不限于巨细胞病毒(CMV)启动子、SV40启动子、二氢叶酸还原酶启动子、β-肌动蛋白启动子、磷酸甘油激酶(PGK)启动子等。

如本文所用，术语“蛋白质”、“肽”和“多肽”可以互换使用并以其最广泛的含义使用，是指两个或更多个亚基的氨基酸、氨基酸类似物或肽模拟物的化合物。亚基可以通过肽键连接。在另一方面中，亚基可以通过其他键连接，例如，酯、醚等。蛋白质或肽必须包含至少两个氨基酸，并且对构成蛋白质或肽序列的最大氨基酸数没有限制。已知蛋白质和肽具有C-末端和N-末端，所述C-末端指代在该末端氨基酸上存在未结合羧基的末端，所述N-末端指代在该末端氨基酸上存在未结合氨基的末端。本文所用术语“氨基酸”指天然和/或非天然或合成的氨基酸，包括甘氨酸，以及D和L光学异构体，氨基酸类似物和肽模拟物。术语“融合”在蛋白质或多肽上下文中是指两个或更多个蛋白质或多肽(或其结构域)末端之间形成融合蛋白的连接。

在进一步描述本发明具体实施方式之前，应理解，本发明的保护范围不局限于下述特定的具体实施方案；还应当理解，本发明实施例中使用的术语是为了描述特定的具体实施方案，而不是为了限制本发明的保护范围；在本发明说明书和权利要求书中，除非文中另外明确指出，单数形式“一个”、“一”和“这个”包括复数形式。

当实施例给出数值范围时，应理解，除非本发明另有说明，每个数值范围的两个端点以及两个端点之间任何一个数值均可选用。除非另外定义，本发明中使用的所有技术和科学术语与本技术领域技术人员通常理解的意义相同。除实施例中使用的具体方法、设备、材料外，根据本技术领域的技术人员对现有技术的掌握及本发明的记载，还可以使用与本发明实施例中所述的方法、设备、材料相似或等同的现有技术的任何方法、设备和材料来实现本发明。

除非另外说明，本发明中所公开的实验方法、检测方法、制备方法均采用本技术领域常规的分子生物学、生物化学、染色质结构和分析、分析化学、细胞培养、重组DNA技术及相关领域的常规技术。这些技术在现有文献中已有完善说明。

实施例1：用于测试C-G编辑系统编辑效率的荧光报告系统

在哺乳动物细胞系中，利用碱基颠换编辑器对获得性报告系统进行碱基编辑，通过荧光信号，判断编辑的有无与效率的高低。具体实施如下：

1、报告系统的构建

报告蛋白的编码核苷酸序列如SEQ ID NO.31所示。当所述报告蛋白的编码核酸的编码链上的第67位氨基酸密码子为GAT(对应非编码链上的ATC)，所述报告系统编码的报告蛋白不显示荧光。当利用碱基编辑系统靶向非编码链引入突变，将非编码链上的ATC突变为ATG，相应地将第67位氨基酸的密码子突变为CAT时，表达组氨酸(密码子CAT)，如SEQ IDNO.31所示的核苷酸编码的报告蛋白氨基酸如SEQ ID NO.1所示，所述报告系统显示出蓝色荧光；当利用碱基编辑引入突变，将第67位氨基酸的密码子突变为TAT时，表达酪氨酸(密码子TAT)，所述报告系统显示出绿色荧光。通过流式细胞术分析蓝色荧光比例，可以推算出C-G碱基的颠换比例。本实施例构建的含有该报告蛋白的质粒(报告系统)的核苷酸序列如SEQID NO.2所示，相应的报告系统命名为BFP-CG报告系统。

2、sgRNA表达载体的构建

设计对应的sgRNA，靶向非编码链，靶序列为：SEQ ID NO.3：catCggtcagggtggtcacgagg，并构建相应的sgRNA表达载体，构建过程如下：

根据靶向位点序列设计碱基互补配对的上下游引物SEQ ID NO.4：accgcatCggtcagggtggtcacg；SEQ ID NO.5：aaaccgtgaccaccctgaccGatg，加灭菌水溶解至100μM。经退火后连接到pGL3-U6-sgRNA(Addgene#51133)载体上，以构建靶向特异性sgRNA。

2.1退火产物

退火体系如下所示：

表1

上游引物	4.5μL
		下游引物	4.5μL
10×NEB buffer2	1μL

退火程序如下：

表2

95℃	5min
		95-85℃	-2℃/s
85-25℃	-0.1℃/s
		4℃	∞

2.2线性化载体

利用BsaI(NEB,R0535S)对pGL3-U6(Addgene#51133)质粒进行酶切以得到线性化sgRNA载体。酶切体系如下所示：

表3

水	补水至50μL
		PGL3-U6质粒	10μg
10×cutsmart buffer	5μL
		BsaI酶	5μL

2.3退火产物与线性化载体连接

以上反应体系配置好后，置于37℃条件下反应5h，酶切产物用AxyPrep DNA凝胶回收试剂盒(Axygen,AP-GX-250G)做割胶回收得到线性化载体。取50ng线性化载体与3μL退火产物通过T4连接酶(NEB,M0202S)连接，16℃孵育2小时后并转化涂板，经Sanger测序得到正确的靶向特异性sgRNA。连接体系如下：

表4

水	补水至10μL
		PGL3-U6-BsaI酶切线性片段	20ng
退火产物	1μL
		Solution I	5μL

连接产物随后进行转化，复苏30min，涂板于氨卞抗性的LB琼脂平板，37℃培养过夜。挑选单克隆进行测序验证。连接成功且无误后，对其进行质粒抽提。

3、碱基编辑蛋白表达质粒的构建

利用AncBE4max载体(addgene：112094)为骨架，用Apobec3A脱氨酶替换其中的Apobec1,构建成A3A-CBEmax的表达载体(SEQ ID NO.6)(图2所示)，构建方法利用GibsonAssembly Master Mix重组试剂盒(NEB，E2611S)。如上所述，UGI是抑制UNG的作用，防止尿嘧啶的切除，为了提高C-G突变效率，进一步地，本发明构建了去掉UGI的编辑蛋白，A3A-CBEmax-△UGI编辑蛋白的表达载体(SEQ ID NO.7)。在此基础上，本发明添加了UdgX(来自Mycobacterium smegmatis)，构建了A3A-Udgx-CBEmax-△UGI编辑蛋白的表达载体(SEQ IDNO.8)。

4、哺乳动物细胞系转染C-G编辑系统和获得型报告系统

(1)HEK293T细胞接种培养在含10％FBS的DMEM培养液中(HyClone,SH30022.01B)，其中含penicillin(100U/ml)和streptomycin(100μg/ml)。

(2)转染前一天将细胞分至6孔板。次日，待密度至70％-80％时进行转染。

(3)按照Lipofectamine^TM2000 Transfection Reagent(Invitrogen,11668-019)的操作手册，将2μg碱基编辑蛋白质粒、1μg sgRNA表达载体与100ng的BFP-CG报告系统混匀，共转染至细胞中，6-8小时后换液，分别在24小时、48小时、72小时与96小时后进行荧光数据检测。

4、荧光报告系统分析碱基颠换效率

利用flowjo软件对流式细胞术结果进行分析，本发明发现，A3A-CBEmax-△UGI与A3A-Udgx-CBEmax-△UGI可以实现BFP荧光的转换，而A3A-CBEmax无法实现BFP荧光的转换(图3)；且A3A-Udgx-CBEmax-△UGI相比A3A-CBEmax-△UGI具有更高的编辑效率。同时，荧光强度在48小时最高，后续实验将以48小时作为时间点检测荧光强度。

上述结果表明，本发明构建的报告系统可以较为准确地反应C-G编辑效率。

实施例2：不同类型的尿嘧啶结合蛋白UdgX对C-G编辑器的作用效果

1、不同种类UdgX的构建

目前报道的UdgX来自耻垢分枝杆菌，为了进一步提高碱基编辑系统的编辑效果，本发明利用进化树分析了19种不同类型的UdgX(图4)，并分别构建了19种A3A-Udgx-CBEmax-△UGI编辑器蛋白。相应的每种UdgX来源的微生物名称分别为类胞内分枝杆菌(Mycobacterium paraintracellulare)、少动鞘氨醇单胞菌(Sphingomonaspaucimobilis)、球形红假单胞菌(Cereibacter sphaeroides)、柑桔溃疡病菌(Xanthomonas citri)、苜蓿中华根瘤菌(Sinorhizobium meliloti)、皮诺卡氏菌(Nocardia farcinica)、盖尔森基兴奴卡氏菌(Nocardia cyriacigeorgica)、哥伦布分枝杆菌(Mycobacterium colombiense)、贪铜菌(Cupriavidus taiwanensis)、阿氟曼链霉菌(Streptomyces scabiei)、欺诈伯克霍尔德菌(Burkholderia dolosa)、梅洛尼鞘氨醇单胞菌(Sphingomonas melonis)、沼泽红假单胞菌(Rhodopseudomonas palustris)、耻垢分枝杆菌(Mycolicibacterium smegmatis)、胞内分枝杆菌(Mycobacterium intracellulare)、多噬伯克霍尔德菌(Burkholderia multivorans)、慢生型大豆根瘤菌(Bradyrhizobiumjaponicum)、高效固氮慢生根瘤菌(Bradyrhizobium diazoefficiens)、草莓角斑病菌(Xanthomonas fragariae)，分析不同类型的UdgX对编辑效果的影响。

2、哺乳动物细胞系转染C-G编辑系统和获得型报告系统

(1)HEK293T细胞接种培养在含10％FBS的DMEM培养液中(HyClone,SH30022.01B)，其中含penicillin(100U/ml)和streptomycin(100μg/ml)。

(2)转染前一天将细胞分至6孔板。次日，待密度至70％-80％时进行转染。

(3)按照Lipofectamine^TM2000 Transfection Reagent(Invitrogen,11668-019)的操作手册，将2μg碱基编辑蛋白质粒、1μg sgRNA表达载体与100ng对应的BFP-CG报告系统混匀，共转染至细胞中，6-8小时后换液，48小时后对BFP荧光信号进行检测分析。

3、不同碱基编辑器对C-G突变效率影响分析

利用flowjo软件对流式细胞术结果进行分析，通过比对BFP荧光信号，判断C-G碱基颠换的效果。结果表明来自皮诺卡氏菌(Nocardia farcinica)的UdgX(nfUdgx)展示出了更高的C-G编辑效果(图5)。后续实验本发明将A3A-nfUdgx-CBEmax-△UGI编辑蛋白的表达载体(SEQ ID NO.8)对应的碱基编辑器命名为CGBE。

实施例3：嵌合型CGBE的构建

已有研究表明脱氨酶容易造成随机脱靶，对碱基编辑器的安全构成了威胁。为此，本发明通过将脱氨酶与UdgX复合蛋白，插入到Cas9蛋白结构域内，可以有效避免脱氨酶的随机脱氨，起到降低脱靶的效果(图6)。

1、pET-nCas9-gRNA-AmpR(Y260X)-KanR质粒载体的构建

利用Gibson Assembly Master Mix重组试剂盒(NEB，E2611S)构建pET-nCas9-gRNA-AmpR(Y260X)-KanR质粒载体(SEQ ID NO.9)。该质粒氨苄抗性基因在260位氨基酸处含有终止密码子TAG，当TAG通过碱基编辑器编辑成TAC时，氨苄抗性才发挥作用，相应的菌在氨苄抗生素的平板上才能生长。

2、用于MuA转座酶构建随机插入的受体质粒

利用扩增出的APOBEC-nfUdgX(SEQ ID NO.10)，pET-nCas9-gRNA-AmpR(Y260X)-KanR)质粒(SEQ ID NO.9)，在MuA转座酶(Thermo Fisher，F-701)作用下，在体外构建不同位置随机插入的载体。具体反应体系如下：

表5

APOBEC-nfUdgX片段	250ng
		质粒(SEQ ID NO.9)	500ng
MuA转座酶	1μL
		5×Reaction Buffer for MuA Transposase	4μL
水	补水至20μL

反应液在30℃孵育1小时以实现随机插入，然后在75℃孵育10分钟以灭活MuA转座酶。随后用异丙醇沉淀纯化出DNA，并重悬于5μL去离子水中，然后电转到100μL BL21(DE3)Electro(Shanghai Weidi Biotechnology，EE1002)感受态细胞中。

3、大肠杆菌中筛选功能性嵌入融合CGBE蛋白的表达质粒

将上述电转后的感受态细胞在SOC培养基中复苏1小时后的细菌涂布在数个含10μg/mL卡那霉素的LB琼脂板上，并在37℃下孵育16小时。然后刮下来平板上的菌落，并将其重悬于含有500μM IPTG的100mL LB中。将培养物温育10-12h以诱导nCas9表达并修复AmpR(Y260X)上的突变。然后将减少量的细胞(5mL，1mL，500μL，100μL)接种到含有氨苄青霉素(10μg/mL)和卡那霉素(10μg/mL)的15cm LB琼脂平板上。37℃孵育过夜后，挑选菌落并进行Sanger测序，以评估AmpR(Y260X)上的碱基编辑并确定APOBEC-nfUdgX插入位点。筛选到以下插入位点，具体位置为328，645，698，794，960，979，993，998，1003，1009，1027，1039，1043，1053，1098，1102，1298，1300。

4、嵌合型CGBE对报告系统的编辑

通过上述获得插入位点，本发明构建了相应的哺乳动物碱基编辑蛋白表达质粒，分别命名为CE-CGBE-328，CE-CGBE-645，CE-CGBE-698，CE-CGBE-794，CE-CGBE-960，CE-CGBE-979，CE-CGBE-993，CE-CGBE-998，CE-CGBE-1003，CE-CGBE-1009，CE-CGBE-1027，CE-CGBE-1039，CE-CGBE-1043，CE-CGBE-1053，CE-CGBE-1098，CE-CGBE-1102，CE-CGBE-1298，CE-CGBE-1300。CE-CGBE-1003、CE-CGBE-1027和CE-CGBE-1039相对应表达的碱基编辑蛋白氨基酸序列分别为SEQ ID NO.11、SEQ ID NO.12和SEQ ID NO.13。

按照Lipofectamine^TM2000 Transfection Reagent(Invitrogen,11668-019)的操作手册，将2μg编辑蛋白表达质粒、1μg的sgRNA表达载体与100ng对应的BFP-CG报告系统混匀，共转染至HEK293T细胞中，6-8小时后换液，48小时后对BFP荧光信号进行分析。利用flowjo软件对流式细胞术结果进行分析，通过比对BFP效率，判断C-G碱基颠换的效果。结果表明，编辑效率最高的三个位点为1003，1027以及1039。后续本发明将以1027位置为插入位点，构建碱基编辑蛋白表达质粒，进行内源基因编辑，获得的嵌合型CGBE编辑器命名为CE-CGBE。

实施例4：CE-CGBE可以对内源基因实现高效的C-G突变

为进一步研究C-G碱基颠换编辑器的作用特点及效率，本发明针对5个内源基因进行编辑。具体实施如下：

1、靶位点的选择与相应sgRNA表达载体的构建。

5个位点选择如下：

FAM171b：ACAACAACAGCAAAAGCAGCTGG(SEQ ID NO.14)；

POLR2A:ACTTCAAGAACTAGTGCGCAGG(SEQ ID NO.15)；

VISTA:GCGGTACCACGTCTTGTAGAAGG(SEQ ID NO.16)；

NANOG:ACCAGAGAATGAAATCTAAGAGG(SEQ ID NO.17)；

DDX53:TGATCAAGAGCGAGCAGTAGAGG(SEQ ID NO.18)。

设计5个位点相应的sgRNA上下游引物，序列为SEQ ID NO.19-SEQ ID NO.28。上下游序列通过程序(95℃，5min；95℃-85℃(以2℃/s速度降温)；85℃-25℃(以0.1℃/s的速度降温)；维持4℃)退火，连接到经过BsaI(NEB：R0539L)线性化的PGL3-U6-sgRNA-EGFP(addgene：107721)载体上。对阳性克隆摇菌，提取质粒(Axygene：AP-MN-P-250G)，测定浓度备用。

2、不同类型脱氨酶的CE-CGBE编辑器构建

利用Gibson Assembly Master Mix重组试剂盒(NEB，E2611S)构建不同脱氨酶的CE-CGBE编辑器(图8)。APOBEC-nfUdgX插入位点为1027。本发明利用了ancAPOBEC1与A3A两种脱氨酶分别构建碱基编辑蛋白，分别命名为CE-CGBE-ancApobec1(碱基编辑蛋白的氨基酸序列如SEQ ID NO.29所示)，与CE-CGBE-A3A(碱基编辑蛋白的氨基酸序列如SEQ IDNO.12所示)。按照实施例1第3节“碱基编辑蛋白表达质粒的构建”的方法，构建相应的碱基编辑蛋白表达质粒。

2、CE-CGBE对内源基因的编辑

将HEK293T细胞接种培养于添加10％FBS的DMEM高糖培养液中(HyClone,SH30022.01B)，其中含penicillin(100U/ml)和streptomycin(100μg/ml)。转染前两个小时换成无抗生素的培养基，按照Lipofectamine^TM2000 Transfection Reagent(Invitrogen,11668-019)的操作手册，将2μg编辑蛋白表达质粒、1μg的sgRNA表达载体混匀，共转染至细胞中，6-8小时后换液，72小时后分选出10000个GFP阳性细胞。细胞通过裂解鉴定基因型，裂解液的成分为50mM KCl，1.5mM MgCl₂，10mM Tris pH 8.0，0.5％Nonidet P-40，0.5％Tween 20，100μg/ml protease K。

3、CE-CGBE对内源基因的编辑效果分析

利用一代sanger测序，本发明对两种编辑器和相应的5个位点进行了分析(图9)，并进一步统计了相应的编辑效率(图10)。两种编辑器均能实现C-G的编辑，其中CE-CGBE-A3A最高能实现70％的C-G编辑，CE-CGBE-AncApobec1最高实现55％的C-G编辑。

实施例5：CE-CGBE可以显著降低DNA和RNA的脱靶

胞嘧啶碱基编辑器在DNA与RNA水平均被报道易发生脱靶，本实施例将分析CE-CGBE-A3A对内源基因编辑后DNA与RNA水平的脱靶。

HEK293T细胞接种培养于添加10％FBS的DMEM高糖培养液中(HyClone,SH30022.01B)，其中含penicillin(100U/ml)和streptomycin(100μg/ml)。转染前两个小时换成无抗生素的培养基，按照Lipofectamine^TM2000 Transfection Reagent(Invitrogen,11668-019)的操作手册，将4μg碱基编辑蛋白表达质粒(实施例4构建的4μg CE-CGBE-A3A编辑蛋白表达质粒、实施例1构建的A3A-Udgx-CBEmax-△UGI碱基编辑蛋白)、2μg的sgRNA表达载体(对应FAM171B位点)混匀，共转染至细胞中，6-8小时后换液，72小时后分选出500000个GFP阳性细胞。

分选的细胞通过提取DNA与RNA，送全基因组测序与RNA-测序。通过与未转染编辑器的阴性对照比较，发现CE-CGBE在DNA与RNA水平的脱靶，与参考基因组没有显著差异，与A3A-Udgx-CBEmax-△UGI相比，具有显著的降低(图11)。表明本发明提出的碱基颠换编辑器具有高效、安全的特点。

综上所述，本发明高效特异实现碱基颠换的编辑系统、方法及用途，可以对靶位点中的C颠换成G。本发明所述的融合蛋白、编辑系统或方法可以在多种细胞如哺乳动物的细胞系及原代细胞中高效实现C-G碱基的颠换，为修复致病突变，研究基因功能以及改善细胞功能等提供了有效的工具，具备良好的应用前景。所以，本发明有效克服了现有技术中的种种缺点而具高度产业利用价值。

以上的实施例是为了说明本发明公开的实施方案，并不能理解为对本发明的限制。此外，本文所列出的各种修改以及发明中方法、组合物的变化，在不脱离本发明的范围和精神的前提下对本领域内的技术人员来说是显而易见的。虽然已结合本发明的多种具体优选实施例对本发明进行了具体的描述，但应当理解，本发明不应仅限于这些具体实施例。事实上，各种如上所述的对本领域内的技术人员来说显而易见的修改来获取发明都应包括在本发明的范围内。

序列表

<110> 上海贝斯昂科生物科技有限公司

<120> 一种高效特异实现碱基颠换的编辑系统、方法及用途

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gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 60

ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 120

cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 180

ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 240

gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 300

tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 360

cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 420

attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 480

atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 540

atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 600

tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 660

actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 720

aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 780

gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca 840

ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc 900

atggtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt cgagctggac 960

ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga tgccacctac 1020

ggcaagctga ccctgaagtt catctgcacc accggcaaac tgcccgtgcc ctggcccacc 1080

ctcgtgacca ccctgaccga tggcgtgcag tgcttcagcc gctaccccga ccacatgaag 1140

cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg caccatcttc 1200

ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg cgacaccctg 1260

gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat cctggggcac 1320

aagctggagt acaactacaa cagccacaac gtctatatca tggccgacaa gcagaagaac 1380

ggcatcaagg tgaacttcaa gatccgccac aacatcgagg acggcagcgt gcagctcgcc 1440

gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc cgacaaccac 1500

tacctgagca cccagtccaa gctgagcaaa gaccccaacg agaagcgcga tcacatggtc 1560

ctgctggagt tcgtgaccgc cgccgggatc actctcggca tggacgagct gtacaagtga 1620

aagcttggta ccgagctcgg atccactagt ccagtgtggt ggaattctgc agatatccag 1680

cacagtggcg gccgctcgag tctagagggc ccgtttaaac ccgctgatca gcctcgactg 1740

tgccttctag ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg 1800

aaggtgccac tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga 1860

gtaggtgtca ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg 1920

aagacaatag caggcatgct ggggatgcgg tgggctctat ggcttctgag gcggaaagaa 1980

ccagctgggg ctctaggggg tatccccacg cgccctgtag cggcgcatta agcgcggcgg 2040

gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt 2100

tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc 2160

gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg 2220

attagggtga tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga 2280

cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc 2340

ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa 2400

aaaatgagct gatttaacaa aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt 2460

agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa 2520

ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag 2580

catgcatctc aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct 2640

aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc 2700

agaggccgag gccgcctctg cctctgagct attccagaag tagtgaggag gcttttttgg 2760

aggcctaggc ttttgcaaaa agctcccggg agcttgtata tccattttcg gatctgatca 2820

agagacagga tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg caggttctcc 2880

ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa tcggctgctc 2940

tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg tcaagaccga 3000

cctgtccggt gccctgaatg aactgcagga cgaggcagcg cggctatcgt ggctggccac 3060

gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa gggactggct 3120

gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc ctgccgagaa 3180

agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg ctacctgccc 3240

attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg aagccggtct 3300

tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg aactgttcgc 3360

caggctcaag gcgcgcatgc ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg 3420

cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact gtggccggct 3480

gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg ctgaagagct 3540

tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc ccgattcgca 3600

gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga gcgggactct ggggttcgaa 3660

atgaccgacc aagcgacgcc caacctgcca tcacgagatt tcgattccac cgccgccttc 3720

tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc 3780

ggggatctca tgctggagtt cttcgcccac cccaacttgt ttattgcagc ttataatggt 3840

tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct 3900

agttgtggtt tgtccaaact catcaatgta tcttatcatg tctgtatacc gtcgacctct 3960

agctagagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc 4020

acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga 4080

gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg 4140

tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg 4200

cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg 4260

gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga 4320

aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg 4380

gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag 4440

aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc 4500

gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg 4560

ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt 4620

cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc 4680

ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc 4740

actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg 4800

tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca 4860

gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc 4920

ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct 4980

ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg 5040

gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt 5100

aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt 5160

gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc 5220

gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg 5280

cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc 5340

gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg 5400

gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca 5460

ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga 5520

tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct 5580

ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg 5640

cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca 5700

accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata 5760

cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct 5820

tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact 5880

cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa 5940

acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc 6000

atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga 6060

tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga 6120

aaagtgccac ctgacgtc 6138

<210> 3

<211> 23

<212> DNA

<213> 人工序列(Artificial Sequence)

<400> 3

catcggtcag ggtggtcacg agg 23

<210> 4

<211> 24

<212> DNA

<213> 人工序列(Artificial Sequence)

<400> 4

accgcatcgg tcagggtggt cacg 24

<210> 5

<211> 24

<212> DNA

<213> 人工序列(Artificial Sequence)

<400> 5

aaaccgtgac caccctgacc gatg 24

<210> 6

<211> 8877

<212> DNA

<213> 人工序列(Artificial Sequence)

<400> 6

gcactaatct gagcgacatc attgagaagg agactgggaa acagctggtc attcaggagt 60

ccatcctgat gctgcctgag gaggtggagg aagtgatcgg caacaagcca gagtctgaca 120

tcctggtgca caccgcctac gacgagtcca cagatgagaa tgtgatgctg ctgacctctg 180

acgcccccga gtataagcct tgggccctgg tcatccagga ttctaacggc gagaataaga 240

tcaagatgct gagcggagga tccggaggat ctggaggcag caccaacctg tctgacatca 300

tcgagaagga gacaggcaag cagctggtca tccaggagag catcctgatg ctgcccgaag 360

aagtcgaaga agtgatcgga aacaagcctg agagcgatat cctggtccat accgcctacg 420

acgagagtac cgacgaaaat gtgatgctgc tgacatccga cgccccagag tataagccct 480

gggctctggt catccaggat tccaacggag agaacaaaat caaaatgctg tctggcggct 540

caaaaagaac cgccgacggc agcgaattcg agcccaagaa gaagaggaaa gtctaaccgg 600

tcatcatcac catcaccatt gagtttaaac ccgctgatca gcctcgactg tgccttctag 660

ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac 720

tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca 780

ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg aagacaatag 840

caggcatgct ggggatgcgg tgggctctat ggcttctgag gcggaaagaa ccagctgggg 900

ctcgataccg tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt 960

gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa 1020

agcctaggat gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc 1080

tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcgggaag 1140

aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt 1200

cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga 1260

atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg 1320

taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa 1380

aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt 1440

tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct 1500

gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct 1560

cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc 1620

cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt 1680

atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc 1740

tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat 1800

ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa 1860

acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa 1920

aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacactc agtggaacga 1980

aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct 2040

tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga 2100

cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc 2160

catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg 2220

ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat 2280

aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat 2340

ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg 2400

caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc 2460

attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa 2520

agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc 2580

actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt 2640

ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag 2700

ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt 2760

gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag 2820

atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac 2880

cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc 2940

gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca 3000

gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg 3060

ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc gacggatcgg gagatcgatc 3120

tcccgatccc ctagggtcga ctctcagtac aatctgctct gatgccgcat agttaagcca 3180

gtatctgctc cctgcttgtg tgttggaggt cgctgagtag tgcgcgagca aaatttaagc 3240

tacaacaagg caaggcttga ccgacaattg catgaagaat ctgcttaggg ttaggcgttt 3300

tgcgctgctt cgcgatgtac gggccagata tacgcgttga cattgattat tgactagtta 3360

ttaatagtaa tcaattacgg ggtcattagt tcatagccca tatatggagt tccgcgttac 3420

ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc cattgacgtc 3480

aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac gtcaatgggt 3540

ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata tgccaagtac 3600

gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattatgccc agtacatgac 3660

cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta ttaccatggt 3720

gatgcggttt tggcagtaca tcaatgggcg tggatagcgg tttgactcac ggggatttcc 3780

aagtctccac cccattgacg tcaatgggag tttgttttgg caccaaaatc aacgggactt 3840

tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg 3900

ggaggtctat ataagcagag ctggtttagt gaaccgtcag atccgctaga gatccgcggc 3960

cgctaatacg actcactata gggagagccg ccaccatgaa acggacagcc gacggaagcg 4020

agttcgagtc accaaagaag aagcggaaag tcagcagtga ggcatctcca gcaagcggac 4080

caaggcacct gatggacccc cacatcttca cctctaactt taacaatggc atcggcaggc 4140

acaagacata cctgtgctat gaggtggagc gcctggacaa tggcaccagc gtgaagatgg 4200

atcagcacag aggcttcctg cacaaccagg ccaagaatct gctgtgcggc ttctacggcc 4260

ggcacgcaga gctgagattt ctggacctgg tgcctagcct gcagctggat ccagcccaga 4320

tctatagggt gacctggttc atcagctggt ccccatgctt ttcctgggga tgtgcaggag 4380

aggtgcgcgc ctttctgcag gagaacacac acgtgcggct gagaatcttc gccgcccgga 4440

tctttgacta cgatcctctg tataaggagg ccctgcagat gctgagagac gcaggagccc 4500

aggtgtccat catgacctac gatgagttca agcactgctg ggacacattt gtggatcacc 4560

agggctgtcc cttccagcct tgggacggac tggatgagca ctcccaggcc ctgtctggca 4620

ggctgagggc catcctgcag aaccagggca attctggagg atctagcgga ggatcctctg 4680

gcagcgagac accaggaaca agcgagtcag caacaccaga gagcagtggc ggcagcagcg 4740

gcggcagcga caagaagtac agcatcggcc tggccatcgg caccaactct gtgggctggg 4800

ccgtgatcac cgacgagtac aaggtgccca gcaagaaatt caaggtgctg ggcaacaccg 4860

accggcacag catcaagaag aacctgatcg gagccctgct gttcgacagc ggcgaaacag 4920

ccgaggccac ccggctgaag agaaccgcca gaagaagata caccagacgg aagaaccgga 4980

tctgctatct gcaagagatc ttcagcaacg agatggccaa ggtggacgac agcttcttcc 5040

acagactgga agagtccttc ctggtggaag aggataagaa gcacgagcgg caccccatct 5100

tcggcaacat cgtggacgag gtggcctacc acgagaagta ccccaccatc taccacctga 5160

gaaagaaact ggtggacagc accgacaagg ccgacctgcg gctgatctat ctggccctgg 5220

cccacatgat caagttccgg ggccacttcc tgatcgaggg cgacctgaac cccgacaaca 5280

gcgacgtgga caagctgttc atccagctgg tgcagaccta caaccagctg ttcgaggaaa 5340

accccatcaa cgccagcggc gtggacgcca aggccatcct gtctgccaga ctgagcaaga 5400

gcagacggct ggaaaatctg atcgcccagc tgcccggcga gaagaagaat ggcctgttcg 5460

gaaacctgat tgccctgagc ctgggcctga cccccaactt caagagcaac ttcgacctgg 5520

ccgaggatgc caaactgcag ctgagcaagg acacctacga cgacgacctg gacaacctgc 5580

tggcccagat cggcgaccag tacgccgacc tgtttctggc cgccaagaac ctgtccgacg 5640

ccatcctgct gagcgacatc ctgagagtga acaccgagat caccaaggcc cccctgagcg 5700

cctctatgat caagagatac gacgagcacc accaggacct gaccctgctg aaagctctcg 5760

tgcggcagca gctgcctgag aagtacaaag agattttctt cgaccagagc aagaacggct 5820

acgccggcta cattgacggc ggagccagcc aggaagagtt ctacaagttc atcaagccca 5880

tcctggaaaa gatggacggc accgaggaac tgctcgtgaa gctgaacaga gaggacctgc 5940

tgcggaagca gcggaccttc gacaacggca gcatccccca ccagatccac ctgggagagc 6000

tgcacgccat tctgcggcgg caggaagatt tttacccatt cctgaaggac aaccgggaaa 6060

agatcgagaa gatcctgacc ttccgcatcc cctactacgt gggccctctg gccaggggaa 6120

acagcagatt cgcctggatg accagaaaga gcgaggaaac catcaccccc tggaacttcg 6180

aggaagtggt ggacaagggc gcttccgccc agagcttcat cgagcggatg accaacttcg 6240

ataagaacct gcccaacgag aaggtgctgc ccaagcacag cctgctgtac gagtacttca 6300

ccgtgtataa cgagctgacc aaagtgaaat acgtgaccga gggaatgaga aagcccgcct 6360

tcctgagcgg cgagcagaaa aaggccatcg tggacctgct gttcaagacc aaccggaaag 6420

tgaccgtgaa gcagctgaaa gaggactact tcaagaaaat cgagtgcttc gactccgtgg 6480

aaatctccgg cgtggaagat cggttcaacg cctccctggg cacataccac gatctgctga 6540

aaattatcaa ggacaaggac ttcctggaca atgaggaaaa cgaggacatt ctggaagata 6600

tcgtgctgac cctgacactg tttgaggaca gagagatgat cgaggaacgg ctgaaaacct 6660

atgcccacct gttcgacgac aaagtgatga agcagctgaa gcggcggaga tacaccggct 6720

ggggcaggct gagccggaag ctgatcaacg gcatccggga caagcagtcc ggcaagacaa 6780

tcctggattt cctgaagtcc gacggcttcg ccaacagaaa cttcatgcag ctgatccacg 6840

acgacagcct gacctttaaa gaggacatcc agaaagccca ggtgtccggc cagggcgata 6900

gcctgcacga gcacattgcc aatctggccg gcagccccgc cattaagaag ggcatcctgc 6960

agacagtgaa ggtggtggac gagctcgtga aagtgatggg ccggcacaag cccgagaaca 7020

tcgtgatcga aatggccaga gagaaccaga ccacccagaa gggacagaag aacagccgcg 7080

agagaatgaa gcggatcgaa gagggcatca aagagctggg cagccagatc ctgaaagaac 7140

accccgtgga aaacacccag ctgcagaacg agaagctgta cctgtactac ctgcagaatg 7200

ggcgggatat gtacgtggac caggaactgg acatcaaccg gctgtccgac tacgatgtgg 7260

accatatcgt gcctcagagc tttctgaagg acgactccat cgacaacaag gtgctgacca 7320

gaagcgacaa gaaccggggc aagagcgaca acgtgccctc cgaagaggtc gtgaagaaga 7380

tgaagaacta ctggcggcag ctgctgaacg ccaagctgat tacccagaga aagttcgaca 7440

atctgaccaa ggccgagaga ggcggcctga gcgaactgga taaggccggc ttcatcaaga 7500

gacagctggt ggaaacccgg cagatcacaa agcacgtggc acagatcctg gactcccgga 7560

tgaacactaa gtacgacgag aatgacaagc tgatccggga agtgaaagtg atcaccctga 7620

agtccaagct ggtgtccgat ttccggaagg atttccagtt ttacaaagtg cgcgagatca 7680

acaactacca ccacgcccac gacgcctacc taaacgccgt cgtgggaacc gccctgatca 7740

aaaagtaccc taagctggaa agcgagttcg tgtacggcga ctacaaggtg tacgacgtgc 7800

ggaagatgat cgccaagagc gagcaggaaa tcggcaaggc taccgccaag tacttcttct 7860

acagcaacat catgaacttt ttcaagaccg agattaccct ggccaacggc gagatccgga 7920

agcggcctct gatcgagaca aacggcgaaa ccggggagat cgtgtgggat aagggccggg 7980

attttgccac cgtgcggaaa gtgctgagca tgccccaagt gaatatcgtg aaaaagaccg 8040

aggtgcagac aggcggcttc agcaaagagt ctatcctgcc caagaggaac agcgataagc 8100

tgatcgccag aaagaaggac tgggacccta agaagtacgg cggcttcgac agccccaccg 8160

tggcctattc tgtgctggtg gtggccaaag tggaaaaggg caagtccaag aaactgaaga 8220

gtgtgaaaga gctgctgggg atcaccatca tggaaagaag cagcttcgag aagaatccca 8280

tcgactttct ggaagccaag ggctacaaag aagtgaaaaa ggacctgatc atcaagctgc 8340

ctaagtactc cctgttcgag ctggaaaacg gccggaagag aatgctggcc tctgccggcg 8400

aactgcagaa gggaaacgaa ctggccctgc cctccaaata tgtgaacttc ctgtacctgg 8460

ccagccacta tgagaagctg aagggctccc ccgaggataa tgagcagaaa cagctgtttg 8520

tggaacagca caagcactac ctggacgaga tcatcgagca gatcagcgag ttctccaaga 8580

gagtgatcct ggccgacgct aatctggaca aagtgctgtc cgcctacaac aagcaccggg 8640

ataagcccat cagagagcag gccgagaata tcatccacct gtttaccctg accaatctgg 8700

gagcccctgc cgccttcaag tactttgaca ccaccatcga ccggaagagg tacaccagca 8760

ccaaagaggt gctggacgcc accctgatcc accagagcat caccggcctg tacgagacac 8820

ggatcgacct gtctcagctg ggaggtgaca gcggcgggag cggcgggagc gggggga 8877

<210> 7

<211> 8319

<212> DNA

<213> 人工序列(Artificial Sequence)

<400> 7

atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa 60

cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg ggaagaggcg gtttgcgtat 120

tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg 180

agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc 240

aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt 300

gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag 360

tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc 420

cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc 480

ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt 540

cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt 600

atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc 660

agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa 720

gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa 780

gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg 840

tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga 900

agatcctttg atcttttcta cggggtctga cactcagtgg aacgaaaact cacgttaagg 960

gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg 1020

aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt 1080

aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact 1140

ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat 1200

gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg 1260

aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg 1320

ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat 1380

tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc 1440

ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt 1500

cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc 1560

agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga 1620

gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc 1680

gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa 1740

acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta 1800

acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg 1860

agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg 1920

aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat 1980

gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt 2040

tccccgaaaa gtgccacctg acgtcgacgg atcgggagat cgatctcccg atcccctagg 2100

gtcgactctc agtacaatct gctctgatgc cgcatagtta agccagtatc tgctccctgc 2160

ttgtgtgttg gaggtcgctg agtagtgcgc gagcaaaatt taagctacaa caaggcaagg 2220

cttgaccgac aattgcatga agaatctgct tagggttagg cgttttgcgc tgcttcgcga 2280

tgtacgggcc agatatacgc gttgacattg attattgact agttattaat agtaatcaat 2340

tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa 2400

tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt 2460

tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta 2520

aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt 2580

caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc 2640

tacttggcag tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca 2700

gtacatcaat gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat 2760

tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa 2820

caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag 2880

cagagctggt ttagtgaacc gtcagatccg ctagagatcc gcggccgcta atacgactca 2940

ctatagggag agccgccacc atgaaacgga cagccgacgg aagcgagttc gagtcaccaa 3000

agaagaagcg gaaagtcagc agtgaggcat ctccagcaag cggaccaagg cacctgatgg 3060

acccccacat cttcacctct aactttaaca atggcatcgg caggcacaag acatacctgt 3120

gctatgaggt ggagcgcctg gacaatggca ccagcgtgaa gatggatcag cacagaggct 3180

tcctgcacaa ccaggccaag aatctgctgt gcggcttcta cggccggcac gcagagctga 3240

gatttctgga cctggtgcct agcctgcagc tggatccagc ccagatctat agggtgacct 3300

ggttcatcag ctggtcccca tgcttttcct ggggatgtgc aggagaggtg cgcgcctttc 3360

tgcaggagaa cacacacgtg cggctgagaa tcttcgccgc ccggatcttt gactacgatc 3420

ctctgtataa ggaggccctg cagatgctga gagacgcagg agcccaggtg tccatcatga 3480

cctacgatga gttcaagcac tgctgggaca catttgtgga tcaccagggc tgtcccttcc 3540

agccttggga cggactggat gagcactccc aggccctgtc tggcaggctg agggccatcc 3600

tgcagaacca gggcaattct ggaggatcta gcggaggatc ctctggcagc gagacaccag 3660

gaacaagcga gtcagcaaca ccagagagca gtggcggcag cagcggcggc agcgacaaga 3720

agtacagcat cggcctggcc atcggcacca actctgtggg ctgggccgtg atcaccgacg 3780

agtacaaggt gcccagcaag aaattcaagg tgctgggcaa caccgaccgg cacagcatca 3840

agaagaacct gatcggagcc ctgctgttcg acagcggcga aacagccgag gccacccggc 3900

tgaagagaac cgccagaaga agatacacca gacggaagaa ccggatctgc tatctgcaag 3960

agatcttcag caacgagatg gccaaggtgg acgacagctt cttccacaga ctggaagagt 4020

ccttcctggt ggaagaggat aagaagcacg agcggcaccc catcttcggc aacatcgtgg 4080

acgaggtggc ctaccacgag aagtacccca ccatctacca cctgagaaag aaactggtgg 4140

acagcaccga caaggccgac ctgcggctga tctatctggc cctggcccac atgatcaagt 4200

tccggggcca cttcctgatc gagggcgacc tgaaccccga caacagcgac gtggacaagc 4260

tgttcatcca gctggtgcag acctacaacc agctgttcga ggaaaacccc atcaacgcca 4320

gcggcgtgga cgccaaggcc atcctgtctg ccagactgag caagagcaga cggctggaaa 4380

atctgatcgc ccagctgccc ggcgagaaga agaatggcct gttcggaaac ctgattgccc 4440

tgagcctggg cctgaccccc aacttcaaga gcaacttcga cctggccgag gatgccaaac 4500

tgcagctgag caaggacacc tacgacgacg acctggacaa cctgctggcc cagatcggcg 4560

accagtacgc cgacctgttt ctggccgcca agaacctgtc cgacgccatc ctgctgagcg 4620

acatcctgag agtgaacacc gagatcacca aggcccccct gagcgcctct atgatcaaga 4680

gatacgacga gcaccaccag gacctgaccc tgctgaaagc tctcgtgcgg cagcagctgc 4740

ctgagaagta caaagagatt ttcttcgacc agagcaagaa cggctacgcc ggctacattg 4800

acggcggagc cagccaggaa gagttctaca agttcatcaa gcccatcctg gaaaagatgg 4860

acggcaccga ggaactgctc gtgaagctga acagagagga cctgctgcgg aagcagcgga 4920

ccttcgacaa cggcagcatc ccccaccaga tccacctggg agagctgcac gccattctgc 4980

ggcggcagga agatttttac ccattcctga aggacaaccg ggaaaagatc gagaagatcc 5040

tgaccttccg catcccctac tacgtgggcc ctctggccag gggaaacagc agattcgcct 5100

ggatgaccag aaagagcgag gaaaccatca ccccctggaa cttcgaggaa gtggtggaca 5160

agggcgcttc cgcccagagc ttcatcgagc ggatgaccaa cttcgataag aacctgccca 5220

acgagaaggt gctgcccaag cacagcctgc tgtacgagta cttcaccgtg tataacgagc 5280

tgaccaaagt gaaatacgtg accgagggaa tgagaaagcc cgccttcctg agcggcgagc 5340

agaaaaaggc catcgtggac ctgctgttca agaccaaccg gaaagtgacc gtgaagcagc 5400

tgaaagagga ctacttcaag aaaatcgagt gcttcgactc cgtggaaatc tccggcgtgg 5460

aagatcggtt caacgcctcc ctgggcacat accacgatct gctgaaaatt atcaaggaca 5520

aggacttcct ggacaatgag gaaaacgagg acattctgga agatatcgtg ctgaccctga 5580

cactgtttga ggacagagag atgatcgagg aacggctgaa aacctatgcc cacctgttcg 5640

acgacaaagt gatgaagcag ctgaagcggc ggagatacac cggctggggc aggctgagcc 5700

ggaagctgat caacggcatc cgggacaagc agtccggcaa gacaatcctg gatttcctga 5760

agtccgacgg cttcgccaac agaaacttca tgcagctgat ccacgacgac agcctgacct 5820

ttaaagagga catccagaaa gcccaggtgt ccggccaggg cgatagcctg cacgagcaca 5880

ttgccaatct ggccggcagc cccgccatta agaagggcat cctgcagaca gtgaaggtgg 5940

tggacgagct cgtgaaagtg atgggccggc acaagcccga gaacatcgtg atcgaaatgg 6000

ccagagagaa ccagaccacc cagaagggac agaagaacag ccgcgagaga atgaagcgga 6060

tcgaagaggg catcaaagag ctgggcagcc agatcctgaa agaacacccc gtggaaaaca 6120

cccagctgca gaacgagaag ctgtacctgt actacctgca gaatgggcgg gatatgtacg 6180

tggaccagga actggacatc aaccggctgt ccgactacga tgtggaccat atcgtgcctc 6240

agagctttct gaaggacgac tccatcgaca acaaggtgct gaccagaagc gacaagaacc 6300

ggggcaagag cgacaacgtg ccctccgaag aggtcgtgaa gaagatgaag aactactggc 6360

ggcagctgct gaacgccaag ctgattaccc agagaaagtt cgacaatctg accaaggccg 6420

agagaggcgg cctgagcgaa ctggataagg ccggcttcat caagagacag ctggtggaaa 6480

cccggcagat cacaaagcac gtggcacaga tcctggactc ccggatgaac actaagtacg 6540

acgagaatga caagctgatc cgggaagtga aagtgatcac cctgaagtcc aagctggtgt 6600

ccgatttccg gaaggatttc cagttttaca aagtgcgcga gatcaacaac taccaccacg 6660

cccacgacgc ctacctaaac gccgtcgtgg gaaccgccct gatcaaaaag taccctaagc 6720

tggaaagcga gttcgtgtac ggcgactaca aggtgtacga cgtgcggaag atgatcgcca 6780

agagcgagca ggaaatcggc aaggctaccg ccaagtactt cttctacagc aacatcatga 6840

actttttcaa gaccgagatt accctggcca acggcgagat ccggaagcgg cctctgatcg 6900

agacaaacgg cgaaaccggg gagatcgtgt gggataaggg ccgggatttt gccaccgtgc 6960

ggaaagtgct gagcatgccc caagtgaata tcgtgaaaaa gaccgaggtg cagacaggcg 7020

gcttcagcaa agagtctatc ctgcccaaga ggaacagcga taagctgatc gccagaaaga 7080

aggactggga ccctaagaag tacggcggct tcgacagccc caccgtggcc tattctgtgc 7140

tggtggtggc caaagtggaa aagggcaagt ccaagaaact gaagagtgtg aaagagctgc 7200

tggggatcac catcatggaa agaagcagct tcgagaagaa tcccatcgac tttctggaag 7260

ccaagggcta caaagaagtg aaaaaggacc tgatcatcaa gctgcctaag tactccctgt 7320

tcgagctgga aaacggccgg aagagaatgc tggcctctgc cggcgaactg cagaagggaa 7380

acgaactggc cctgccctcc aaatatgtga acttcctgta cctggccagc cactatgaga 7440

agctgaaggg ctcccccgag gataatgagc agaaacagct gtttgtggaa cagcacaagc 7500

actacctgga cgagatcatc gagcagatca gcgagttctc caagagagtg atcctggccg 7560

acgctaatct ggacaaagtg ctgtccgcct acaacaagca ccgggataag cccatcagag 7620

agcaggccga gaatatcatc cacctgttta ccctgaccaa tctgggagcc cctgccgcct 7680

tcaagtactt tgacaccacc atcgaccgga agaggtacac cagcaccaaa gaggtgctgg 7740

acgccaccct gatccaccag agcatcaccg gcctgtacga gacacggatc gacctgtctc 7800

agctgggagg tgactctggc ggctcaaaaa gaaccgccga cggcagcgaa ttcgagccca 7860

agaagaagag gaaagtctaa ccggtcatca tcaccatcac cattgagttt aaacccgctg 7920

atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc 7980

ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc 8040

atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa 8100

gggggaggat tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggcttc 8160

tgaggcggaa agaaccagct ggggctcgat accgtcgacc tctagctaga gcttggcgta 8220

atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat 8280

acgagccgga agcataaagt gtaaagccta ggatgccta 8319

<210> 8

<211> 8976

<212> DNA

<213> 人工序列(Artificial Sequence)

<400> 8

atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa 60

cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg ggaagaggcg gtttgcgtat 120

tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg 180

agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc 240

aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt 300

gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag 360

tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc 420

cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc 480

ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt 540

cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt 600

atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc 660

agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa 720

gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa 780

gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg 840

tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga 900

agatcctttg atcttttcta cggggtctga cactcagtgg aacgaaaact cacgttaagg 960

gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg 1020

aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt 1080

aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact 1140

ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat 1200

gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg 1260

aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg 1320

ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat 1380

tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc 1440

ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt 1500

cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc 1560

agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga 1620

gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc 1680

gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa 1740

acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta 1800

acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg 1860

agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg 1920

aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat 1980

gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt 2040

tccccgaaaa gtgccacctg acgtcgacgg atcgggagat cgatctcccg atcccctagg 2100

gtcgactctc agtacaatct gctctgatgc cgcatagtta agccagtatc tgctccctgc 2160

ttgtgtgttg gaggtcgctg agtagtgcgc gagcaaaatt taagctacaa caaggcaagg 2220

cttgaccgac aattgcatga agaatctgct tagggttagg cgttttgcgc tgcttcgcga 2280

tgtacgggcc agatatacgc gttgacattg attattgact agttattaat agtaatcaat 2340

tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa 2400

tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt 2460

tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta 2520

aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt 2580

caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc 2640

tacttggcag tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca 2700

gtacatcaat gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat 2760

tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa 2820

caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag 2880

cagagctggt ttagtgaacc gtcagatccg ctagagatcc gcggccgcta atacgactca 2940

ctatagggag agccgccacc atgaaacgga cagccgacgg aagcgagttc gagtcaccaa 3000

agaagaagcg gaaagtcagc agtgaggcat ctccagcaag cggaccaagg cacctgatgg 3060

acccccacat cttcacctct aactttaaca atggcatcgg caggcacaag acatacctgt 3120

gctatgaggt ggagcgcctg gacaatggca ccagcgtgaa gatggatcag cacagaggct 3180

tcctgcacaa ccaggccaag aatctgctgt gcggcttcta cggccggcac gcagagctga 3240

gatttctgga cctggtgcct agcctgcagc tggatccagc ccagatctat agggtgacct 3300

ggttcatcag ctggtcccca tgcttttcct ggggatgtgc aggagaggtg cgcgcctttc 3360

tgcaggagaa cacacacgtg cggctgagaa tcttcgccgc ccggatcttt gactacgatc 3420

ctctgtataa ggaggccctg cagatgctga gagacgcagg agcccaggtg tccatcatga 3480

cctacgatga gttcaagcac tgctgggaca catttgtgga tcaccagggc tgtcccttcc 3540

agccttggga cggactggat gagcactccc aggccctgtc tggcaggctg agggccatcc 3600

tgcagaacca gggcaatagc ggaggatccg gaggatctgg aggcagcatg gccggagcac 3660

aggattttgt cccacatact gccgatctgg ctgagctggc tgccgccgct ggagagtgta 3720

gaggatgcgg gctgtaccgg gatgccacac aggccgtgtt cggagcaggc ggcaggagcg 3780

cccgcatcat gatgatcggc gagcagccag gcgataagga ggacctggcc ggcctgccat 3840

ttgtgggacc agcaggccgg ctgctggaca gagccctgga ggccgccgac atcgataggg 3900

acgccctgta cgtgaccaac gccgtgaagc acttcaagtt tacacgcgcc gcaggaggca 3960

agaggagaat ccacaagacc ccctctcgga cagaggtggt ggcctgcaga ccttggctga 4020

tcgccgagat gaccagcgtg gagccagatg tggtggtgct gctgggagca acagcagcaa 4080

aggccctgct gggcaatgac ttcagggtga cccagcacag gggagaggtg ctgcacgtgg 4140

acgatgtgcc aggcgatcct gccctggtgg caaccgtgca ccctagctcc ctgctgagag 4200

gcccaaagga ggagcgggaa tccgccttcg ctgggctggt ggacgacctg agagtggctg 4260

ccgatgtgag accttctgga ggatctagcg gaggatcctc tggcagcgag acaccaggaa 4320

caagcgagtc agcaacacca gagagcagtg gcggcagcag cggcggcagc gacaagaagt 4380

acagcatcgg cctggccatc ggcaccaact ctgtgggctg ggccgtgatc accgacgagt 4440

acaaggtgcc cagcaagaaa ttcaaggtgc tgggcaacac cgaccggcac agcatcaaga 4500

agaacctgat cggagccctg ctgttcgaca gcggcgaaac agccgaggcc acccggctga 4560

agagaaccgc cagaagaaga tacaccagac ggaagaaccg gatctgctat ctgcaagaga 4620

tcttcagcaa cgagatggcc aaggtggacg acagcttctt ccacagactg gaagagtcct 4680

tcctggtgga agaggataag aagcacgagc ggcaccccat cttcggcaac atcgtggacg 4740

aggtggccta ccacgagaag taccccacca tctaccacct gagaaagaaa ctggtggaca 4800

gcaccgacaa ggccgacctg cggctgatct atctggccct ggcccacatg atcaagttcc 4860

ggggccactt cctgatcgag ggcgacctga accccgacaa cagcgacgtg gacaagctgt 4920

tcatccagct ggtgcagacc tacaaccagc tgttcgagga aaaccccatc aacgccagcg 4980

gcgtggacgc caaggccatc ctgtctgcca gactgagcaa gagcagacgg ctggaaaatc 5040

tgatcgccca gctgcccggc gagaagaaga atggcctgtt cggaaacctg attgccctga 5100

gcctgggcct gacccccaac ttcaagagca acttcgacct ggccgaggat gccaaactgc 5160

agctgagcaa ggacacctac gacgacgacc tggacaacct gctggcccag atcggcgacc 5220

agtacgccga cctgtttctg gccgccaaga acctgtccga cgccatcctg ctgagcgaca 5280

tcctgagagt gaacaccgag atcaccaagg cccccctgag cgcctctatg atcaagagat 5340

acgacgagca ccaccaggac ctgaccctgc tgaaagctct cgtgcggcag cagctgcctg 5400

agaagtacaa agagattttc ttcgaccaga gcaagaacgg ctacgccggc tacattgacg 5460

gcggagccag ccaggaagag ttctacaagt tcatcaagcc catcctggaa aagatggacg 5520

gcaccgagga actgctcgtg aagctgaaca gagaggacct gctgcggaag cagcggacct 5580

tcgacaacgg cagcatcccc caccagatcc acctgggaga gctgcacgcc attctgcggc 5640

ggcaggaaga tttttaccca ttcctgaagg acaaccggga aaagatcgag aagatcctga 5700

ccttccgcat cccctactac gtgggccctc tggccagggg aaacagcaga ttcgcctgga 5760

tgaccagaaa gagcgaggaa accatcaccc cctggaactt cgaggaagtg gtggacaagg 5820

gcgcttccgc ccagagcttc atcgagcgga tgaccaactt cgataagaac ctgcccaacg 5880

agaaggtgct gcccaagcac agcctgctgt acgagtactt caccgtgtat aacgagctga 5940

ccaaagtgaa atacgtgacc gagggaatga gaaagcccgc cttcctgagc ggcgagcaga 6000

aaaaggccat cgtggacctg ctgttcaaga ccaaccggaa agtgaccgtg aagcagctga 6060

aagaggacta cttcaagaaa atcgagtgct tcgactccgt ggaaatctcc ggcgtggaag 6120

atcggttcaa cgcctccctg ggcacatacc acgatctgct gaaaattatc aaggacaagg 6180

acttcctgga caatgaggaa aacgaggaca ttctggaaga tatcgtgctg accctgacac 6240

tgtttgagga cagagagatg atcgaggaac ggctgaaaac ctatgcccac ctgttcgacg 6300

acaaagtgat gaagcagctg aagcggcgga gatacaccgg ctggggcagg ctgagccgga 6360

agctgatcaa cggcatccgg gacaagcagt ccggcaagac aatcctggat ttcctgaagt 6420

ccgacggctt cgccaacaga aacttcatgc agctgatcca cgacgacagc ctgaccttta 6480

aagaggacat ccagaaagcc caggtgtccg gccagggcga tagcctgcac gagcacattg 6540

ccaatctggc cggcagcccc gccattaaga agggcatcct gcagacagtg aaggtggtgg 6600

acgagctcgt gaaagtgatg ggccggcaca agcccgagaa catcgtgatc gaaatggcca 6660

gagagaacca gaccacccag aagggacaga agaacagccg cgagagaatg aagcggatcg 6720

aagagggcat caaagagctg ggcagccaga tcctgaaaga acaccccgtg gaaaacaccc 6780

agctgcagaa cgagaagctg tacctgtact acctgcagaa tgggcgggat atgtacgtgg 6840

accaggaact ggacatcaac cggctgtccg actacgatgt ggaccatatc gtgcctcaga 6900

gctttctgaa ggacgactcc atcgacaaca aggtgctgac cagaagcgac aagaaccggg 6960

gcaagagcga caacgtgccc tccgaagagg tcgtgaagaa gatgaagaac tactggcggc 7020

agctgctgaa cgccaagctg attacccaga gaaagttcga caatctgacc aaggccgaga 7080

gaggcggcct gagcgaactg gataaggccg gcttcatcaa gagacagctg gtggaaaccc 7140

ggcagatcac aaagcacgtg gcacagatcc tggactcccg gatgaacact aagtacgacg 7200

agaatgacaa gctgatccgg gaagtgaaag tgatcaccct gaagtccaag ctggtgtccg 7260

atttccggaa ggatttccag ttttacaaag tgcgcgagat caacaactac caccacgccc 7320

acgacgccta cctaaacgcc gtcgtgggaa ccgccctgat caaaaagtac cctaagctgg 7380

aaagcgagtt cgtgtacggc gactacaagg tgtacgacgt gcggaagatg atcgccaaga 7440

gcgagcagga aatcggcaag gctaccgcca agtacttctt ctacagcaac atcatgaact 7500

ttttcaagac cgagattacc ctggccaacg gcgagatccg gaagcggcct ctgatcgaga 7560

caaacggcga aaccggggag atcgtgtggg ataagggccg ggattttgcc accgtgcgga 7620

aagtgctgag catgccccaa gtgaatatcg tgaaaaagac cgaggtgcag acaggcggct 7680

tcagcaaaga gtctatcctg cccaagagga acagcgataa gctgatcgcc agaaagaagg 7740

actgggaccc taagaagtac ggcggcttcg acagccccac cgtggcctat tctgtgctgg 7800

tggtggccaa agtggaaaag ggcaagtcca agaaactgaa gagtgtgaaa gagctgctgg 7860

ggatcaccat catggaaaga agcagcttcg agaagaatcc catcgacttt ctggaagcca 7920

agggctacaa agaagtgaaa aaggacctga tcatcaagct gcctaagtac tccctgttcg 7980

agctggaaaa cggccggaag agaatgctgg cctctgccgg cgaactgcag aagggaaacg 8040

aactggccct gccctccaaa tatgtgaact tcctgtacct ggccagccac tatgagaagc 8100

tgaagggctc ccccgaggat aatgagcaga aacagctgtt tgtggaacag cacaagcact 8160

acctggacga gatcatcgag cagatcagcg agttctccaa gagagtgatc ctggccgacg 8220

ctaatctgga caaagtgctg tccgcctaca acaagcaccg ggataagccc atcagagagc 8280

aggccgagaa tatcatccac ctgtttaccc tgaccaatct gggagcccct gccgccttca 8340

agtactttga caccaccatc gaccggaaga ggtacaccag caccaaagag gtgctggacg 8400

ccaccctgat ccaccagagc atcaccggcc tgtacgagac acggatcgac ctgtctcagc 8460

tgggaggtga ctctggcggc tcaaaaagaa ccgccgacgg cagcgaattc gagcccaaga 8520

agaagaggaa agtctaaccg gtcatcatca ccatcaccat tgagtttaaa cccgctgatc 8580

agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc 8640

cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc 8700

gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg 8760

ggaggattgg gaagacaata gcaggcatgc tggggatgcg gtgggctcta tggcttctga 8820

ggcggaaaga accagctggg gctcgatacc gtcgacctct agctagagct tggcgtaatc 8880

atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg 8940

agccggaagc ataaagtgta aagcctagga tgccta 8976

<210> 9

<211> 10864

<212> DNA

<213> 人工序列(Artificial Sequence)

<400> 9

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 ctctggcggc tcaaaaagaa ccgccgacgg cagcgaattc 4140

gagcccaaga agaagaggaa agtctaaccg gtcatcatca ccatcaccat tgagtttaaa 4200

cccgctgatc agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc 4260

ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg 4320

aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg 4380

acagcaaggg ggaggattgg gaagacaata gcaggcatgc tggggatgcg gtgggctcta 4440

tggcttctga ggcggaaaga accagctggg gctcgttgac agctagctca gtcctaggta 4500

taatactagt gtcgtctaga taactacgat agttttagag ctagaaatag caagttaaaa 4560

taaggctagt ccgttatcaa cttgaaaaag tggcaccgag tcggtgcttt ttttgatccg 4620

gctgctaaca aagcccgaaa ggaagctgag ttggctgctg ccaccgctga gcaataacta 4680

gcataacccc ttggggcctc taaacgggtc ttgaggggtt ttttgctgaa aggaggaact 4740

atatccggat tggcgaatgg gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg 4800

tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt 4860

tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc 4920

tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg 4980

gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg 5040

agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct 5100

cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg 5160

agctgattta acaaaaattt aacgcgaatt ttaacaaaat attaacgttt acaatttcag 5220

gtggcacttt tcggggaaat gtgggaaatg tgcgcggaac ccctatttgt ttatttttct 5280

aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat 5340

attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg 5400

cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg 5460

aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc 5520

ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat 5580

gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact 5640

attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca 5700

tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact 5760

tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg 5820

atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg 5880

agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg 5940

aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg 6000

caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag 6060

ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc 6120

gtatcgtagt tatctagacg acggggagtc aggcaactat ggatgaacga aatagacaga 6180

tcgctgagat aggtgcctca ctgattaagc attggtaagc gcggaacccc tatttgttta 6240

tttttctaaa tacattcaaa tatgtatccg ctcatgaatt aattcttaga aaaactcatc 6300

gagcatcaaa tgaaactgca atttattcat atcaggatta tcaataccat atttttgaaa 6360

aagccgtttc tgtaatgaag gagaaaactc accgaggcag ttccatagga tggcaagatc 6420

ctggtatcgg tctgcgattc cgactcgtcc aacatcaata caacctatta atttcccctc 6480

gtcaaaaata aggttatcaa gtgagaaatc accatgagtg acgactgaat ccggtgagaa 6540

tggcaaaagt ttatgcattt ctttccagac ttgttcaaca ggccagccat tacgctcgtc 6600

atcaaaatca ctcgcatcaa ccaaaccgtt attcattcgt gattgcgcct gagcgagacg 6660

aaatacgcga tcgctgttaa aaggacaatt acaaacagga atcgaatgca accggcgcag 6720

gaacactgcc agcgcatcaa caatattttc acctgaatca ggatattctt ctaatacctg 6780

gaatgctgtt ttcccgggga tcgcagtggt gagtaaccat gcatcatcag gagtacggat 6840

aaaatgcttg atggtcggaa gaggcataaa ttccgtcagc cagtttagtc tgaccatctc 6900

atctgtaaca tcattggcaa cgctaccttt gccatgtttc agaaacaact ctggcgcatc 6960

gggcttccca tacaatcgat agattgtcgc acctgattgc ccgacattat cgcgagccca 7020

tttataccca tataaatcag catccatgtt ggaatttaat cgcggcctag agcaagacgt 7080

ttcccgttga atatggctca taacacccct tgtattactg tttatgtaag cagacagttt 7140

tattgttcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg 7200

tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc 7260

aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc 7320

tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc cttctagtgt 7380

agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc 7440

taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact 7500

caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac 7560

agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag 7620

aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg 7680

gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg 7740

tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga 7800

gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt 7860

ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct 7920

ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg 7980

aggaagcgga agagcgcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac 8040

accgcatata tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagta 8100

tacactccgc tatcgctacg tgactgggtc atggctgcgc cccgacaccc gccaacaccc 8160

gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc 8220

gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgaggcag 8280

ctgcggtaaa gctcatcagc gtggtcgtga agcgattcac agatgtctgc ctgttcatcc 8340

gcgtccagct cgttgagttt ctccagaagc gttaatgtct ggcttctgat aaagcgggcc 8400

atgttaaggg cggttttttc ctgtttggtc actgatgcct ccgtgtaagg gggatttctg 8460

ttcatggggg taatgatacc gatgaaacga gagaggatgc tcacgatacg ggttactgat 8520

gatgaacatg cccggttact ggaacgttgt gagggtaaac aactggcggt atggatgcgg 8580

cgggaccaga gaaaaatcac tcagggtcaa tgccagcgct tcgttaatac agatgtaggt 8640

gttccacagg gtagccagca gcatcctgcg atgcagatcc ggaacataat ggtgcagggc 8700

gctgacttcc gcgtttccag actttacgaa acacggaaac cgaagaccat tcatgttgtt 8760

gctcaggtcg cagacgtttt gcagcagcag tcgcttcacg ttcgctcgcg tatcggtgat 8820

tcattctgct aaccagtaag gcaaccccgc cagcctagcc gggtcctcaa cgacaggagc 8880

acgatcatgc gcacccgtgg ggccgccatg ccggcgataa tggcctgctt ctcgccgaaa 8940

cgtttggtgg cgggaccagt gacgaaggct tgagcgaggg cgtgcaagat tccgaatacc 9000

gcaagcgaca ggccgatcat cgtcgcgctc cagcgaaagc ggtcctcgcc gaaaatgacc 9060

cagagcgctg ccggcacctg tcctacgagt tgcatgataa agaagacagt cataagtgcg 9120

gcgacgatag tcatgccccg cgcccaccgg aaggagctga ctgggttgaa ggctctcaag 9180

ggcatcggtc gagatcccgg tgcctaatga gtgagctaac ttacattaat tgcgttgcgc 9240

tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa 9300

cgcgcgggga gaggcggttt gcgtattggg cgccagggtg gtttttcttt tcaccagtga 9360

gacgggcaac agctgattgc ccttcaccgc ctggccctga gagagttgca gcaagcggtc 9420

cacgctggtt tgccccagca ggcgaaaatc ctgtttgatg gtggttaacg gcgggatata 9480

acatgagctg tcttcggtat cgtcgtatcc cactaccgag atgtccgcac caacgcgcag 9540

cccggactcg gtaatggcgc gcattgcgcc cagcgccatc tgatcgttgg caaccagcat 9600

cgcagtggga acgatgccct cattcagcat ttgcatggtt tgttgaaaac cggacatggc 9660

actccagtcg ccttcccgtt ccgctatcgg ctgaatttga ttgcgagtga gatatttatg 9720

ccagccagcc agacgcagac gcgccgagac agaacttaat gggcccgcta acagcgcgat 9780

ttgctggtga cccaatgcga ccagatgctc cacgcccagt cgcgtaccgt cttcatggga 9840

gaaaataata ctgttgatgg gtgtctggtc agagacatca agaaataacg ccggaacatt 9900

agtgcaggca gcttccacag caatggcatc ctggtcatcc agcggatagt taatgatcag 9960

cccactgacg cgttgcgcga gaagattgtg caccgccgct ttacaggctt cgacgccgct 10020

tcgttctacc atcgacacca ccacgctggc acccagttga tcggcgcgag atttaatcgc 10080

cgcgacaatt tgcgacggcg cgtgcagggc cagactggag gtggcaacgc caatcagcaa 10140

cgactgtttg cccgccagtt gttgtgccac gcggttggga atgtaattca gctccgccat 10200

cgccgcttcc actttttccc gcgttttcgc agaaacgtgg ctggcctggt tcaccacgcg 10260

ggaaacggtc tgataagaga caccggcata ctctgcgaca tcgtataacg ttactggttt 10320

cacattcacc accctgaatt gactctcttc cgggcgctat catgccatac cgcgaaaggt 10380

tttgcgccat tcgatggtgt ccgggatctc gacgctctcc cttatgcgac tcctgcatta 10440

ggaagcagcc cagtagtagg ttgaggccgt tgagcaccgc cgccgcaagg aatggtgcat 10500

gcaaggagat ggcgcccaac agtcccccgg ccacggggcc tgccaccata cccacgccga 10560

aacaagcgct catgagcccg aagtggcgag cccgatcttc cccatcggtg atgtcggcga 10620

tataggcgcc agcaaccgca cctgtggcgc cggtgatgcc ggccacgatg cgtccggcgt 10680

agaggatcga gatcgatctc gatcccgcga aattaatacg actcactata ggggaattgt 10740

gagcggataa caattcccct ctagaaataa ttttgtttaa ctttaagaag gagatataca 10800

tgccaccatg aaacggacag ccgacggaag cgagttcgag tcaccaaaga agaagcggaa 10860

agtc 10864

<210> 10

<211> 1251

<212> DNA

<213> 人工序列(Artificial Sequence)

<400> 10

gaggcatctc cagcaagcgg accaaggcac ctgatggacc cccacatctt cacctctaac 60

tttaacaatg gcatcggcag gcacaagaca tacctgtgct atgaggtgga gcgcctggac 120

aatggcacca gcgtgaagat ggatcagcac agaggcttcc tgcacaacca ggccaagaat 180

ctgctgtgcg gcttctacgg ccggcacgca gagctgagat ttctggacct ggtgcctagc 240

ctgcagctgg atccagccca gatctatagg gtgacctggt tcatcagctg gtccccatgc 300

ttttcctggg gatgtgcagg agaggtgcgc gcctttctgc aggagaacac acacgtgcgg 360

ctgagaatct tcgccgcccg gatctttgac tacgatcctc tgtataagga ggccctgcag 420

atgctgagag acgcaggagc ccaggtgtcc atcatgacct acgatgagtt caagcactgc 480

tgggacacat ttgtggatca ccagggctgt cccttccagc cttgggacgg actggatgag 540

cactcccagg ccctgtctgg caggctgagg gccatcctgc agaaccaggg caatagcgga 600

ggatccggag gatctggagg cagcatggcc ggagcacagg attttgtccc acatactgcc 660

gatctggctg agctggctgc cgccgctgga gagtgtagag gatgcgggct gtaccgggat 720

gccacacagg ccgtgttcgg agcaggcggc aggagcgccc gcatcatgat gatcggcgag 780

cagccaggcg ataaggagga cctggccggc ctgccatttg tgggaccagc aggccggctg 840

ctggacagag ccctggaggc cgccgacatc gatagggacg ccctgtacgt gaccaacgcc 900

gtgaagcact tcaagtttac acgcgccgca ggaggcaaga ggagaatcca caagaccccc 960

tctcggacag aggtggtggc ctgcagacct tggctgatcg ccgagatgac cagcgtggag 1020

ccagatgtgg tggtgctgct gggagcaaca gcagcaaagg ccctgctggg caatgacttc 1080

agggtgaccc agcacagggg agaggtgctg cacgtggacg atgtgccagg cgatcctgcc 1140

ctggtggcaa ccgtgcaccc tagctccctg ctgagaggcc caaaggagga gcgggaatcc 1200

gccttcgctg ggctggtgga cgacctgaga gtggctgccg atgtgagacc t 1251

<210> 11

<211> 1876

<212> PRT

<213> 人工序列(Artificial Sequence)

<400> 11

Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys

1 5 10 15

Arg Lys Val Asp Lys Lys Tyr Ser Ile Gly Leu Ala Ile Gly Thr Asn

20 25 30

Ser Val Gly Trp Ala Val Ile Thr Asp Glu Tyr Lys Val Pro Ser Lys

35 40 45

Lys Phe Lys Val Leu Gly Asn Thr Asp Arg His Ser Ile Lys Lys Asn

50 55 60

Leu Ile Gly Ala Leu Leu Phe Asp Ser Gly Glu Thr Ala Glu Ala Thr

65 70 75 80

Arg Leu Lys Arg Thr Ala Arg Arg Arg Tyr Thr Arg Arg Lys Asn Arg

85 90 95

Ile Cys Tyr Leu Gln Glu Ile Phe Ser Asn Glu Met Ala Lys Val Asp

100 105 110

Asp Ser Phe Phe His Arg Leu Glu Glu Ser Phe Leu Val Glu Glu Asp

115 120 125

Lys Lys His Glu Arg His Pro Ile Phe Gly Asn Ile Val Asp Glu Val

130 135 140

Ala Tyr His Glu Lys Tyr Pro Thr Ile Tyr His Leu Arg Lys Lys Leu

145 150 155 160

Val Asp Ser Thr Asp Lys Ala Asp Leu Arg Leu Ile Tyr Leu Ala Leu

165 170 175

Ala His Met Ile Lys Phe Arg Gly His Phe Leu Ile Glu Gly Asp Leu

180 185 190

Asn Pro Asp Asn Ser Asp Val Asp Lys Leu Phe Ile Gln Leu Val Gln

195 200 205

Thr Tyr Asn Gln Leu Phe Glu Glu Asn Pro Ile Asn Ala Ser Gly Val

210 215 220

Asp Ala Lys Ala Ile Leu Ser Ala Arg Leu Ser Lys Ser Arg Arg Leu

225 230 235 240

Glu Asn Leu Ile Ala Gln Leu Pro Gly Glu Lys Lys Asn Gly Leu Phe

245 250 255

Gly Asn Leu Ile Ala Leu Ser Leu Gly Leu Thr Pro Asn Phe Lys Ser

260 265 270

Asn Phe Asp Leu Ala Glu Asp Ala Lys Leu Gln Leu Ser Lys Asp Thr

275 280 285

Tyr Asp Asp Asp Leu Asp Asn Leu Leu Ala Gln Ile Gly Asp Gln Tyr

290 295 300

Ala Asp Leu Phe Leu Ala Ala Lys Asn Leu Ser Asp Ala Ile Leu Leu

305 310 315 320

Ser Asp Ile Leu Arg Val Asn Thr Glu Ile Thr Lys Ala Pro Leu Ser

325 330 335

Ala Ser Met Ile Lys Arg Tyr Asp Glu His His Gln Asp Leu Thr Leu

340 345 350

Leu Lys Ala Leu Val Arg Gln Gln Leu Pro Glu Lys Tyr Lys Glu Ile

355 360 365

Phe Phe Asp Gln Ser Lys Asn Gly Tyr Ala Gly Tyr Ile Asp Gly Gly

370 375 380

Ala Ser Gln Glu Glu Phe Tyr Lys Phe Ile Lys Pro Ile Leu Glu Lys

385 390 395 400

Met Asp Gly Thr Glu Glu Leu Leu Val Lys Leu Asn Arg Glu Asp Leu

405 410 415

Leu Arg Lys Gln Arg Thr Phe Asp Asn Gly Ser Ile Pro His Gln Ile

420 425 430

His Leu Gly Glu Leu His Ala Ile Leu Arg Arg Gln Glu Asp Phe Tyr

435 440 445

Pro Phe Leu Lys Asp Asn Arg Glu Lys Ile Glu Lys Ile Leu Thr Phe

450 455 460

Arg Ile Pro Tyr Tyr Val Gly Pro Leu Ala Arg Gly Asn Ser Arg Phe

465 470 475 480

Ala Trp Met Thr Arg Lys Ser Glu Glu Thr Ile Thr Pro Trp Asn Phe

485 490 495

Glu Glu Val Val Asp Lys Gly Ala Ser Ala Gln Ser Phe Ile Glu Arg

500 505 510

Met Thr Asn Phe Asp Lys Asn Leu Pro Asn Glu Lys Val Leu Pro Lys

515 520 525

His Ser Leu Leu Tyr Glu Tyr Phe Thr Val Tyr Asn Glu Leu Thr Lys

530 535 540

Val Lys Tyr Val Thr Glu Gly Met Arg Lys Pro Ala Phe Leu Ser Gly

545 550 555 560

Glu Gln Lys Lys Ala Ile Val Asp Leu Leu Phe Lys Thr Asn Arg Lys

565 570 575

Val Thr Val Lys Gln Leu Lys Glu Asp Tyr Phe Lys Lys Ile Glu Cys

580 585 590

Phe Asp Ser Val Glu Ile Ser Gly Val Glu Asp Arg Phe Asn Ala Ser

595 600 605

Leu Gly Thr Tyr His Asp Leu Leu Lys Ile Ile Lys Asp Lys Asp Phe

610 615 620

Leu Asp Asn Glu Glu Asn Glu Asp Ile Leu Glu Asp Ile Val Leu Thr

625 630 635 640

Leu Thr Leu Phe Glu Asp Arg Glu Met Ile Glu Glu Arg Leu Lys Thr

645 650 655

Tyr Ala His Leu Phe Asp Asp Lys Val Met Lys Gln Leu Lys Arg Arg

660 665 670

Arg Tyr Thr Gly Trp Gly Arg Leu Ser Arg Lys Leu Ile Asn Gly Ile

675 680 685

Arg Asp Lys Gln Ser Gly Lys Thr Ile Leu Asp Phe Leu Lys Ser Asp

690 695 700

Gly Phe Ala Asn Arg Asn Phe Met Gln Leu Ile His Asp Asp Ser Leu

705 710 715 720

Thr Phe Lys Glu Asp Ile Gln Lys Ala Gln Val Ser Gly Gln Gly Asp

725 730 735

Ser Leu His Glu His Ile Ala Asn Leu Ala Gly Ser Pro Ala Ile Lys

740 745 750

Lys Gly Ile Leu Gln Thr Val Lys Val Val Asp Glu Leu Val Lys Val

755 760 765

Met Gly Arg His Lys Pro Glu Asn Ile Val Ile Glu Met Ala Arg Glu

770 775 780

Asn Gln Thr Thr Gln Lys Gly Gln Lys Asn Ser Arg Glu Arg Met Lys

785 790 795 800

Arg Ile Glu Glu Gly Ile Lys Glu Leu Gly Ser Gln Ile Leu Lys Glu

805 810 815

His Pro Val Glu Asn Thr Gln Leu Gln Asn Glu Lys Leu Tyr Leu Tyr

820 825 830

Tyr Leu Gln Asn Gly Arg Asp Met Tyr Val Asp Gln Glu Leu Asp Ile

835 840 845

Asn Arg Leu Ser Asp Tyr Asp Val Asp His Ile Val Pro Gln Ser Phe

850 855 860

Leu Lys Asp Asp Ser Ile Asp Asn Lys Val Leu Thr Arg Ser Asp Lys

865 870 875 880

Asn Arg Gly Lys Ser Asp Asn Val Pro Ser Glu Glu Val Val Lys Lys

885 890 895

Met Lys Asn Tyr Trp Arg Gln Leu Leu Asn Ala Lys Leu Ile Thr Gln

900 905 910

Arg Lys Phe Asp Asn Leu Thr Lys Ala Glu Arg Gly Gly Leu Ser Glu

915 920 925

Leu Asp Lys Ala Gly Phe Ile Lys Arg Gln Leu Val Glu Thr Arg Gln

930 935 940

Ile Thr Lys His Val Ala Gln Ile Leu Asp Ser Arg Met Asn Thr Lys

945 950 955 960

Tyr Asp Glu Asn Asp Lys Leu Ile Arg Glu Val Lys Val Ile Thr Leu

965 970 975

Lys Ser Lys Leu Val Ser Asp Phe Arg Lys Asp Phe Gln Phe Tyr Lys

980 985 990

Val Arg Glu Ile Asn Asn Tyr His His Ala His Asp Ala Tyr Leu Asn

995 1000 1005

Ala Val Val Gly Thr Ala Leu Ile Lys Lys Tyr Pro Lys Ser Gly Ser

1010 1015 1020

Glu Thr Pro Gly Thr Ser Glu Ser Ala Thr Pro Glu Ser Gly Ser Glu

1025 1030 1035 1040

Ala Ser Pro Ala Ser Gly Pro Arg His Leu Met Asp Pro His Ile Phe

1045 1050 1055

Thr Ser Asn Phe Asn Asn Gly Ile Gly Arg His Lys Thr Tyr Leu Cys

1060 1065 1070

Tyr Glu Val Glu Arg Leu Asp Asn Gly Thr Ser Val Lys Met Asp Gln

1075 1080 1085

His Arg Gly Phe Leu His Asn Gln Ala Lys Asn Leu Leu Cys Gly Phe

1090 1095 1100

Tyr Gly Arg His Ala Glu Leu Arg Phe Leu Asp Leu Val Pro Ser Leu

1105 1110 1115 1120

Gln Leu Asp Pro Ala Gln Ile Tyr Arg Val Thr Trp Phe Ile Ser Trp

1125 1130 1135

Ser Pro Cys Phe Ser Trp Gly Cys Ala Gly Glu Val Arg Ala Phe Leu

1140 1145 1150

Gln Glu Asn Thr His Val Arg Leu Arg Ile Phe Ala Ala Arg Ile Phe

1155 1160 1165

Asp Tyr Asp Pro Leu Tyr Lys Glu Ala Leu Gln Met Leu Arg Asp Ala

1170 1175 1180

Gly Ala Gln Val Ser Ile Met Thr Tyr Asp Glu Phe Lys His Cys Trp

1185 1190 1195 1200

Asp Thr Phe Val Asp His Gln Gly Cys Pro Phe Gln Pro Trp Asp Gly

1205 1210 1215

Leu Asp Glu His Ser Gln Ala Leu Ser Gly Arg Leu Arg Ala Ile Leu

1220 1225 1230

Gln Asn Gln Gly Asn Ser Gly Gly Ser Gly Gly Ser Gly Gly Ser Met

1235 1240 1245

Ala Ala Ala Lys Ala Pro Gly Ala Ala Glu Phe Val Pro Ala Asp Ala

1250 1255 1260

Asp Leu Asp Thr Leu Arg Thr Ala Val Gln Gly Cys Arg Gly Cys Glu

1265 1270 1275 1280

Leu Tyr Arg Gly Ala Thr Gln Ala Val Phe Gly Glu Gly Pro Ala His

1285 1290 1295

Ala Pro Val Phe Val Val Gly Glu Gln Pro Gly Asp Arg Glu Asp Val

1300 1305 1310

Ala Gly His Pro Phe Val Gly Pro Ala Gly Arg Leu Leu Asp Lys Ala

1315 1320 1325

Leu Thr Glu Ala Asp Ile Asp Arg Glu Ala Val Tyr Leu Thr Asn Ala

1330 1335 1340

Val Lys His Phe Lys Phe Glu Glu Arg Gly Lys Arg Arg Ile His Lys

1345 1350 1355 1360

Gln Pro Gly Arg Thr Glu Val Val Ala Cys Ser Pro Trp Leu Thr Ala

1365 1370 1375

Glu Leu Asp Ala Val Arg Pro Gln Leu Val Val Cys Leu Gly Ala Val

1380 1385 1390

Ala Ala Lys Ala Val Leu Gly Pro Ser Phe Lys Val Ser Glu Arg Arg

1395 1400 1405

Gly Glu Val Val Glu Ala Gly Glu His Arg Val Ile Ala Thr Val His

1410 1415 1420

Pro Ser Ser Val Leu Arg Ala Pro Asp Arg Ala Ala Ala Tyr Ala Asp

1425 1430 1435 1440

Phe Leu Ala Asp Leu Arg Lys Val Arg Thr Ala Ala Gly Glu Leu His

1445 1450 1455

Arg Ala Ser Gly Gly Ser Ser Gly Gly Ser Ser Gly Ser Glu Thr Pro

1460 1465 1470

Gly Thr Ser Glu Ser Ala Thr Pro Glu Ser Ser Gly Gly Ser Ser Gly

1475 1480 1485

Gly Ser Leu Glu Ser Glu Phe Val Tyr Gly Asp Tyr Lys Val Tyr Asp

1490 1495 1500

Val Arg Lys Met Ile Ala Lys Ser Glu Gln Glu Ile Gly Lys Ala Thr

1505 1510 1515 1520

Ala Lys Tyr Phe Phe Tyr Ser Asn Ile Met Asn Phe Phe Lys Thr Glu

1525 1530 1535

Ile Thr Leu Ala Asn Gly Glu Ile Arg Lys Arg Pro Leu Ile Glu Thr

1540 1545 1550

Asn Gly Glu Thr Gly Glu Ile Val Trp Asp Lys Gly Arg Asp Phe Ala

1555 1560 1565

Thr Val Arg Lys Val Leu Ser Met Pro Gln Val Asn Ile Val Lys Lys

1570 1575 1580

Thr Glu Val Gln Thr Gly Gly Phe Ser Lys Glu Ser Ile Leu Pro Lys

1585 1590 1595 1600

Arg Asn Ser Asp Lys Leu Ile Ala Arg Lys Lys Asp Trp Asp Pro Lys

1605 1610 1615

Lys Tyr Gly Gly Phe Asp Ser Pro Thr Val Ala Tyr Ser Val Leu Val

1620 1625 1630

Val Ala Lys Val Glu Lys Gly Lys Ser Lys Lys Leu Lys Ser Val Lys

1635 1640 1645

Glu Leu Leu Gly Ile Thr Ile Met Glu Arg Ser Ser Phe Glu Lys Asn

1650 1655 1660

Pro Ile Asp Phe Leu Glu Ala Lys Gly Tyr Lys Glu Val Lys Lys Asp

1665 1670 1675 1680

Leu Ile Ile Lys Leu Pro Lys Tyr Ser Leu Phe Glu Leu Glu Asn Gly

1685 1690 1695

Arg Lys Arg Met Leu Ala Ser Ala Gly Glu Leu Gln Lys Gly Asn Glu

1700 1705 1710

Leu Ala Leu Pro Ser Lys Tyr Val Asn Phe Leu Tyr Leu Ala Ser His

1715 1720 1725

Tyr Glu Lys Leu Lys Gly Ser Pro Glu Asp Asn Glu Gln Lys Gln Leu

1730 1735 1740

Phe Val Glu Gln His Lys His Tyr Leu Asp Glu Ile Ile Glu Gln Ile

1745 1750 1755 1760

Ser Glu Phe Ser Lys Arg Val Ile Leu Ala Asp Ala Asn Leu Asp Lys

1765 1770 1775

Val Leu Ser Ala Tyr Asn Lys His Arg Asp Lys Pro Ile Arg Glu Gln

1780 1785 1790

Ala Glu Asn Ile Ile His Leu Phe Thr Leu Thr Asn Leu Gly Ala Pro

1795 1800 1805

Ala Ala Phe Lys Tyr Phe Asp Thr Thr Ile Asp Arg Lys Arg Tyr Thr

1810 1815 1820

Ser Thr Lys Glu Val Leu Asp Ala Thr Leu Ile His Gln Ser Ile Thr

1825 1830 1835 1840

Gly Leu Tyr Glu Thr Arg Ile Asp Leu Ser Gln Leu Gly Gly Asp Ser

1845 1850 1855

Gly Gly Ser Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Pro Lys Lys

1860 1865 1870

Lys Arg Lys Val

1875

<210> 12

<211> 1876

<212> PRT

<213> 人工序列(Artificial Sequence)

<400> 12

Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys

1 5 10 15

Arg Lys Val Asp Lys Lys Tyr Ser Ile Gly Leu Ala Ile Gly Thr Asn

20 25 30

Ser Val Gly Trp Ala Val Ile Thr Asp Glu Tyr Lys Val Pro Ser Lys

35 40 45

Lys Phe Lys Val Leu Gly Asn Thr Asp Arg His Ser Ile Lys Lys Asn

50 55 60

Leu Ile Gly Ala Leu Leu Phe Asp Ser Gly Glu Thr Ala Glu Ala Thr

65 70 75 80

Arg Leu Lys Arg Thr Ala Arg Arg Arg Tyr Thr Arg Arg Lys Asn Arg

85 90 95

Ile Cys Tyr Leu Gln Glu Ile Phe Ser Asn Glu Met Ala Lys Val Asp

100 105 110

Asp Ser Phe Phe His Arg Leu Glu Glu Ser Phe Leu Val Glu Glu Asp

115 120 125

Lys Lys His Glu Arg His Pro Ile Phe Gly Asn Ile Val Asp Glu Val

130 135 140

Ala Tyr His Glu Lys Tyr Pro Thr Ile Tyr His Leu Arg Lys Lys Leu

145 150 155 160

Val Asp Ser Thr Asp Lys Ala Asp Leu Arg Leu Ile Tyr Leu Ala Leu

165 170 175

Ala His Met Ile Lys Phe Arg Gly His Phe Leu Ile Glu Gly Asp Leu

180 185 190

Asn Pro Asp Asn Ser Asp Val Asp Lys Leu Phe Ile Gln Leu Val Gln

195 200 205

Thr Tyr Asn Gln Leu Phe Glu Glu Asn Pro Ile Asn Ala Ser Gly Val

210 215 220

Asp Ala Lys Ala Ile Leu Ser Ala Arg Leu Ser Lys Ser Arg Arg Leu

225 230 235 240

Glu Asn Leu Ile Ala Gln Leu Pro Gly Glu Lys Lys Asn Gly Leu Phe

245 250 255

Gly Asn Leu Ile Ala Leu Ser Leu Gly Leu Thr Pro Asn Phe Lys Ser

260 265 270

Asn Phe Asp Leu Ala Glu Asp Ala Lys Leu Gln Leu Ser Lys Asp Thr

275 280 285

Tyr Asp Asp Asp Leu Asp Asn Leu Leu Ala Gln Ile Gly Asp Gln Tyr

290 295 300

Ala Asp Leu Phe Leu Ala Ala Lys Asn Leu Ser Asp Ala Ile Leu Leu

305 310 315 320

Ser Asp Ile Leu Arg Val Asn Thr Glu Ile Thr Lys Ala Pro Leu Ser

325 330 335

Ala Ser Met Ile Lys Arg Tyr Asp Glu His His Gln Asp Leu Thr Leu

340 345 350

Leu Lys Ala Leu Val Arg Gln Gln Leu Pro Glu Lys Tyr Lys Glu Ile

355 360 365

Phe Phe Asp Gln Ser Lys Asn Gly Tyr Ala Gly Tyr Ile Asp Gly Gly

370 375 380

Ala Ser Gln Glu Glu Phe Tyr Lys Phe Ile Lys Pro Ile Leu Glu Lys

385 390 395 400

Met Asp Gly Thr Glu Glu Leu Leu Val Lys Leu Asn Arg Glu Asp Leu

405 410 415

Leu Arg Lys Gln Arg Thr Phe Asp Asn Gly Ser Ile Pro His Gln Ile

420 425 430

His Leu Gly Glu Leu His Ala Ile Leu Arg Arg Gln Glu Asp Phe Tyr

435 440 445

Pro Phe Leu Lys Asp Asn Arg Glu Lys Ile Glu Lys Ile Leu Thr Phe

450 455 460

Arg Ile Pro Tyr Tyr Val Gly Pro Leu Ala Arg Gly Asn Ser Arg Phe

465 470 475 480

Ala Trp Met Thr Arg Lys Ser Glu Glu Thr Ile Thr Pro Trp Asn Phe

485 490 495

Glu Glu Val Val Asp Lys Gly Ala Ser Ala Gln Ser Phe Ile Glu Arg

500 505 510

Met Thr Asn Phe Asp Lys Asn Leu Pro Asn Glu Lys Val Leu Pro Lys

515 520 525

His Ser Leu Leu Tyr Glu Tyr Phe Thr Val Tyr Asn Glu Leu Thr Lys

530 535 540

Val Lys Tyr Val Thr Glu Gly Met Arg Lys Pro Ala Phe Leu Ser Gly

545 550 555 560

Glu Gln Lys Lys Ala Ile Val Asp Leu Leu Phe Lys Thr Asn Arg Lys

565 570 575

Val Thr Val Lys Gln Leu Lys Glu Asp Tyr Phe Lys Lys Ile Glu Cys

580 585 590

Phe Asp Ser Val Glu Ile Ser Gly Val Glu Asp Arg Phe Asn Ala Ser

595 600 605

Leu Gly Thr Tyr His Asp Leu Leu Lys Ile Ile Lys Asp Lys Asp Phe

610 615 620

Leu Asp Asn Glu Glu Asn Glu Asp Ile Leu Glu Asp Ile Val Leu Thr

625 630 635 640

Leu Thr Leu Phe Glu Asp Arg Glu Met Ile Glu Glu Arg Leu Lys Thr

645 650 655

Tyr Ala His Leu Phe Asp Asp Lys Val Met Lys Gln Leu Lys Arg Arg

660 665 670

Arg Tyr Thr Gly Trp Gly Arg Leu Ser Arg Lys Leu Ile Asn Gly Ile

675 680 685

Arg Asp Lys Gln Ser Gly Lys Thr Ile Leu Asp Phe Leu Lys Ser Asp

690 695 700

Gly Phe Ala Asn Arg Asn Phe Met Gln Leu Ile His Asp Asp Ser Leu

705 710 715 720

Thr Phe Lys Glu Asp Ile Gln Lys Ala Gln Val Ser Gly Gln Gly Asp

725 730 735

Ser Leu His Glu His Ile Ala Asn Leu Ala Gly Ser Pro Ala Ile Lys

740 745 750

Lys Gly Ile Leu Gln Thr Val Lys Val Val Asp Glu Leu Val Lys Val

755 760 765

Met Gly Arg His Lys Pro Glu Asn Ile Val Ile Glu Met Ala Arg Glu

770 775 780

Asn Gln Thr Thr Gln Lys Gly Gln Lys Asn Ser Arg Glu Arg Met Lys

785 790 795 800

Arg Ile Glu Glu Gly Ile Lys Glu Leu Gly Ser Gln Ile Leu Lys Glu

805 810 815

His Pro Val Glu Asn Thr Gln Leu Gln Asn Glu Lys Leu Tyr Leu Tyr

820 825 830

Tyr Leu Gln Asn Gly Arg Asp Met Tyr Val Asp Gln Glu Leu Asp Ile

835 840 845

Asn Arg Leu Ser Asp Tyr Asp Val Asp His Ile Val Pro Gln Ser Phe

850 855 860

Leu Lys Asp Asp Ser Ile Asp Asn Lys Val Leu Thr Arg Ser Asp Lys

865 870 875 880

Asn Arg Gly Lys Ser Asp Asn Val Pro Ser Glu Glu Val Val Lys Lys

885 890 895

Met Lys Asn Tyr Trp Arg Gln Leu Leu Asn Ala Lys Leu Ile Thr Gln

900 905 910

Arg Lys Phe Asp Asn Leu Thr Lys Ala Glu Arg Gly Gly Leu Ser Glu

915 920 925

Leu Asp Lys Ala Gly Phe Ile Lys Arg Gln Leu Val Glu Thr Arg Gln

930 935 940

Ile Thr Lys His Val Ala Gln Ile Leu Asp Ser Arg Met Asn Thr Lys

945 950 955 960

Tyr Asp Glu Asn Asp Lys Leu Ile Arg Glu Val Lys Val Ile Thr Leu

965 970 975

Lys Ser Lys Leu Val Ser Asp Phe Arg Lys Asp Phe Gln Phe Tyr Lys

980 985 990

Val Arg Glu Ile Asn Asn Tyr His His Ala His Asp Ala Tyr Leu Asn

995 1000 1005

Ala Val Val Gly Thr Ala Leu Ile Lys Lys Tyr Pro Lys Leu Glu Ser

1010 1015 1020

Glu Phe Val Tyr Gly Asp Tyr Lys Val Tyr Asp Val Arg Lys Met Ile

1025 1030 1035 1040

Ala Lys Ser Glu Gln Ser Gly Ser Glu Thr Pro Gly Thr Ser Glu Ser

1045 1050 1055

Ala Thr Pro Glu Ser Gly Ser Glu Ala Ser Pro Ala Ser Gly Pro Arg

1060 1065 1070

His Leu Met Asp Pro His Ile Phe Thr Ser Asn Phe Asn Asn Gly Ile

1075 1080 1085

Gly Arg His Lys Thr Tyr Leu Cys Tyr Glu Val Glu Arg Leu Asp Asn

1090 1095 1100

Gly Thr Ser Val Lys Met Asp Gln His Arg Gly Phe Leu His Asn Gln

1105 1110 1115 1120

Ala Lys Asn Leu Leu Cys Gly Phe Tyr Gly Arg His Ala Glu Leu Arg

1125 1130 1135

Phe Leu Asp Leu Val Pro Ser Leu Gln Leu Asp Pro Ala Gln Ile Tyr

1140 1145 1150

Arg Val Thr Trp Phe Ile Ser Trp Ser Pro Cys Phe Ser Trp Gly Cys

1155 1160 1165

Ala Gly Glu Val Arg Ala Phe Leu Gln Glu Asn Thr His Val Arg Leu

1170 1175 1180

Arg Ile Phe Ala Ala Arg Ile Phe Asp Tyr Asp Pro Leu Tyr Lys Glu

1185 1190 1195 1200

Ala Leu Gln Met Leu Arg Asp Ala Gly Ala Gln Val Ser Ile Met Thr

1205 1210 1215

Tyr Asp Glu Phe Lys His Cys Trp Asp Thr Phe Val Asp His Gln Gly

1220 1225 1230

Cys Pro Phe Gln Pro Trp Asp Gly Leu Asp Glu His Ser Gln Ala Leu

1235 1240 1245

Ser Gly Arg Leu Arg Ala Ile Leu Gln Asn Gln Gly Asn Ser Gly Gly

1250 1255 1260

Ser Gly Gly Ser Gly Gly Ser Met Ala Ala Ala Lys Ala Pro Gly Ala

1265 1270 1275 1280

Ala Glu Phe Val Pro Ala Asp Ala Asp Leu Asp Thr Leu Arg Thr Ala

1285 1290 1295

Val Gln Gly Cys Arg Gly Cys Glu Leu Tyr Arg Gly Ala Thr Gln Ala

1300 1305 1310

Val Phe Gly Glu Gly Pro Ala His Ala Pro Val Phe Val Val Gly Glu

1315 1320 1325

Gln Pro Gly Asp Arg Glu Asp Val Ala Gly His Pro Phe Val Gly Pro

1330 1335 1340

Ala Gly Arg Leu Leu Asp Lys Ala Leu Thr Glu Ala Asp Ile Asp Arg

1345 1350 1355 1360

Glu Ala Val Tyr Leu Thr Asn Ala Val Lys His Phe Lys Phe Glu Glu

1365 1370 1375

Arg Gly Lys Arg Arg Ile His Lys Gln Pro Gly Arg Thr Glu Val Val

1380 1385 1390

Ala Cys Ser Pro Trp Leu Thr Ala Glu Leu Asp Ala Val Arg Pro Gln

1395 1400 1405

Leu Val Val Cys Leu Gly Ala Val Ala Ala Lys Ala Val Leu Gly Pro

1410 1415 1420

Ser Phe Lys Val Ser Glu Arg Arg Gly Glu Val Val Glu Ala Gly Glu

1425 1430 1435 1440

His Arg Val Ile Ala Thr Val His Pro Ser Ser Val Leu Arg Ala Pro

1445 1450 1455

Asp Arg Ala Ala Ala Tyr Ala Asp Phe Leu Ala Asp Leu Arg Lys Val

1460 1465 1470

Arg Thr Ala Ala Gly Glu Leu His Arg Ala Ser Gly Gly Ser Ser Gly

1475 1480 1485

Gly Ser Ser Gly Ser Glu Thr Pro Gly Thr Ser Glu Ser Ala Thr Pro

1490 1495 1500

Glu Ser Ser Gly Gly Ser Ser Gly Gly Ser Glu Ile Gly Lys Ala Thr

1505 1510 1515 1520

Ala Lys Tyr Phe Phe Tyr Ser Asn Ile Met Asn Phe Phe Lys Thr Glu

1525 1530 1535

Ile Thr Leu Ala Asn Gly Glu Ile Arg Lys Arg Pro Leu Ile Glu Thr

1540 1545 1550

Asn Gly Glu Thr Gly Glu Ile Val Trp Asp Lys Gly Arg Asp Phe Ala

1555 1560 1565

Thr Val Arg Lys Val Leu Ser Met Pro Gln Val Asn Ile Val Lys Lys

1570 1575 1580

Thr Glu Val Gln Thr Gly Gly Phe Ser Lys Glu Ser Ile Leu Pro Lys

1585 1590 1595 1600

Arg Asn Ser Asp Lys Leu Ile Ala Arg Lys Lys Asp Trp Asp Pro Lys

1605 1610 1615

Lys Tyr Gly Gly Phe Asp Ser Pro Thr Val Ala Tyr Ser Val Leu Val

1620 1625 1630

Val Ala Lys Val Glu Lys Gly Lys Ser Lys Lys Leu Lys Ser Val Lys

1635 1640 1645

Glu Leu Leu Gly Ile Thr Ile Met Glu Arg Ser Ser Phe Glu Lys Asn

1650 1655 1660

Pro Ile Asp Phe Leu Glu Ala Lys Gly Tyr Lys Glu Val Lys Lys Asp

1665 1670 1675 1680

Leu Ile Ile Lys Leu Pro Lys Tyr Ser Leu Phe Glu Leu Glu Asn Gly

1685 1690 1695

Arg Lys Arg Met Leu Ala Ser Ala Gly Glu Leu Gln Lys Gly Asn Glu

1700 1705 1710

Leu Ala Leu Pro Ser Lys Tyr Val Asn Phe Leu Tyr Leu Ala Ser His

1715 1720 1725

Tyr Glu Lys Leu Lys Gly Ser Pro Glu Asp Asn Glu Gln Lys Gln Leu

1730 1735 1740

Phe Val Glu Gln His Lys His Tyr Leu Asp Glu Ile Ile Glu Gln Ile

1745 1750 1755 1760

Ser Glu Phe Ser Lys Arg Val Ile Leu Ala Asp Ala Asn Leu Asp Lys

1765 1770 1775

Val Leu Ser Ala Tyr Asn Lys His Arg Asp Lys Pro Ile Arg Glu Gln

1780 1785 1790

Ala Glu Asn Ile Ile His Leu Phe Thr Leu Thr Asn Leu Gly Ala Pro

1795 1800 1805

Ala Ala Phe Lys Tyr Phe Asp Thr Thr Ile Asp Arg Lys Arg Tyr Thr

1810 1815 1820

Ser Thr Lys Glu Val Leu Asp Ala Thr Leu Ile His Gln Ser Ile Thr

1825 1830 1835 1840

Gly Leu Tyr Glu Thr Arg Ile Asp Leu Ser Gln Leu Gly Gly Asp Ser

1845 1850 1855

Gly Gly Ser Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Pro Lys Lys

1860 1865 1870

Lys Arg Lys Val

1875

<210> 13

<211> 1876

<212> PRT

<213> 人工序列(Artificial Sequence)

<400> 13

Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys

1 5 10 15

Arg Lys Val Asp Lys Lys Tyr Ser Ile Gly Leu Ala Ile Gly Thr Asn

20 25 30

Ser Val Gly Trp Ala Val Ile Thr Asp Glu Tyr Lys Val Pro Ser Lys

35 40 45

Lys Phe Lys Val Leu Gly Asn Thr Asp Arg His Ser Ile Lys Lys Asn

50 55 60

Leu Ile Gly Ala Leu Leu Phe Asp Ser Gly Glu Thr Ala Glu Ala Thr

65 70 75 80

Arg Leu Lys Arg Thr Ala Arg Arg Arg Tyr Thr Arg Arg Lys Asn Arg

85 90 95

Ile Cys Tyr Leu Gln Glu Ile Phe Ser Asn Glu Met Ala Lys Val Asp

100 105 110

Asp Ser Phe Phe His Arg Leu Glu Glu Ser Phe Leu Val Glu Glu Asp

115 120 125

Lys Lys His Glu Arg His Pro Ile Phe Gly Asn Ile Val Asp Glu Val

130 135 140

Ala Tyr His Glu Lys Tyr Pro Thr Ile Tyr His Leu Arg Lys Lys Leu

145 150 155 160

Val Asp Ser Thr Asp Lys Ala Asp Leu Arg Leu Ile Tyr Leu Ala Leu

165 170 175

Ala His Met Ile Lys Phe Arg Gly His Phe Leu Ile Glu Gly Asp Leu

180 185 190

Asn Pro Asp Asn Ser Asp Val Asp Lys Leu Phe Ile Gln Leu Val Gln

195 200 205

Thr Tyr Asn Gln Leu Phe Glu Glu Asn Pro Ile Asn Ala Ser Gly Val

210 215 220

Asp Ala Lys Ala Ile Leu Ser Ala Arg Leu Ser Lys Ser Arg Arg Leu

225 230 235 240

Glu Asn Leu Ile Ala Gln Leu Pro Gly Glu Lys Lys Asn Gly Leu Phe

245 250 255

Gly Asn Leu Ile Ala Leu Ser Leu Gly Leu Thr Pro Asn Phe Lys Ser

260 265 270

Asn Phe Asp Leu Ala Glu Asp Ala Lys Leu Gln Leu Ser Lys Asp Thr

275 280 285

Tyr Asp Asp Asp Leu Asp Asn Leu Leu Ala Gln Ile Gly Asp Gln Tyr

290 295 300

Ala Asp Leu Phe Leu Ala Ala Lys Asn Leu Ser Asp Ala Ile Leu Leu

305 310 315 320

Ser Asp Ile Leu Arg Val Asn Thr Glu Ile Thr Lys Ala Pro Leu Ser

325 330 335

Ala Ser Met Ile Lys Arg Tyr Asp Glu His His Gln Asp Leu Thr Leu

340 345 350

Leu Lys Ala Leu Val Arg Gln Gln Leu Pro Glu Lys Tyr Lys Glu Ile

355 360 365

Phe Phe Asp Gln Ser Lys Asn Gly Tyr Ala Gly Tyr Ile Asp Gly Gly

370 375 380

Ala Ser Gln Glu Glu Phe Tyr Lys Phe Ile Lys Pro Ile Leu Glu Lys

385 390 395 400

Met Asp Gly Thr Glu Glu Leu Leu Val Lys Leu Asn Arg Glu Asp Leu

405 410 415

Leu Arg Lys Gln Arg Thr Phe Asp Asn Gly Ser Ile Pro His Gln Ile

420 425 430

His Leu Gly Glu Leu His Ala Ile Leu Arg Arg Gln Glu Asp Phe Tyr

435 440 445

Pro Phe Leu Lys Asp Asn Arg Glu Lys Ile Glu Lys Ile Leu Thr Phe

450 455 460

Arg Ile Pro Tyr Tyr Val Gly Pro Leu Ala Arg Gly Asn Ser Arg Phe

465 470 475 480

Ala Trp Met Thr Arg Lys Ser Glu Glu Thr Ile Thr Pro Trp Asn Phe

485 490 495

Glu Glu Val Val Asp Lys Gly Ala Ser Ala Gln Ser Phe Ile Glu Arg

500 505 510

Met Thr Asn Phe Asp Lys Asn Leu Pro Asn Glu Lys Val Leu Pro Lys

515 520 525

His Ser Leu Leu Tyr Glu Tyr Phe Thr Val Tyr Asn Glu Leu Thr Lys

530 535 540

Val Lys Tyr Val Thr Glu Gly Met Arg Lys Pro Ala Phe Leu Ser Gly

545 550 555 560

Glu Gln Lys Lys Ala Ile Val Asp Leu Leu Phe Lys Thr Asn Arg Lys

565 570 575

Val Thr Val Lys Gln Leu Lys Glu Asp Tyr Phe Lys Lys Ile Glu Cys

580 585 590

Phe Asp Ser Val Glu Ile Ser Gly Val Glu Asp Arg Phe Asn Ala Ser

595 600 605

Leu Gly Thr Tyr His Asp Leu Leu Lys Ile Ile Lys Asp Lys Asp Phe

610 615 620

Leu Asp Asn Glu Glu Asn Glu Asp Ile Leu Glu Asp Ile Val Leu Thr

625 630 635 640

Leu Thr Leu Phe Glu Asp Arg Glu Met Ile Glu Glu Arg Leu Lys Thr

645 650 655

Tyr Ala His Leu Phe Asp Asp Lys Val Met Lys Gln Leu Lys Arg Arg

660 665 670

Arg Tyr Thr Gly Trp Gly Arg Leu Ser Arg Lys Leu Ile Asn Gly Ile

675 680 685

Arg Asp Lys Gln Ser Gly Lys Thr Ile Leu Asp Phe Leu Lys Ser Asp

690 695 700

Gly Phe Ala Asn Arg Asn Phe Met Gln Leu Ile His Asp Asp Ser Leu

705 710 715 720

Thr Phe Lys Glu Asp Ile Gln Lys Ala Gln Val Ser Gly Gln Gly Asp

725 730 735

Ser Leu His Glu His Ile Ala Asn Leu Ala Gly Ser Pro Ala Ile Lys

740 745 750

Lys Gly Ile Leu Gln Thr Val Lys Val Val Asp Glu Leu Val Lys Val

755 760 765

Met Gly Arg His Lys Pro Glu Asn Ile Val Ile Glu Met Ala Arg Glu

770 775 780

Asn Gln Thr Thr Gln Lys Gly Gln Lys Asn Ser Arg Glu Arg Met Lys

785 790 795 800

Arg Ile Glu Glu Gly Ile Lys Glu Leu Gly Ser Gln Ile Leu Lys Glu

805 810 815

His Pro Val Glu Asn Thr Gln Leu Gln Asn Glu Lys Leu Tyr Leu Tyr

820 825 830

Tyr Leu Gln Asn Gly Arg Asp Met Tyr Val Asp Gln Glu Leu Asp Ile

835 840 845

Asn Arg Leu Ser Asp Tyr Asp Val Asp His Ile Val Pro Gln Ser Phe

850 855 860

Leu Lys Asp Asp Ser Ile Asp Asn Lys Val Leu Thr Arg Ser Asp Lys

865 870 875 880

Asn Arg Gly Lys Ser Asp Asn Val Pro Ser Glu Glu Val Val Lys Lys

885 890 895

Met Lys Asn Tyr Trp Arg Gln Leu Leu Asn Ala Lys Leu Ile Thr Gln

900 905 910

Arg Lys Phe Asp Asn Leu Thr Lys Ala Glu Arg Gly Gly Leu Ser Glu

915 920 925

Leu Asp Lys Ala Gly Phe Ile Lys Arg Gln Leu Val Glu Thr Arg Gln

930 935 940

Ile Thr Lys His Val Ala Gln Ile Leu Asp Ser Arg Met Asn Thr Lys

945 950 955 960

Tyr Asp Glu Asn Asp Lys Leu Ile Arg Glu Val Lys Val Ile Thr Leu

965 970 975

Lys Ser Lys Leu Val Ser Asp Phe Arg Lys Asp Phe Gln Phe Tyr Lys

980 985 990

Val Arg Glu Ile Asn Asn Tyr His His Ala His Asp Ala Tyr Leu Asn

995 1000 1005

Ala Val Val Gly Thr Ala Leu Ile Lys Lys Tyr Pro Lys Leu Glu Ser

1010 1015 1020

Glu Phe Val Tyr Gly Asp Tyr Lys Val Tyr Asp Val Arg Lys Met Ile

1025 1030 1035 1040

Ala Lys Ser Glu Gln Glu Ile Gly Lys Ala Thr Ala Lys Tyr Phe Phe

1045 1050 1055

Tyr Ser Gly Ser Glu Thr Pro Gly Thr Ser Glu Ser Ala Thr Pro Glu

1060 1065 1070

Ser Gly Ser Glu Ala Ser Pro Ala Ser Gly Pro Arg His Leu Met Asp

1075 1080 1085

Pro His Ile Phe Thr Ser Asn Phe Asn Asn Gly Ile Gly Arg His Lys

1090 1095 1100

Thr Tyr Leu Cys Tyr Glu Val Glu Arg Leu Asp Asn Gly Thr Ser Val

1105 1110 1115 1120

Lys Met Asp Gln His Arg Gly Phe Leu His Asn Gln Ala Lys Asn Leu

1125 1130 1135

Leu Cys Gly Phe Tyr Gly Arg His Ala Glu Leu Arg Phe Leu Asp Leu

1140 1145 1150

Val Pro Ser Leu Gln Leu Asp Pro Ala Gln Ile Tyr Arg Val Thr Trp

1155 1160 1165

Phe Ile Ser Trp Ser Pro Cys Phe Ser Trp Gly Cys Ala Gly Glu Val

1170 1175 1180

Arg Ala Phe Leu Gln Glu Asn Thr His Val Arg Leu Arg Ile Phe Ala

1185 1190 1195 1200

Ala Arg Ile Phe Asp Tyr Asp Pro Leu Tyr Lys Glu Ala Leu Gln Met

1205 1210 1215

Leu Arg Asp Ala Gly Ala Gln Val Ser Ile Met Thr Tyr Asp Glu Phe

1220 1225 1230

Lys His Cys Trp Asp Thr Phe Val Asp His Gln Gly Cys Pro Phe Gln

1235 1240 1245

Pro Trp Asp Gly Leu Asp Glu His Ser Gln Ala Leu Ser Gly Arg Leu

1250 1255 1260

Arg Ala Ile Leu Gln Asn Gln Gly Asn Ser Gly Gly Ser Gly Gly Ser

1265 1270 1275 1280

Gly Gly Ser Met Ala Ala Ala Lys Ala Pro Gly Ala Ala Glu Phe Val

1285 1290 1295

Pro Ala Asp Ala Asp Leu Asp Thr Leu Arg Thr Ala Val Gln Gly Cys

1300 1305 1310

Arg Gly Cys Glu Leu Tyr Arg Gly Ala Thr Gln Ala Val Phe Gly Glu

1315 1320 1325

Gly Pro Ala His Ala Pro Val Phe Val Val Gly Glu Gln Pro Gly Asp

1330 1335 1340

Arg Glu Asp Val Ala Gly His Pro Phe Val Gly Pro Ala Gly Arg Leu

1345 1350 1355 1360

Leu Asp Lys Ala Leu Thr Glu Ala Asp Ile Asp Arg Glu Ala Val Tyr

1365 1370 1375

Leu Thr Asn Ala Val Lys His Phe Lys Phe Glu Glu Arg Gly Lys Arg

1380 1385 1390

Arg Ile His Lys Gln Pro Gly Arg Thr Glu Val Val Ala Cys Ser Pro

1395 1400 1405

Trp Leu Thr Ala Glu Leu Asp Ala Val Arg Pro Gln Leu Val Val Cys

1410 1415 1420

Leu Gly Ala Val Ala Ala Lys Ala Val Leu Gly Pro Ser Phe Lys Val

1425 1430 1435 1440

Ser Glu Arg Arg Gly Glu Val Val Glu Ala Gly Glu His Arg Val Ile

1445 1450 1455

Ala Thr Val His Pro Ser Ser Val Leu Arg Ala Pro Asp Arg Ala Ala

1460 1465 1470

Ala Tyr Ala Asp Phe Leu Ala Asp Leu Arg Lys Val Arg Thr Ala Ala

1475 1480 1485

Gly Glu Leu His Arg Ala Ser Gly Gly Ser Ser Gly Gly Ser Ser Gly

1490 1495 1500

Ser Glu Thr Pro Gly Thr Ser Glu Ser Ala Thr Pro Glu Ser Ser Gly

1505 1510 1515 1520

Gly Ser Ser Gly Gly Ser Ser Asn Ile Met Asn Phe Phe Lys Thr Glu

1525 1530 1535

Ile Thr Leu Ala Asn Gly Glu Ile Arg Lys Arg Pro Leu Ile Glu Thr

1540 1545 1550

Asn Gly Glu Thr Gly Glu Ile Val Trp Asp Lys Gly Arg Asp Phe Ala

1555 1560 1565

Thr Val Arg Lys Val Leu Ser Met Pro Gln Val Asn Ile Val Lys Lys

1570 1575 1580

Thr Glu Val Gln Thr Gly Gly Phe Ser Lys Glu Ser Ile Leu Pro Lys

1585 1590 1595 1600

Arg Asn Ser Asp Lys Leu Ile Ala Arg Lys Lys Asp Trp Asp Pro Lys

1605 1610 1615

Lys Tyr Gly Gly Phe Asp Ser Pro Thr Val Ala Tyr Ser Val Leu Val

1620 1625 1630

Val Ala Lys Val Glu Lys Gly Lys Ser Lys Lys Leu Lys Ser Val Lys

1635 1640 1645

Glu Leu Leu Gly Ile Thr Ile Met Glu Arg Ser Ser Phe Glu Lys Asn

1650 1655 1660

Pro Ile Asp Phe Leu Glu Ala Lys Gly Tyr Lys Glu Val Lys Lys Asp

1665 1670 1675 1680

Leu Ile Ile Lys Leu Pro Lys Tyr Ser Leu Phe Glu Leu Glu Asn Gly

1685 1690 1695

Arg Lys Arg Met Leu Ala Ser Ala Gly Glu Leu Gln Lys Gly Asn Glu

1700 1705 1710

Leu Ala Leu Pro Ser Lys Tyr Val Asn Phe Leu Tyr Leu Ala Ser His

1715 1720 1725

Tyr Glu Lys Leu Lys Gly Ser Pro Glu Asp Asn Glu Gln Lys Gln Leu

1730 1735 1740

Phe Val Glu Gln His Lys His Tyr Leu Asp Glu Ile Ile Glu Gln Ile

1745 1750 1755 1760

Ser Glu Phe Ser Lys Arg Val Ile Leu Ala Asp Ala Asn Leu Asp Lys

1765 1770 1775

Val Leu Ser Ala Tyr Asn Lys His Arg Asp Lys Pro Ile Arg Glu Gln

1780 1785 1790

Ala Glu Asn Ile Ile His Leu Phe Thr Leu Thr Asn Leu Gly Ala Pro

1795 1800 1805

Ala Ala Phe Lys Tyr Phe Asp Thr Thr Ile Asp Arg Lys Arg Tyr Thr

1810 1815 1820

Ser Thr Lys Glu Val Leu Asp Ala Thr Leu Ile His Gln Ser Ile Thr

1825 1830 1835 1840

Gly Leu Tyr Glu Thr Arg Ile Asp Leu Ser Gln Leu Gly Gly Asp Ser

1845 1850 1855

Gly Gly Ser Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Pro Lys Lys

1860 1865 1870

Lys Arg Lys Val

1875

<210> 14

<211> 23

<212> DNA

<213> 人工序列(Artificial Sequence)

<400> 14

acaacaacag caaaagcagc tgg 23

<210> 15

<211> 22

<212> DNA

<213> 人工序列(Artificial Sequence)

<400> 15

acttcaagaa ctagtgcgca gg 22

<210> 16

<211> 23

<212> DNA

<213> 人工序列(Artificial Sequence)

<400> 16

gcggtaccac gtcttgtaga agg 23

<210> 17

<211> 23

<212> DNA

<213> 人工序列(Artificial Sequence)

<400> 17

accagagaat gaaatctaag agg 23

<210> 18

<211> 23

<212> DNA

<213> 人工序列(Artificial Sequence)

<400> 18

tgatcaagag cgagcagtag agg 23

<210> 19

<211> 24

<212> DNA

<213> 人工序列(Artificial Sequence)

<400> 19

accgacaaca acagcaaaag cagc 24

<210> 20

<211> 24

<212> DNA

<213> 人工序列(Artificial Sequence)

<400> 20

aaacgctgct tttgctgttg ttgt 24

<210> 21

<211> 23

<212> DNA

<213> 人工序列(Artificial Sequence)

<400> 21

accgacttca agaactagtg cgc 23

<210> 22

<211> 23

<212> DNA

<213> 人工序列(Artificial Sequence)

<400> 22

aaacgcgcac tagttcttga agt 23

<210> 23

<211> 24

<212> DNA

<213> 人工序列(Artificial Sequence)

<400> 23

accggcggta ccacgtcttg taga 24

<210> 24

<211> 24

<212> DNA

<213> 人工序列(Artificial Sequence)

<400> 24

aaactctaca agacgtggta ccgc 24

<210> 25

<211> 24

<212> DNA

<213> 人工序列(Artificial Sequence)

<400> 25

accgaccaga gaatgaaatc taag 24

<210> 26

<211> 24

<212> DNA

<213> 人工序列(Artificial Sequence)

<400> 26

aaaccttaga tttcattctc tggt 24

<210> 27

<211> 24

<212> DNA

<213> 人工序列(Artificial Sequence)

<400> 27

accgtgatca agagcgagca gtag 24

<210> 28

<211> 24

<212> DNA

<213> 人工序列(Artificial Sequence)

<400> 28

aaacctactg ctcgctcttg atca 24

<210> 29

<211> 1904

<212> PRT

<213> 人工序列(Artificial Sequence)

<400> 29

Met Lys Arg Thr Ala Asp Gly Ser Glu Phe Glu Ser Pro Lys Lys Lys

1 5 10 15

Arg Lys Val Asp Lys Lys Tyr Ser Ile Gly Leu Ala Ile Gly Thr Asn

20 25 30

Ser Val Gly Trp Ala Val Ile Thr Asp Glu Tyr Lys Val Pro Ser Lys

35 40 45

Lys Phe Lys Val Leu Gly Asn Thr Asp Arg His Ser Ile Lys Lys Asn

50 55 60

Leu Ile Gly Ala Leu Leu Phe Asp Ser Gly Glu Thr Ala Glu Ala Thr

65 70 75 80

Arg Leu Lys Arg Thr Ala Arg Arg Arg Tyr Thr Arg Arg Lys Asn Arg

85 90 95

Ile Cys Tyr Leu Gln Glu Ile Phe Ser Asn Glu Met Ala Lys Val Asp

100 105 110

Asp Ser Phe Phe His Arg Leu Glu Glu Ser Phe Leu Val Glu Glu Asp

115 120 125

Lys Lys His Glu Arg His Pro Ile Phe Gly Asn Ile Val Asp Glu Val

130 135 140

Ala Tyr His Glu Lys Tyr Pro Thr Ile Tyr His Leu Arg Lys Lys Leu

145 150 155 160

Val Asp Ser Thr Asp Lys Ala Asp Leu Arg Leu Ile Tyr Leu Ala Leu

165 170 175

Ala His Met Ile Lys Phe Arg Gly His Phe Leu Ile Glu Gly Asp Leu

180 185 190

Asn Pro Asp Asn Ser Asp Val Asp Lys Leu Phe Ile Gln Leu Val Gln

195 200 205

Thr Tyr Asn Gln Leu Phe Glu Glu Asn Pro Ile Asn Ala Ser Gly Val

210 215 220

Asp Ala Lys Ala Ile Leu Ser Ala Arg Leu Ser Lys Ser Arg Arg Leu

225 230 235 240

Glu Asn Leu Ile Ala Gln Leu Pro Gly Glu Lys Lys Asn Gly Leu Phe

245 250 255

Gly Asn Leu Ile Ala Leu Ser Leu Gly Leu Thr Pro Asn Phe Lys Ser

260 265 270

Asn Phe Asp Leu Ala Glu Asp Ala Lys Leu Gln Leu Ser Lys Asp Thr

275 280 285

Tyr Asp Asp Asp Leu Asp Asn Leu Leu Ala Gln Ile Gly Asp Gln Tyr

290 295 300

Ala Asp Leu Phe Leu Ala Ala Lys Asn Leu Ser Asp Ala Ile Leu Leu

305 310 315 320

Ser Asp Ile Leu Arg Val Asn Thr Glu Ile Thr Lys Ala Pro Leu Ser

325 330 335

Ala Ser Met Ile Lys Arg Tyr Asp Glu His His Gln Asp Leu Thr Leu

340 345 350

Leu Lys Ala Leu Val Arg Gln Gln Leu Pro Glu Lys Tyr Lys Glu Ile

355 360 365

Phe Phe Asp Gln Ser Lys Asn Gly Tyr Ala Gly Tyr Ile Asp Gly Gly

370 375 380

Ala Ser Gln Glu Glu Phe Tyr Lys Phe Ile Lys Pro Ile Leu Glu Lys

385 390 395 400

Met Asp Gly Thr Glu Glu Leu Leu Val Lys Leu Asn Arg Glu Asp Leu

405 410 415

Leu Arg Lys Gln Arg Thr Phe Asp Asn Gly Ser Ile Pro His Gln Ile

420 425 430

His Leu Gly Glu Leu His Ala Ile Leu Arg Arg Gln Glu Asp Phe Tyr

435 440 445

Pro Phe Leu Lys Asp Asn Arg Glu Lys Ile Glu Lys Ile Leu Thr Phe

450 455 460

Arg Ile Pro Tyr Tyr Val Gly Pro Leu Ala Arg Gly Asn Ser Arg Phe

465 470 475 480

Ala Trp Met Thr Arg Lys Ser Glu Glu Thr Ile Thr Pro Trp Asn Phe

485 490 495

Glu Glu Val Val Asp Lys Gly Ala Ser Ala Gln Ser Phe Ile Glu Arg

500 505 510

Met Thr Asn Phe Asp Lys Asn Leu Pro Asn Glu Lys Val Leu Pro Lys

515 520 525

His Ser Leu Leu Tyr Glu Tyr Phe Thr Val Tyr Asn Glu Leu Thr Lys

530 535 540

Val Lys Tyr Val Thr Glu Gly Met Arg Lys Pro Ala Phe Leu Ser Gly

545 550 555 560

Glu Gln Lys Lys Ala Ile Val Asp Leu Leu Phe Lys Thr Asn Arg Lys

565 570 575

Val Thr Val Lys Gln Leu Lys Glu Asp Tyr Phe Lys Lys Ile Glu Cys

580 585 590

Phe Asp Ser Val Glu Ile Ser Gly Val Glu Asp Arg Phe Asn Ala Ser

595 600 605

Leu Gly Thr Tyr His Asp Leu Leu Lys Ile Ile Lys Asp Lys Asp Phe

610 615 620

Leu Asp Asn Glu Glu Asn Glu Asp Ile Leu Glu Asp Ile Val Leu Thr

625 630 635 640

Leu Thr Leu Phe Glu Asp Arg Glu Met Ile Glu Glu Arg Leu Lys Thr

645 650 655

Tyr Ala His Leu Phe Asp Asp Lys Val Met Lys Gln Leu Lys Arg Arg

660 665 670

Arg Tyr Thr Gly Trp Gly Arg Leu Ser Arg Lys Leu Ile Asn Gly Ile

675 680 685

Arg Asp Lys Gln Ser Gly Lys Thr Ile Leu Asp Phe Leu Lys Ser Asp

690 695 700

Gly Phe Ala Asn Arg Asn Phe Met Gln Leu Ile His Asp Asp Ser Leu

705 710 715 720

Thr Phe Lys Glu Asp Ile Gln Lys Ala Gln Val Ser Gly Gln Gly Asp

725 730 735

Ser Leu His Glu His Ile Ala Asn Leu Ala Gly Ser Pro Ala Ile Lys

740 745 750

Lys Gly Ile Leu Gln Thr Val Lys Val Val Asp Glu Leu Val Lys Val

755 760 765

Met Gly Arg His Lys Pro Glu Asn Ile Val Ile Glu Met Ala Arg Glu

770 775 780

Asn Gln Thr Thr Gln Lys Gly Gln Lys Asn Ser Arg Glu Arg Met Lys

785 790 795 800

Arg Ile Glu Glu Gly Ile Lys Glu Leu Gly Ser Gln Ile Leu Lys Glu

805 810 815

His Pro Val Glu Asn Thr Gln Leu Gln Asn Glu Lys Leu Tyr Leu Tyr

820 825 830

Tyr Leu Gln Asn Gly Arg Asp Met Tyr Val Asp Gln Glu Leu Asp Ile

835 840 845

Asn Arg Leu Ser Asp Tyr Asp Val Asp His Ile Val Pro Gln Ser Phe

850 855 860

Leu Lys Asp Asp Ser Ile Asp Asn Lys Val Leu Thr Arg Ser Asp Lys

865 870 875 880

Asn Arg Gly Lys Ser Asp Asn Val Pro Ser Glu Glu Val Val Lys Lys

885 890 895

Met Lys Asn Tyr Trp Arg Gln Leu Leu Asn Ala Lys Leu Ile Thr Gln

900 905 910

Arg Lys Phe Asp Asn Leu Thr Lys Ala Glu Arg Gly Gly Leu Ser Glu

915 920 925

Leu Asp Lys Ala Gly Phe Ile Lys Arg Gln Leu Val Glu Thr Arg Gln

930 935 940

Ile Thr Lys His Val Ala Gln Ile Leu Asp Ser Arg Met Asn Thr Lys

945 950 955 960

Tyr Asp Glu Asn Asp Lys Leu Ile Arg Glu Val Lys Val Ile Thr Leu

965 970 975

Lys Ser Lys Leu Val Ser Asp Phe Arg Lys Asp Phe Gln Phe Tyr Lys

980 985 990

Val Arg Glu Ile Asn Asn Tyr His His Ala His Asp Ala Tyr Leu Asn

995 1000 1005

Ala Val Val Gly Thr Ala Leu Ile Lys Lys Tyr Pro Lys Leu Glu Ser

1010 1015 1020

Glu Phe Val Tyr Gly Asp Tyr Lys Val Tyr Asp Val Arg Lys Met Ile

1025 1030 1035 1040

Ala Lys Ser Glu Gln Ser Gly Ser Glu Thr Pro Gly Thr Ser Glu Ser

1045 1050 1055

Ala Thr Pro Glu Ser Gly Ser Glu Thr Gly Pro Val Ala Val Asp Pro

1060 1065 1070

Thr Leu Arg Arg Arg Ile Glu Pro His Glu Phe Glu Val Phe Phe Asp

1075 1080 1085

Pro Arg Glu Leu Arg Lys Glu Thr Cys Leu Leu Tyr Glu Ile Lys Trp

1090 1095 1100

Gly Thr Ser His Lys Ile Trp Arg His Ser Ser Lys Asn Thr Thr Lys

1105 1110 1115 1120

His Val Glu Val Asn Phe Ile Glu Lys Phe Thr Ser Glu Arg His Phe

1125 1130 1135

Cys Pro Ser Thr Ser Cys Ser Ile Thr Trp Phe Leu Ser Trp Ser Pro

1140 1145 1150

Cys Gly Glu Cys Ser Lys Ala Ile Thr Glu Phe Leu Ser Gln His Pro

1155 1160 1165

Asn Val Thr Leu Val Ile Tyr Val Ala Arg Leu Tyr His His Met Asp

1170 1175 1180

Gln Gln Asn Arg Gln Gly Leu Arg Asp Leu Val Asn Ser Gly Val Thr

1185 1190 1195 1200

Ile Gln Ile Met Thr Ala Pro Glu Tyr Asp Tyr Cys Trp Arg Asn Phe

1205 1210 1215

Val Asn Tyr Pro Pro Gly Lys Glu Ala His Trp Pro Arg Tyr Pro Pro

1220 1225 1230

Leu Trp Met Lys Leu Tyr Ala Leu Glu Leu His Ala Gly Ile Leu Gly

1235 1240 1245

Leu Pro Pro Cys Leu Asn Ile Leu Arg Arg Lys Gln Pro Gln Leu Thr

1250 1255 1260

Phe Phe Thr Ile Ala Leu Gln Ser Cys His Tyr Gln Arg Leu Pro Pro

1265 1270 1275 1280

His Ile Leu Trp Ala Thr Gly Leu Lys Ser Gly Gly Ser Gly Gly Ser

1285 1290 1295

Gly Gly Ser Met Ala Ala Ala Lys Ala Pro Gly Ala Ala Glu Phe Val

1300 1305 1310

Pro Ala Asp Ala Asp Leu Asp Thr Leu Arg Thr Ala Val Gln Gly Cys

1315 1320 1325

Arg Gly Cys Glu Leu Tyr Arg Gly Ala Thr Gln Ala Val Phe Gly Glu

1330 1335 1340

Gly Pro Ala His Ala Pro Val Phe Val Val Gly Glu Gln Pro Gly Asp

1345 1350 1355 1360

Arg Glu Asp Val Ala Gly His Pro Phe Val Gly Pro Ala Gly Arg Leu

1365 1370 1375

Leu Asp Lys Ala Leu Thr Glu Ala Asp Ile Asp Arg Glu Ala Val Tyr

1380 1385 1390

Leu Thr Asn Ala Val Lys His Phe Lys Phe Glu Glu Arg Gly Lys Arg

1395 1400 1405

Arg Ile His Lys Gln Pro Gly Arg Thr Glu Val Val Ala Cys Ser Pro

1410 1415 1420

Trp Leu Thr Ala Glu Leu Asp Ala Val Arg Pro Gln Leu Val Val Cys

1425 1430 1435 1440

Leu Gly Ala Val Ala Ala Lys Ala Val Leu Gly Pro Ser Phe Lys Val

1445 1450 1455

Ser Glu Arg Arg Gly Glu Val Val Glu Ala Gly Glu His Arg Val Ile

1460 1465 1470

Ala Thr Val His Pro Ser Ser Val Leu Arg Ala Pro Asp Arg Ala Ala

1475 1480 1485

Ala Tyr Ala Asp Phe Leu Ala Asp Leu Arg Lys Val Arg Thr Ala Ala

1490 1495 1500

Gly Glu Leu His Arg Ala Ser Gly Gly Ser Ser Gly Gly Ser Ser Gly

1505 1510 1515 1520

Ser Glu Thr Pro Gly Thr Ser Glu Ser Ala Thr Pro Glu Ser Ser Gly

1525 1530 1535

Gly Ser Ser Gly Gly Ser Glu Ile Gly Lys Ala Thr Ala Lys Tyr Phe

1540 1545 1550

Phe Tyr Ser Asn Ile Met Asn Phe Phe Lys Thr Glu Ile Thr Leu Ala

1555 1560 1565

Asn Gly Glu Ile Arg Lys Arg Pro Leu Ile Glu Thr Asn Gly Glu Thr

1570 1575 1580

Gly Glu Ile Val Trp Asp Lys Gly Arg Asp Phe Ala Thr Val Arg Lys

1585 1590 1595 1600

Val Leu Ser Met Pro Gln Val Asn Ile Val Lys Lys Thr Glu Val Gln

1605 1610 1615

Thr Gly Gly Phe Ser Lys Glu Ser Ile Leu Pro Lys Arg Asn Ser Asp

1620 1625 1630

Lys Leu Ile Ala Arg Lys Lys Asp Trp Asp Pro Lys Lys Tyr Gly Gly

1635 1640 1645

Phe Asp Ser Pro Thr Val Ala Tyr Ser Val Leu Val Val Ala Lys Val

1650 1655 1660

Glu Lys Gly Lys Ser Lys Lys Leu Lys Ser Val Lys Glu Leu Leu Gly

1665 1670 1675 1680

Ile Thr Ile Met Glu Arg Ser Ser Phe Glu Lys Asn Pro Ile Asp Phe

1685 1690 1695

Leu Glu Ala Lys Gly Tyr Lys Glu Val Lys Lys Asp Leu Ile Ile Lys

1700 1705 1710

Leu Pro Lys Tyr Ser Leu Phe Glu Leu Glu Asn Gly Arg Lys Arg Met

1715 1720 1725

Leu Ala Ser Ala Gly Glu Leu Gln Lys Gly Asn Glu Leu Ala Leu Pro

1730 1735 1740

Ser Lys Tyr Val Asn Phe Leu Tyr Leu Ala Ser His Tyr Glu Lys Leu

1745 1750 1755 1760

Lys Gly Ser Pro Glu Asp Asn Glu Gln Lys Gln Leu Phe Val Glu Gln

1765 1770 1775

His Lys His Tyr Leu Asp Glu Ile Ile Glu Gln Ile Ser Glu Phe Ser

1780 1785 1790

Lys Arg Val Ile Leu Ala Asp Ala Asn Leu Asp Lys Val Leu Ser Ala

1795 1800 1805

Tyr Asn Lys His Arg Asp Lys Pro Ile Arg Glu Gln Ala Glu Asn Ile

1810 1815 1820

Ile His Leu Phe Thr Leu Thr Asn Leu Gly Ala Pro Ala Ala Phe Lys

1825 1830 1835 1840

Tyr Phe Asp Thr Thr Ile Asp Arg Lys Arg Tyr Thr Ser Thr Lys Glu

1845 1850 1855

Val Leu Asp Ala Thr Leu Ile His Gln Ser Ile Thr Gly Leu Tyr Glu

1860 1865 1870

Thr Arg Ile Asp Leu Ser Gln Leu Gly Gly Asp Ser Gly Gly Ser Lys

1875 1880 1885

Arg Thr Ala Asp Gly Ser Glu Phe Glu Pro Lys Lys Lys Arg Lys Val

1890 1895 1900

<210> 30

<211> 56

<212> DNA

<213> 人工序列(Artificial Sequence)

<400> 30

cctggcccac cctcgtgacc accctgaccc atggcgtgca gtgcttcagc cgctac 56

<210> 31

<211> 56

<212> DNA

<213> 人工序列(Artificial Sequence)

<400> 31

cctggcccac cctcgtgacc accctgaccg atggcgtgca gtgcttcagc cgctac 56

<210> 32

<211> 56

<212> DNA

<213> 人工序列(Artificial Sequence)

<400> 32

cctggcccac cctcgtgacc accctgacct atggcgtgca gtgcttcagc cgctac 56

<210> 33

<211> 56

<212> DNA

<213> 人工序列(Artificial Sequence)

<400> 33

cctggcccac cctcgtgacc accctgacca atggcgtgca gtgcttcagc cgctac 56

<210> 34

<211> 1003

<212> PRT

<213> 人工序列(Artificial Sequence)

<400> 34

Met Asp Lys Lys Tyr Ser Ile Gly Leu Ala Ile Gly Thr Asn Ser Val

1 5 10 15

Gly Trp Ala Val Ile Thr Asp Glu Tyr Lys Val Pro Ser Lys Lys Phe

20 25 30

Lys Val Leu Gly Asn Thr Asp Arg His Ser Ile Lys Lys Asn Leu Ile

35 40 45

Gly Ala Leu Leu Phe Asp Ser Gly Glu Thr Ala Glu Ala Thr Arg Leu

50 55 60

Lys Arg Thr Ala Arg Arg Arg Tyr Thr Arg Arg Lys Asn Arg Ile Cys

65 70 75 80

Tyr Leu Gln Glu Ile Phe Ser Asn Glu Met Ala Lys Val Asp Asp Ser

85 90 95

Phe Phe His Arg Leu Glu Glu Ser Phe Leu Val Glu Glu Asp Lys Lys

100 105 110

His Glu Arg His Pro Ile Phe Gly Asn Ile Val Asp Glu Val Ala Tyr

115 120 125

His Glu Lys Tyr Pro Thr Ile Tyr His Leu Arg Lys Lys Leu Val Asp

130 135 140

Ser Thr Asp Lys Ala Asp Leu Arg Leu Ile Tyr Leu Ala Leu Ala His

145 150 155 160

Met Ile Lys Phe Arg Gly His Phe Leu Ile Glu Gly Asp Leu Asn Pro

165 170 175

Asp Asn Ser Asp Val Asp Lys Leu Phe Ile Gln Leu Val Gln Thr Tyr

180 185 190

Asn Gln Leu Phe Glu Glu Asn Pro Ile Asn Ala Ser Gly Val Asp Ala

195 200 205

Lys Ala Ile Leu Ser Ala Arg Leu Ser Lys Ser Arg Arg Leu Glu Asn

210 215 220

Leu Ile Ala Gln Leu Pro Gly Glu Lys Lys Asn Gly Leu Phe Gly Asn

225 230 235 240

Leu Ile Ala Leu Ser Leu Gly Leu Thr Pro Asn Phe Lys Ser Asn Phe

245 250 255

Asp Leu Ala Glu Asp Ala Lys Leu Gln Leu Ser Lys Asp Thr Tyr Asp

260 265 270

Asp Asp Leu Asp Asn Leu Leu Ala Gln Ile Gly Asp Gln Tyr Ala Asp

275 280 285

Leu Phe Leu Ala Ala Lys Asn Leu Ser Asp Ala Ile Leu Leu Ser Asp

290 295 300

Ile Leu Arg Val Asn Thr Glu Ile Thr Lys Ala Pro Leu Ser Ala Ser

305 310 315 320

Met Ile Lys Arg Tyr Asp Glu His His Gln Asp Leu Thr Leu Leu Lys

325 330 335

Ala Leu Val Arg Gln Gln Leu Pro Glu Lys Tyr Lys Glu Ile Phe Phe

340 345 350

Asp Gln Ser Lys Asn Gly Tyr Ala Gly Tyr Ile Asp Gly Gly Ala Ser

355 360 365

Gln Glu Glu Phe Tyr Lys Phe Ile Lys Pro Ile Leu Glu Lys Met Asp

370 375 380

Gly Thr Glu Glu Leu Leu Val Lys Leu Asn Arg Glu Asp Leu Leu Arg

385 390 395 400

Lys Gln Arg Thr Phe Asp Asn Gly Ser Ile Pro His Gln Ile His Leu

405 410 415

Gly Glu Leu His Ala Ile Leu Arg Arg Gln Glu Asp Phe Tyr Pro Phe

420 425 430

Leu Lys Asp Asn Arg Glu Lys Ile Glu Lys Ile Leu Thr Phe Arg Ile

435 440 445

Pro Tyr Tyr Val Gly Pro Leu Ala Arg Gly Asn Ser Arg Phe Ala Trp

450 455 460

Met Thr Arg Lys Ser Glu Glu Thr Ile Thr Pro Trp Asn Phe Glu Glu

465 470 475 480

Val Val Asp Lys Gly Ala Ser Ala Gln Ser Phe Ile Glu Arg Met Thr

485 490 495

Asn Phe Asp Lys Asn Leu Pro Asn Glu Lys Val Leu Pro Lys His Ser

500 505 510

Leu Leu Tyr Glu Tyr Phe Thr Val Tyr Asn Glu Leu Thr Lys Val Lys

515 520 525

Tyr Val Thr Glu Gly Met Arg Lys Pro Ala Phe Leu Ser Gly Glu Gln

530 535 540

Lys Lys Ala Ile Val Asp Leu Leu Phe Lys Thr Asn Arg Lys Val Thr

545 550 555 560

Val Lys Gln Leu Lys Glu Asp Tyr Phe Lys Lys Ile Glu Cys Phe Asp

565 570 575

Ser Val Glu Ile Ser Gly Val Glu Asp Arg Phe Asn Ala Ser Leu Gly

580 585 590

Thr Tyr His Asp Leu Leu Lys Ile Ile Lys Asp Lys Asp Phe Leu Asp

595 600 605

Asn Glu Glu Asn Glu Asp Ile Leu Glu Asp Ile Val Leu Thr Leu Thr

610 615 620

Leu Phe Glu Asp Arg Glu Met Ile Glu Glu Arg Leu Lys Thr Tyr Ala

625 630 635 640

His Leu Phe Asp Asp Lys Val Met Lys Gln Leu Lys Arg Arg Arg Tyr

645 650 655

Thr Gly Trp Gly Arg Leu Ser Arg Lys Leu Ile Asn Gly Ile Arg Asp

660 665 670

Lys Gln Ser Gly Lys Thr Ile Leu Asp Phe Leu Lys Ser Asp Gly Phe

675 680 685

Ala Asn Arg Asn Phe Met Gln Leu Ile His Asp Asp Ser Leu Thr Phe

690 695 700

Lys Glu Asp Ile Gln Lys Ala Gln Val Ser Gly Gln Gly Asp Ser Leu

705 710 715 720

His Glu His Ile Ala Asn Leu Ala Gly Ser Pro Ala Ile Lys Lys Gly

725 730 735

Ile Leu Gln Thr Val Lys Val Val Asp Glu Leu Val Lys Val Met Gly

740 745 750

Arg His Lys Pro Glu Asn Ile Val Ile Glu Met Ala Arg Glu Asn Gln

755 760 765

Thr Thr Gln Lys Gly Gln Lys Asn Ser Arg Glu Arg Met Lys Arg Ile

770 775 780

Glu Glu Gly Ile Lys Glu Leu Gly Ser Gln Ile Leu Lys Glu His Pro

785 790 795 800

Val Glu Asn Thr Gln Leu Gln Asn Glu Lys Leu Tyr Leu Tyr Tyr Leu

805 810 815

Gln Asn Gly Arg Asp Met Tyr Val Asp Gln Glu Leu Asp Ile Asn Arg

820 825 830

Leu Ser Asp Tyr Asp Val Asp His Ile Val Pro Gln Ser Phe Leu Lys

835 840 845

Asp Asp Ser Ile Asp Asn Lys Val Leu Thr Arg Ser Asp Lys Asn Arg

850 855 860

Gly Lys Ser Asp Asn Val Pro Ser Glu Glu Val Val Lys Lys Met Lys

865 870 875 880

Asn Tyr Trp Arg Gln Leu Leu Asn Ala Lys Leu Ile Thr Gln Arg Lys

885 890 895

Phe Asp Asn Leu Thr Lys Ala Glu Arg Gly Gly Leu Ser Glu Leu Asp

900 905 910

Lys Ala Gly Phe Ile Lys Arg Gln Leu Val Glu Thr Arg Gln Ile Thr

915 920 925

Lys His Val Ala Gln Ile Leu Asp Ser Arg Met Asn Thr Lys Tyr Asp

930 935 940

Glu Asn Asp Lys Leu Ile Arg Glu Val Lys Val Ile Thr Leu Lys Ser

945 950 955 960

Lys Leu Val Ser Asp Phe Arg Lys Asp Phe Gln Phe Tyr Lys Val Arg

965 970 975

Glu Ile Asn Asn Tyr His His Ala His Asp Ala Tyr Leu Asn Ala Val

980 985 990

Val Gly Thr Ala Leu Ile Lys Lys Tyr Pro Lys

995 1000

<210> 35

<211> 1027

<212> PRT

<213> 人工序列(Artificial Sequence)

<400> 35

Met Asp Lys Lys Tyr Ser Ile Gly Leu Ala Ile Gly Thr Asn Ser Val

1 5 10 15

Gly Trp Ala Val Ile Thr Asp Glu Tyr Lys Val Pro Ser Lys Lys Phe

20 25 30

Lys Val Leu Gly Asn Thr Asp Arg His Ser Ile Lys Lys Asn Leu Ile

35 40 45

Gly Ala Leu Leu Phe Asp Ser Gly Glu Thr Ala Glu Ala Thr Arg Leu

50 55 60

Lys Arg Thr Ala Arg Arg Arg Tyr Thr Arg Arg Lys Asn Arg Ile Cys

65 70 75 80

Tyr Leu Gln Glu Ile Phe Ser Asn Glu Met Ala Lys Val Asp Asp Ser

85 90 95

Phe Phe His Arg Leu Glu Glu Ser Phe Leu Val Glu Glu Asp Lys Lys

100 105 110

His Glu Arg His Pro Ile Phe Gly Asn Ile Val Asp Glu Val Ala Tyr

115 120 125

His Glu Lys Tyr Pro Thr Ile Tyr His Leu Arg Lys Lys Leu Val Asp

130 135 140

Ser Thr Asp Lys Ala Asp Leu Arg Leu Ile Tyr Leu Ala Leu Ala His

145 150 155 160

Met Ile Lys Phe Arg Gly His Phe Leu Ile Glu Gly Asp Leu Asn Pro

165 170 175

Asp Asn Ser Asp Val Asp Lys Leu Phe Ile Gln Leu Val Gln Thr Tyr

180 185 190

Asn Gln Leu Phe Glu Glu Asn Pro Ile Asn Ala Ser Gly Val Asp Ala

195 200 205

Lys Ala Ile Leu Ser Ala Arg Leu Ser Lys Ser Arg Arg Leu Glu Asn

210 215 220

Leu Ile Ala Gln Leu Pro Gly Glu Lys Lys Asn Gly Leu Phe Gly Asn

225 230 235 240

Leu Ile Ala Leu Ser Leu Gly Leu Thr Pro Asn Phe Lys Ser Asn Phe

245 250 255

Asp Leu Ala Glu Asp Ala Lys Leu Gln Leu Ser Lys Asp Thr Tyr Asp

260 265 270

Asp Asp Leu Asp Asn Leu Leu Ala Gln Ile Gly Asp Gln Tyr Ala Asp

275 280 285

Leu Phe Leu Ala Ala Lys Asn Leu Ser Asp Ala Ile Leu Leu Ser Asp

290 295 300

Ile Leu Arg Val Asn Thr Glu Ile Thr Lys Ala Pro Leu Ser Ala Ser

305 310 315 320

Met Ile Lys Arg Tyr Asp Glu His His Gln Asp Leu Thr Leu Leu Lys

325 330 335

Ala Leu Val Arg Gln Gln Leu Pro Glu Lys Tyr Lys Glu Ile Phe Phe

340 345 350

Asp Gln Ser Lys Asn Gly Tyr Ala Gly Tyr Ile Asp Gly Gly Ala Ser

355 360 365

Gln Glu Glu Phe Tyr Lys Phe Ile Lys Pro Ile Leu Glu Lys Met Asp

370 375 380

Gly Thr Glu Glu Leu Leu Val Lys Leu Asn Arg Glu Asp Leu Leu Arg

385 390 395 400

Lys Gln Arg Thr Phe Asp Asn Gly Ser Ile Pro His Gln Ile His Leu

405 410 415

Gly Glu Leu His Ala Ile Leu Arg Arg Gln Glu Asp Phe Tyr Pro Phe

420 425 430

Leu Lys Asp Asn Arg Glu Lys Ile Glu Lys Ile Leu Thr Phe Arg Ile

435 440 445

Pro Tyr Tyr Val Gly Pro Leu Ala Arg Gly Asn Ser Arg Phe Ala Trp

450 455 460

Met Thr Arg Lys Ser Glu Glu Thr Ile Thr Pro Trp Asn Phe Glu Glu

465 470 475 480

Val Val Asp Lys Gly Ala Ser Ala Gln Ser Phe Ile Glu Arg Met Thr

485 490 495

Asn Phe Asp Lys Asn Leu Pro Asn Glu Lys Val Leu Pro Lys His Ser

500 505 510

Leu Leu Tyr Glu Tyr Phe Thr Val Tyr Asn Glu Leu Thr Lys Val Lys

515 520 525

Tyr Val Thr Glu Gly Met Arg Lys Pro Ala Phe Leu Ser Gly Glu Gln

530 535 540

Lys Lys Ala Ile Val Asp Leu Leu Phe Lys Thr Asn Arg Lys Val Thr

545 550 555 560

Val Lys Gln Leu Lys Glu Asp Tyr Phe Lys Lys Ile Glu Cys Phe Asp

565 570 575

Ser Val Glu Ile Ser Gly Val Glu Asp Arg Phe Asn Ala Ser Leu Gly

580 585 590

Thr Tyr His Asp Leu Leu Lys Ile Ile Lys Asp Lys Asp Phe Leu Asp

595 600 605

Asn Glu Glu Asn Glu Asp Ile Leu Glu Asp Ile Val Leu Thr Leu Thr

610 615 620

Leu Phe Glu Asp Arg Glu Met Ile Glu Glu Arg Leu Lys Thr Tyr Ala

625 630 635 640

His Leu Phe Asp Asp Lys Val Met Lys Gln Leu Lys Arg Arg Arg Tyr

645 650 655

Thr Gly Trp Gly Arg Leu Ser Arg Lys Leu Ile Asn Gly Ile Arg Asp

660 665 670

Lys Gln Ser Gly Lys Thr Ile Leu Asp Phe Leu Lys Ser Asp Gly Phe

675 680 685

Ala Asn Arg Asn Phe Met Gln Leu Ile His Asp Asp Ser Leu Thr Phe

690 695 700

Lys Glu Asp Ile Gln Lys Ala Gln Val Ser Gly Gln Gly Asp Ser Leu

705 710 715 720

His Glu His Ile Ala Asn Leu Ala Gly Ser Pro Ala Ile Lys Lys Gly

725 730 735

Ile Leu Gln Thr Val Lys Val Val Asp Glu Leu Val Lys Val Met Gly

740 745 750

Arg His Lys Pro Glu Asn Ile Val Ile Glu Met Ala Arg Glu Asn Gln

755 760 765

Thr Thr Gln Lys Gly Gln Lys Asn Ser Arg Glu Arg Met Lys Arg Ile

770 775 780

Glu Glu Gly Ile Lys Glu Leu Gly Ser Gln Ile Leu Lys Glu His Pro

785 790 795 800

Val Glu Asn Thr Gln Leu Gln Asn Glu Lys Leu Tyr Leu Tyr Tyr Leu

805 810 815

Gln Asn Gly Arg Asp Met Tyr Val Asp Gln Glu Leu Asp Ile Asn Arg

820 825 830

Leu Ser Asp Tyr Asp Val Asp His Ile Val Pro Gln Ser Phe Leu Lys

835 840 845

Asp Asp Ser Ile Asp Asn Lys Val Leu Thr Arg Ser Asp Lys Asn Arg

850 855 860

Gly Lys Ser Asp Asn Val Pro Ser Glu Glu Val Val Lys Lys Met Lys

865 870 875 880

Asn Tyr Trp Arg Gln Leu Leu Asn Ala Lys Leu Ile Thr Gln Arg Lys

885 890 895

Phe Asp Asn Leu Thr Lys Ala Glu Arg Gly Gly Leu Ser Glu Leu Asp

900 905 910

Lys Ala Gly Phe Ile Lys Arg Gln Leu Val Glu Thr Arg Gln Ile Thr

915 920 925

Lys His Val Ala Gln Ile Leu Asp Ser Arg Met Asn Thr Lys Tyr Asp

930 935 940

Glu Asn Asp Lys Leu Ile Arg Glu Val Lys Val Ile Thr Leu Lys Ser

945 950 955 960

Lys Leu Val Ser Asp Phe Arg Lys Asp Phe Gln Phe Tyr Lys Val Arg

965 970 975

Glu Ile Asn Asn Tyr His His Ala His Asp Ala Tyr Leu Asn Ala Val

980 985 990

Val Gly Thr Ala Leu Ile Lys Lys Tyr Pro Lys Leu Glu Ser Glu Phe

995 1000 1005

Val Tyr Gly Asp Tyr Lys Val Tyr Asp Val Arg Lys Met Ile Ala Lys

1010 1015 1020

Ser Glu Gln

1025

<210> 36

<211> 1038

<212> PRT

<213> 人工序列(Artificial Sequence)

<400> 36

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 Ser

1010 1015 1020

Glu Gln Glu Ile Gly Lys Ala Thr Ala Lys Tyr Phe Phe Tyr

1025 1030 1035

<210> 37

<211> 365

<212> PRT

<213> 人工序列(Artificial Sequence)

<400> 37

Leu Glu Ser Glu Phe Val Tyr Gly Asp Tyr Lys Val Tyr Asp Val Arg

1 5 10 15

Lys Met Ile Ala Lys Ser Glu Gln Glu Ile Gly Lys Ala Thr Ala Lys

20 25 30

Tyr Phe Phe Tyr Ser Asn Ile Met Asn Phe Phe Lys Thr Glu Ile Thr

35 40 45

Leu Ala Asn Gly Glu Ile Arg Lys Arg Pro Leu Ile Glu Thr Asn Gly

50 55 60

Glu Thr Gly Glu Ile Val Trp Asp Lys Gly Arg Asp Phe Ala Thr Val

65 70 75 80

Arg Lys Val Leu Ser Met Pro Gln Val Asn Ile Val Lys Lys Thr Glu

85 90 95

Val Gln Thr Gly Gly Phe Ser Lys Glu Ser Ile Leu Pro Lys Arg Asn

100 105 110

Ser Asp Lys Leu Ile Ala Arg Lys Lys Asp Trp Asp Pro Lys Lys Tyr

115 120 125

Gly Gly Phe Asp Ser Pro Thr Val Ala Tyr Ser Val Leu Val Val Ala

130 135 140

Lys Val Glu Lys Gly Lys Ser Lys Lys Leu Lys Ser Val Lys Glu Leu

145 150 155 160

Leu Gly Ile Thr Ile Met Glu Arg Ser Ser Phe Glu Lys Asn Pro Ile

165 170 175

Asp Phe Leu Glu Ala Lys Gly Tyr Lys Glu Val Lys Lys Asp Leu Ile

180 185 190

Ile Lys Leu Pro Lys Tyr Ser Leu Phe Glu Leu Glu Asn Gly Arg Lys

195 200 205

Arg Met Leu Ala Ser Ala Gly Glu Leu Gln Lys Gly Asn Glu Leu Ala

210 215 220

Leu Pro Ser Lys Tyr Val Asn Phe Leu Tyr Leu Ala Ser His Tyr Glu

225 230 235 240

Lys Leu Lys Gly Ser Pro Glu Asp Asn Glu Gln Lys Gln Leu Phe Val

245 250 255

Glu Gln His Lys His Tyr Leu Asp Glu Ile Ile Glu Gln Ile Ser Glu

260 265 270

Phe Ser Lys Arg Val Ile Leu Ala Asp Ala Asn Leu Asp Lys Val Leu

275 280 285

Ser Ala Tyr Asn Lys His Arg Asp Lys Pro Ile Arg Glu Gln Ala Glu

290 295 300

Asn Ile Ile His Leu Phe Thr Leu Thr Asn Leu Gly Ala Pro Ala Ala

305 310 315 320

Phe Lys Tyr Phe Asp Thr Thr Ile Asp Arg Lys Arg Tyr Thr Ser Thr

325 330 335

Lys Glu Val Leu Asp Ala Thr Leu Ile His Gln Ser Ile Thr Gly Leu

340 345 350

Tyr Glu Thr Arg Ile Asp Leu Ser Gln Leu Gly Gly Asp

355 360 365

<210> 38

<211> 341

<212> PRT

<213> 人工序列(Artificial Sequence)

<400> 38

Glu Ile Gly Lys Ala Thr Ala Lys Tyr Phe Phe Tyr Ser Asn Ile Met

1 5 10 15

Asn Phe Phe Lys Thr Glu Ile Thr Leu Ala Asn Gly Glu Ile Arg Lys

20 25 30

Arg Pro Leu Ile Glu Thr Asn Gly Glu Thr Gly Glu Ile Val Trp Asp

35 40 45

Lys Gly Arg Asp Phe Ala Thr Val Arg Lys Val Leu Ser Met Pro Gln

50 55 60

Val Asn Ile Val Lys Lys Thr Glu Val Gln Thr Gly Gly Phe Ser Lys

65 70 75 80

Glu Ser Ile Leu Pro Lys Arg Asn Ser Asp Lys Leu Ile Ala Arg Lys

85 90 95

Lys Asp Trp Asp Pro Lys Lys Tyr Gly Gly Phe Asp Ser Pro Thr Val

100 105 110

Ala Tyr Ser Val Leu Val Val Ala Lys Val Glu Lys Gly Lys Ser Lys

115 120 125

Lys Leu Lys Ser Val Lys Glu Leu Leu Gly Ile Thr Ile Met Glu Arg

130 135 140

Ser Ser Phe Glu Lys Asn Pro Ile Asp Phe Leu Glu Ala Lys Gly Tyr

145 150 155 160

Lys Glu Val Lys Lys Asp Leu Ile Ile Lys Leu Pro Lys Tyr Ser Leu

165 170 175

Phe Glu Leu Glu Asn Gly Arg Lys Arg Met Leu Ala Ser Ala Gly Glu

180 185 190

Leu Gln Lys Gly Asn Glu Leu Ala Leu Pro Ser Lys Tyr Val Asn Phe

195 200 205

Leu Tyr Leu Ala Ser His Tyr Glu Lys Leu Lys Gly Ser Pro Glu Asp

210 215 220

Asn Glu Gln Lys Gln Leu Phe Val Glu Gln His Lys His Tyr Leu Asp

225 230 235 240

Glu Ile Ile Glu Gln Ile Ser Glu Phe Ser Lys Arg Val Ile Leu Ala

245 250 255

Asp Ala Asn Leu Asp Lys Val Leu Ser Ala Tyr Asn Lys His Arg Asp

260 265 270

Lys Pro Ile Arg Glu Gln Ala Glu Asn Ile Ile His Leu Phe Thr Leu

275 280 285

Thr Asn Leu Gly Ala Pro Ala Ala Phe Lys Tyr Phe Asp Thr Thr Ile

290 295 300

Asp Arg Lys Arg Tyr Thr Ser Thr Lys Glu Val Leu Asp Ala Thr Leu

305 310 315 320

Ile His Gln Ser Ile Thr Gly Leu Tyr Glu Thr Arg Ile Asp Leu Ser

325 330 335

Gln Leu Gly Gly Asp

340

<210> 39

<211> 329

<212> PRT

<213> 人工序列(Artificial Sequence)

<400> 39

Ser Asn Ile Met Asn Phe Phe Lys Thr Glu Ile Thr Leu Ala Asn Gly

1 5 10 15

Glu Ile Arg Lys Arg Pro Leu Ile Glu Thr Asn Gly Glu Thr Gly Glu

20 25 30

Ile Val Trp Asp Lys Gly Arg Asp Phe Ala Thr Val Arg Lys Val Leu

35 40 45

Ser Met Pro Gln Val Asn Ile Val Lys Lys Thr Glu Val Gln Thr Gly

50 55 60

Gly Phe Ser Lys Glu Ser Ile Leu Pro Lys Arg Asn Ser Asp Lys Leu

65 70 75 80

Ile Ala Arg Lys Lys Asp Trp Asp Pro Lys Lys Tyr Gly Gly Phe Asp

85 90 95

Ser Pro Thr Val Ala Tyr Ser Val Leu Val Val Ala Lys Val Glu Lys

100 105 110

Gly Lys Ser Lys Lys Leu Lys Ser Val Lys Glu Leu Leu Gly Ile Thr

115 120 125

Ile Met Glu Arg Ser Ser Phe Glu Lys Asn Pro Ile Asp Phe Leu Glu

130 135 140

Ala Lys Gly Tyr Lys Glu Val Lys Lys Asp Leu Ile Ile Lys Leu Pro

145 150 155 160

Lys Tyr Ser Leu Phe Glu Leu Glu Asn Gly Arg Lys Arg Met Leu Ala

165 170 175

Ser Ala Gly Glu Leu Gln Lys Gly Asn Glu Leu Ala Leu Pro Ser Lys

180 185 190

Tyr Val Asn Phe Leu Tyr Leu Ala Ser His Tyr Glu Lys Leu Lys Gly

195 200 205

Ser Pro Glu Asp Asn Glu Gln Lys Gln Leu Phe Val Glu Gln His Lys

210 215 220

His Tyr Leu Asp Glu Ile Ile Glu Gln Ile Ser Glu Phe Ser Lys Arg

225 230 235 240

Val Ile Leu Ala Asp Ala Asn Leu Asp Lys Val Leu Ser Ala Tyr Asn

245 250 255

Lys His Arg Asp Lys Pro Ile Arg Glu Gln Ala Glu Asn Ile Ile His

260 265 270

Leu Phe Thr Leu Thr Asn Leu Gly Ala Pro Ala Ala Phe Lys Tyr Phe

275 280 285

Asp Thr Thr Ile Asp Arg Lys Arg Tyr Thr Ser Thr Lys Glu Val Leu

290 295 300

Asp Ala Thr Leu Ile His Gln Ser Ile Thr Gly Leu Tyr Glu Thr Arg

305 310 315 320

Ile Asp Leu Ser Gln Leu Gly Gly Asp

325

<210> 40

<211> 18

<212> PRT

<213> 人工序列(Artificial Sequence)

<400> 40

Ser Gly Ser Glu Thr Pro Gly Thr Ser Glu Ser Ala Thr Pro Glu Ser

1 5 10 15

Gly Ser

<210> 41

<211> 18

<212> PRT

<213> 人工序列(Artificial Sequence)

<400> 41

Ser Gly Ser Gly Ser Glu Thr Pro Gly Thr Ser Glu Ser Ala Thr Pro

1 5 10 15

Glu Ser

<210> 42

<211> 5631

<212> DNA

<213> 人工序列(Artificial Sequence)

<400> 42

atgaaacgga cagccgacgg aagcgagttc gagtcaccaa agaagaagcg gaaagtcgac 60

aagaagtaca gcatcggcct ggccatcggc accaactctg tgggctgggc cgtgatcacc 120

gacgagtaca aggtgcccag caagaaattc aaggtgctgg gcaacaccga ccggcacagc 180

atcaagaaga acctgatcgg agccctgctg ttcgacagcg gcgaaacagc cgaggccacc 240

cggctgaaga gaaccgccag aagaagatac accagacgga agaaccggat ctgctatctg 300

caagagatct tcagcaacga gatggccaag gtggacgaca gcttcttcca cagactggaa 360

gagtccttcc tggtggaaga ggataagaag cacgagcggc accccatctt cggcaacatc 420

gtggacgagg tggcctacca cgagaagtac cccaccatct accacctgag aaagaaactg 480

gtggacagca ccgacaaggc cgacctgcgg ctgatctatc tggccctggc ccacatgatc 540

aagttccggg gccacttcct gatcgagggc gacctgaacc ccgacaacag cgacgtggac 600

aagctgttca tccagctggt gcagacctac aaccagctgt tcgaggaaaa ccccatcaac 660

gccagcggcg tggacgccaa ggccatcctg tctgccagac tgagcaagag cagacggctg 720

gaaaatctga tcgcccagct gcccggcgag aagaagaatg gcctgttcgg aaacctgatt 780

gccctgagcc tgggcctgac ccccaacttc aagagcaact tcgacctggc cgaggatgcc 840

aaactgcagc tgagcaagga cacctacgac gacgacctgg acaacctgct ggcccagatc 900

ggcgaccagt acgccgacct gtttctggcc gccaagaacc tgtccgacgc catcctgctg 960

agcgacatcc tgagagtgaa caccgagatc accaaggccc ccctgagcgc ctctatgatc 1020

aagagatacg acgagcacca ccaggacctg accctgctga aagctctcgt gcggcagcag 1080

ctgcctgaga agtacaaaga gattttcttc gaccagagca agaacggcta cgccggctac 1140

attgacggcg gagccagcca ggaagagttc tacaagttca tcaagcccat cctggaaaag 1200

atggacggca ccgaggaact gctcgtgaag ctgaacagag aggacctgct gcggaagcag 1260

cggaccttcg acaacggcag catcccccac cagatccacc tgggagagct gcacgccatt 1320

ctgcggcggc aggaagattt ttacccattc ctgaaggaca accgggaaaa gatcgagaag 1380

atcctgacct tccgcatccc ctactacgtg ggccctctgg ccaggggaaa cagcagattc 1440

gcctggatga ccagaaagag cgaggaaacc atcaccccct ggaacttcga ggaagtggtg 1500

gacaagggcg cttccgccca gagcttcatc gagcggatga ccaacttcga taagaacctg 1560

cccaacgaga aggtgctgcc caagcacagc ctgctgtacg agtacttcac cgtgtataac 1620

gagctgacca aagtgaaata cgtgaccgag ggaatgagaa agcccgcctt cctgagcggc 1680

gagcagaaaa aggccatcgt ggacctgctg ttcaagacca accggaaagt gaccgtgaag 1740

cagctgaaag aggactactt caagaaaatc gagtgcttcg actccgtgga aatctccggc 1800

gtggaagatc ggttcaacgc ctccctgggc acataccacg atctgctgaa aattatcaag 1860

gacaaggact tcctggacaa tgaggaaaac gaggacattc tggaagatat cgtgctgacc 1920

ctgacactgt ttgaggacag agagatgatc gaggaacggc tgaaaaccta tgcccacctg 1980

ttcgacgaca aagtgatgaa gcagctgaag cggcggagat acaccggctg gggcaggctg 2040

agccggaagc tgatcaacgg catccgggac aagcagtccg gcaagacaat cctggatttc 2100

ctgaagtccg acggcttcgc caacagaaac ttcatgcagc tgatccacga cgacagcctg 2160

acctttaaag aggacatcca gaaagcccag gtgtccggcc agggcgatag cctgcacgag 2220

cacattgcca atctggccgg cagccccgcc attaagaagg gcatcctgca gacagtgaag 2280

gtggtggacg agctcgtgaa agtgatgggc cggcacaagc ccgagaacat cgtgatcgaa 2340

atggccagag agaaccagac cacccagaag ggacagaaga acagccgcga gagaatgaag 2400

cggatcgaag agggcatcaa agagctgggc agccagatcc tgaaagaaca ccccgtggaa 2460

aacacccagc tgcagaacga gaagctgtac ctgtactacc tgcagaatgg gcgggatatg 2520

tacgtggacc aggaactgga catcaaccgg ctgtccgact acgatgtgga ccatatcgtg 2580

cctcagagct ttctgaagga cgactccatc gacaacaagg tgctgaccag aagcgacaag 2640

aaccggggca agagcgacaa cgtgccctcc gaagaggtcg tgaagaagat gaagaactac 2700

tggcggcagc tgctgaacgc caagctgatt acccagagaa agttcgacaa tctgaccaag 2760

gccgagagag gcggcctgag cgaactggat aaggccggct tcatcaagag acagctggtg 2820

gaaacccggc agatcacaaa gcacgtggca cagatcctgg actcccggat gaacactaag 2880

tacgacgaga atgacaagct gatccgggaa gtgaaagtga tcaccctgaa gtccaagctg 2940

gtgtccgatt tccggaagga tttccagttt tacaaagtgc gcgagatcaa caactaccac 3000

cacgcccacg acgcctacct aaacgccgtc gtgggaaccg ccctgatcaa aaagtaccct 3060

aagtctggca gcgagacacc aggaacaagc gagtcagcaa caccagagag cggcagcgag 3120

gcatctccag caagcggacc aaggcacctg atggaccccc acatcttcac ctctaacttt 3180

aacaatggca tcggcaggca caagacatac ctgtgctatg aggtggagcg cctggacaat 3240

ggcaccagcg tgaagatgga tcagcacaga ggcttcctgc acaaccaggc caagaatctg 3300

ctgtgcggct tctacggccg gcacgcagag ctgagatttc tggacctggt gcctagcctg 3360

cagctggatc cagcccagat ctatagggtg acctggttca tcagctggtc cccatgcttt 3420

tcctggggat gtgcaggaga ggtgcgcgcc tttctgcagg agaacacaca cgtgcggctg 3480

agaatcttcg ccgcccggat ctttgactac gatcctctgt ataaggaggc cctgcagatg 3540

ctgagagacg caggagccca ggtgtccatc atgacctacg atgagttcaa gcactgctgg 3600

gacacatttg tggatcacca gggctgtccc ttccagcctt gggacggact ggatgagcac 3660

tcccaggccc tgtctggcag gctgagggcc atcctgcaga accagggcaa tagcggagga 3720

tccggaggat ctggaggcag catggccgct gctaaagccc ccggcgctgc tgaattcgtc 3780

cccgctgatg ccgacctgga tacactgcgg accgccgtgc agggctgcag aggctgtgaa 3840

ctgtacagag gcgccaccca ggccgtgttc ggcgagggcc ctgctcacgc ccctgtgttt 3900

gtggtcggcg agcagcctgg cgaccgggaa gatgtggccg gccacccctt cgtgggcccc 3960

gccggcagac tgctggacaa ggccctgaca gaggccgaca tcgacaggga agccgtgtac 4020

ctgaccaacg ccgttaagca cttcaagttc gaggaaagag gaaaaagaag aatccacaag 4080

caacctggca gaaccgaggt ggtggcatgc agcccttggc tgaccgccga gctggacgcc 4140

gtgcggcctc agctggtggt gtgcctgggc gccgtggccg ccaaggccgt gctgggacca 4200

tcttttaagg tgtccgagcg gcggggcgaa gtggtcgagg ccggagagca cagagtgatc 4260

gccacagtgc accctagcag cgtgctgaga gccccagacc gcgccgctgc ctacgccgac 4320

ttcctggccg atctgagaaa ggtgcggacc gccgctggag agctccatag agcctctgga 4380

ggatctagcg gaggatcctc tggcagcgag acaccaggaa caagcgagtc agcaacacca 4440

gagagcagtg gcggcagcag cggcggcagc ctggaaagcg agttcgtgta cggcgactac 4500

aaggtgtacg acgtgcggaa gatgatcgcc aagagcgagc aggaaatcgg caaggctacc 4560

gccaagtact tcttctacag caacatcatg aactttttca agaccgagat taccctggcc 4620

aacggcgaga tccggaagcg gcctctgatc gagacaaacg gcgaaaccgg ggagatcgtg 4680

tgggataagg gccgggattt tgccaccgtg cggaaagtgc tgagcatgcc ccaagtgaat 4740

atcgtgaaaa agaccgaggt gcagacaggc ggcttcagca aagagtctat cctgcccaag 4800

aggaacagcg ataagctgat cgccagaaag aaggactggg accctaagaa gtacggcggc 4860

ttcgacagcc ccaccgtggc ctattctgtg ctggtggtgg ccaaagtgga aaagggcaag 4920

tccaagaaac tgaagagtgt gaaagagctg ctggggatca ccatcatgga aagaagcagc 4980

ttcgagaaga atcccatcga ctttctggaa gccaagggct acaaagaagt gaaaaaggac 5040

ctgatcatca agctgcctaa gtactccctg ttcgagctgg aaaacggccg gaagagaatg 5100

ctggcctctg ccggcgaact gcagaaggga aacgaactgg ccctgccctc caaatatgtg 5160

aacttcctgt acctggccag ccactatgag aagctgaagg gctcccccga ggataatgag 5220

cagaaacagc tgtttgtgga acagcacaag cactacctgg acgagatcat cgagcagatc 5280

agcgagttct ccaagagagt gatcctggcc gacgctaatc tggacaaagt gctgtccgcc 5340

tacaacaagc accgggataa gcccatcaga gagcaggccg agaatatcat ccacctgttt 5400

accctgacca atctgggagc ccctgccgcc ttcaagtact ttgacaccac catcgaccgg 5460

aagaggtaca ccagcaccaa agaggtgctg gacgccaccc tgatccacca gagcatcacc 5520

ggcctgtacg agacacggat cgacctgtct cagctgggag gtgactctgg cggctcaaaa 5580

agaaccgccg acggcagcga attcgagccc aagaagaaga ggaaagtcta a 5631

<210> 43

<211> 5631

<212> DNA

<213> 人工序列(Artificial Sequence)

<400> 43

atgaaacgga cagccgacgg aagcgagttc gagtcaccaa agaagaagcg gaaagtcgac 60

aagaagtaca gcatcggcct ggccatcggc accaactctg tgggctgggc cgtgatcacc 120

gacgagtaca aggtgcccag caagaaattc aaggtgctgg gcaacaccga ccggcacagc 180

atcaagaaga acctgatcgg agccctgctg ttcgacagcg gcgaaacagc cgaggccacc 240

cggctgaaga gaaccgccag aagaagatac accagacgga agaaccggat ctgctatctg 300

caagagatct tcagcaacga gatggccaag gtggacgaca gcttcttcca cagactggaa 360

gagtccttcc tggtggaaga ggataagaag cacgagcggc accccatctt cggcaacatc 420

gtggacgagg tggcctacca cgagaagtac cccaccatct accacctgag aaagaaactg 480

gtggacagca ccgacaaggc cgacctgcgg ctgatctatc tggccctggc ccacatgatc 540

aagttccggg gccacttcct gatcgagggc gacctgaacc ccgacaacag cgacgtggac 600

aagctgttca tccagctggt gcagacctac aaccagctgt tcgaggaaaa ccccatcaac 660

gccagcggcg tggacgccaa ggccatcctg tctgccagac tgagcaagag cagacggctg 720

gaaaatctga tcgcccagct gcccggcgag aagaagaatg gcctgttcgg aaacctgatt 780

gccctgagcc tgggcctgac ccccaacttc aagagcaact tcgacctggc cgaggatgcc 840

aaactgcagc tgagcaagga cacctacgac gacgacctgg acaacctgct ggcccagatc 900

ggcgaccagt acgccgacct gtttctggcc gccaagaacc tgtccgacgc catcctgctg 960

agcgacatcc tgagagtgaa caccgagatc accaaggccc ccctgagcgc ctctatgatc 1020

aagagatacg acgagcacca ccaggacctg accctgctga aagctctcgt gcggcagcag 1080

ctgcctgaga agtacaaaga gattttcttc gaccagagca agaacggcta cgccggctac 1140

attgacggcg gagccagcca ggaagagttc tacaagttca tcaagcccat cctggaaaag 1200

atggacggca ccgaggaact gctcgtgaag ctgaacagag aggacctgct gcggaagcag 1260

cggaccttcg acaacggcag catcccccac cagatccacc tgggagagct gcacgccatt 1320

ctgcggcggc aggaagattt ttacccattc ctgaaggaca accgggaaaa gatcgagaag 1380

atcctgacct tccgcatccc ctactacgtg ggccctctgg ccaggggaaa cagcagattc 1440

gcctggatga ccagaaagag cgaggaaacc atcaccccct ggaacttcga ggaagtggtg 1500

gacaagggcg cttccgccca gagcttcatc gagcggatga ccaacttcga taagaacctg 1560

cccaacgaga aggtgctgcc caagcacagc ctgctgtacg agtacttcac cgtgtataac 1620

gagctgacca aagtgaaata cgtgaccgag ggaatgagaa agcccgcctt cctgagcggc 1680

gagcagaaaa aggccatcgt ggacctgctg ttcaagacca accggaaagt gaccgtgaag 1740

cagctgaaag aggactactt caagaaaatc gagtgcttcg actccgtgga aatctccggc 1800

gtggaagatc ggttcaacgc ctccctgggc acataccacg atctgctgaa aattatcaag 1860

gacaaggact tcctggacaa tgaggaaaac gaggacattc tggaagatat cgtgctgacc 1920

ctgacactgt ttgaggacag agagatgatc gaggaacggc tgaaaaccta tgcccacctg 1980

ttcgacgaca aagtgatgaa gcagctgaag cggcggagat acaccggctg gggcaggctg 2040

agccggaagc tgatcaacgg catccgggac aagcagtccg gcaagacaat cctggatttc 2100

ctgaagtccg acggcttcgc caacagaaac ttcatgcagc tgatccacga cgacagcctg 2160

acctttaaag aggacatcca gaaagcccag gtgtccggcc agggcgatag cctgcacgag 2220

cacattgcca atctggccgg cagccccgcc attaagaagg gcatcctgca gacagtgaag 2280

gtggtggacg agctcgtgaa agtgatgggc cggcacaagc ccgagaacat cgtgatcgaa 2340

atggccagag agaaccagac cacccagaag ggacagaaga acagccgcga gagaatgaag 2400

cggatcgaag agggcatcaa agagctgggc agccagatcc tgaaagaaca ccccgtggaa 2460

aacacccagc tgcagaacga gaagctgtac ctgtactacc tgcagaatgg gcgggatatg 2520

tacgtggacc aggaactgga catcaaccgg ctgtccgact acgatgtgga ccatatcgtg 2580

cctcagagct ttctgaagga cgactccatc gacaacaagg tgctgaccag aagcgacaag 2640

aaccggggca agagcgacaa cgtgccctcc gaagaggtcg tgaagaagat gaagaactac 2700

tggcggcagc tgctgaacgc caagctgatt acccagagaa agttcgacaa tctgaccaag 2760

gccgagagag gcggcctgag cgaactggat aaggccggct tcatcaagag acagctggtg 2820

gaaacccggc agatcacaaa gcacgtggca cagatcctgg actcccggat gaacactaag 2880

tacgacgaga atgacaagct gatccgggaa gtgaaagtga tcaccctgaa gtccaagctg 2940

gtgtccgatt tccggaagga tttccagttt tacaaagtgc gcgagatcaa caactaccac 3000

cacgcccacg acgcctacct aaacgccgtc gtgggaaccg ccctgatcaa aaagtaccct 3060

aagctggaaa gcgagttcgt gtacggcgac tacaaggtgt acgacgtgcg gaagatgatc 3120

gccaagagcg agcagtctgg cagcgagaca ccaggaacaa gcgagtcagc aacaccagag 3180

agcggcagcg aggcatctcc agcaagcgga ccaaggcacc tgatggaccc ccacatcttc 3240

acctctaact ttaacaatgg catcggcagg cacaagacat acctgtgcta tgaggtggag 3300

cgcctggaca atggcaccag cgtgaagatg gatcagcaca gaggcttcct gcacaaccag 3360

gccaagaatc tgctgtgcgg cttctacggc cggcacgcag agctgagatt tctggacctg 3420

gtgcctagcc tgcagctgga tccagcccag atctataggg tgacctggtt catcagctgg 3480

tccccatgct tttcctgggg atgtgcagga gaggtgcgcg cctttctgca ggagaacaca 3540

cacgtgcggc tgagaatctt cgccgcccgg atctttgact acgatcctct gtataaggag 3600

gccctgcaga tgctgagaga cgcaggagcc caggtgtcca tcatgaccta cgatgagttc 3660

aagcactgct gggacacatt tgtggatcac cagggctgtc ccttccagcc ttgggacgga 3720

ctggatgagc actcccaggc cctgtctggc aggctgaggg ccatcctgca gaaccagggc 3780

aatagcggag gatccggagg atctggaggc agcatggccg ctgctaaagc ccccggcgct 3840

gctgaattcg tccccgctga tgccgacctg gatacactgc ggaccgccgt gcagggctgc 3900

agaggctgtg aactgtacag aggcgccacc caggccgtgt tcggcgaggg ccctgctcac 3960

gcccctgtgt ttgtggtcgg cgagcagcct ggcgaccggg aagatgtggc cggccacccc 4020

ttcgtgggcc ccgccggcag actgctggac aaggccctga cagaggccga catcgacagg 4080

gaagccgtgt acctgaccaa cgccgttaag cacttcaagt tcgaggaaag aggaaaaaga 4140

agaatccaca agcaacctgg cagaaccgag gtggtggcat gcagcccttg gctgaccgcc 4200

gagctggacg ccgtgcggcc tcagctggtg gtgtgcctgg gcgccgtggc cgccaaggcc 4260

gtgctgggac catcttttaa ggtgtccgag cggcggggcg aagtggtcga ggccggagag 4320

cacagagtga tcgccacagt gcaccctagc agcgtgctga gagccccaga ccgcgccgct 4380

gcctacgccg acttcctggc cgatctgaga aaggtgcgga ccgccgctgg agagctccat 4440

agagcctctg gaggatctag cggaggatcc tctggcagcg agacaccagg aacaagcgag 4500

tcagcaacac cagagagcag tggcggcagc agcggcggca gcgaaatcgg caaggctacc 4560

gccaagtact tcttctacag caacatcatg aactttttca agaccgagat taccctggcc 4620

aacggcgaga tccggaagcg gcctctgatc gagacaaacg gcgaaaccgg ggagatcgtg 4680

tgggataagg gccgggattt tgccaccgtg cggaaagtgc tgagcatgcc ccaagtgaat 4740

atcgtgaaaa agaccgaggt gcagacaggc ggcttcagca aagagtctat cctgcccaag 4800

aggaacagcg ataagctgat cgccagaaag aaggactggg accctaagaa gtacggcggc 4860

ttcgacagcc ccaccgtggc ctattctgtg ctggtggtgg ccaaagtgga aaagggcaag 4920

tccaagaaac tgaagagtgt gaaagagctg ctggggatca ccatcatgga aagaagcagc 4980

ttcgagaaga atcccatcga ctttctggaa gccaagggct acaaagaagt gaaaaaggac 5040

ctgatcatca agctgcctaa gtactccctg ttcgagctgg aaaacggccg gaagagaatg 5100

ctggcctctg ccggcgaact gcagaaggga aacgaactgg ccctgccctc caaatatgtg 5160

aacttcctgt acctggccag ccactatgag aagctgaagg gctcccccga ggataatgag 5220

cagaaacagc tgtttgtgga acagcacaag cactacctgg acgagatcat cgagcagatc 5280

agcgagttct ccaagagagt gatcctggcc gacgctaatc tggacaaagt gctgtccgcc 5340

tacaacaagc accgggataa gcccatcaga gagcaggccg agaatatcat ccacctgttt 5400

accctgacca atctgggagc ccctgccgcc ttcaagtact ttgacaccac catcgaccgg 5460

aagaggtaca ccagcaccaa agaggtgctg gacgccaccc tgatccacca gagcatcacc 5520

ggcctgtacg agacacggat cgacctgtct cagctgggag gtgactctgg cggctcaaaa 5580

agaaccgccg acggcagcga attcgagccc aagaagaaga ggaaagtcta a 5631

<210> 44

<211> 5631

<212> DNA

<213> 人工序列(Artificial Sequence)

<400> 44

atgaaacgga cagccgacgg aagcgagttc gagtcaccaa agaagaagcg gaaagtcgac 60

aagaagtaca gcatcggcct ggccatcggc accaactctg tgggctgggc cgtgatcacc 120

gacgagtaca aggtgcccag caagaaattc aaggtgctgg gcaacaccga ccggcacagc 180

atcaagaaga acctgatcgg agccctgctg ttcgacagcg gcgaaacagc cgaggccacc 240

cggctgaaga gaaccgccag aagaagatac accagacgga agaaccggat ctgctatctg 300

caagagatct tcagcaacga gatggccaag gtggacgaca gcttcttcca cagactggaa 360

gagtccttcc tggtggaaga ggataagaag cacgagcggc accccatctt cggcaacatc 420

gtggacgagg tggcctacca cgagaagtac cccaccatct accacctgag aaagaaactg 480

gtggacagca ccgacaaggc cgacctgcgg ctgatctatc tggccctggc ccacatgatc 540

aagttccggg gccacttcct gatcgagggc gacctgaacc ccgacaacag cgacgtggac 600

aagctgttca tccagctggt gcagacctac aaccagctgt tcgaggaaaa ccccatcaac 660

gccagcggcg tggacgccaa ggccatcctg tctgccagac tgagcaagag cagacggctg 720

gaaaatctga tcgcccagct gcccggcgag aagaagaatg gcctgttcgg aaacctgatt 780

gccctgagcc tgggcctgac ccccaacttc aagagcaact tcgacctggc cgaggatgcc 840

aaactgcagc tgagcaagga cacctacgac gacgacctgg acaacctgct ggcccagatc 900

ggcgaccagt acgccgacct gtttctggcc gccaagaacc tgtccgacgc catcctgctg 960

agcgacatcc tgagagtgaa caccgagatc accaaggccc ccctgagcgc ctctatgatc 1020

aagagatacg acgagcacca ccaggacctg accctgctga aagctctcgt gcggcagcag 1080

ctgcctgaga agtacaaaga gattttcttc gaccagagca agaacggcta cgccggctac 1140

attgacggcg gagccagcca ggaagagttc tacaagttca tcaagcccat cctggaaaag 1200

atggacggca ccgaggaact gctcgtgaag ctgaacagag aggacctgct gcggaagcag 1260

cggaccttcg acaacggcag catcccccac cagatccacc tgggagagct gcacgccatt 1320

ctgcggcggc aggaagattt ttacccattc ctgaaggaca accgggaaaa gatcgagaag 1380

atcctgacct tccgcatccc ctactacgtg ggccctctgg ccaggggaaa cagcagattc 1440

gcctggatga ccagaaagag cgaggaaacc atcaccccct ggaacttcga ggaagtggtg 1500

gacaagggcg cttccgccca gagcttcatc gagcggatga ccaacttcga taagaacctg 1560

cccaacgaga aggtgctgcc caagcacagc ctgctgtacg agtacttcac cgtgtataac 1620

gagctgacca aagtgaaata cgtgaccgag ggaatgagaa agcccgcctt cctgagcggc 1680

gagcagaaaa aggccatcgt ggacctgctg ttcaagacca accggaaagt gaccgtgaag 1740

cagctgaaag aggactactt caagaaaatc gagtgcttcg actccgtgga aatctccggc 1800

gtggaagatc ggttcaacgc ctccctgggc acataccacg atctgctgaa aattatcaag 1860

gacaaggact tcctggacaa tgaggaaaac gaggacattc tggaagatat cgtgctgacc 1920

ctgacactgt ttgaggacag agagatgatc gaggaacggc tgaaaaccta tgcccacctg 1980

ttcgacgaca aagtgatgaa gcagctgaag cggcggagat acaccggctg gggcaggctg 2040

agccggaagc tgatcaacgg catccgggac aagcagtccg gcaagacaat cctggatttc 2100

ctgaagtccg acggcttcgc caacagaaac ttcatgcagc tgatccacga cgacagcctg 2160

acctttaaag aggacatcca gaaagcccag gtgtccggcc agggcgatag cctgcacgag 2220

cacattgcca atctggccgg cagccccgcc attaagaagg gcatcctgca gacagtgaag 2280

gtggtggacg agctcgtgaa agtgatgggc cggcacaagc ccgagaacat cgtgatcgaa 2340

atggccagag agaaccagac cacccagaag ggacagaaga acagccgcga gagaatgaag 2400

cggatcgaag agggcatcaa agagctgggc agccagatcc tgaaagaaca ccccgtggaa 2460

aacacccagc tgcagaacga gaagctgtac ctgtactacc tgcagaatgg gcgggatatg 2520

tacgtggacc aggaactgga catcaaccgg ctgtccgact acgatgtgga ccatatcgtg 2580

cctcagagct ttctgaagga cgactccatc gacaacaagg tgctgaccag aagcgacaag 2640

aaccggggca agagcgacaa cgtgccctcc gaagaggtcg tgaagaagat gaagaactac 2700

tggcggcagc tgctgaacgc caagctgatt acccagagaa agttcgacaa tctgaccaag 2760

gccgagagag gcggcctgag cgaactggat aaggccggct tcatcaagag acagctggtg 2820

gaaacccggc agatcacaaa gcacgtggca cagatcctgg actcccggat gaacactaag 2880

tacgacgaga atgacaagct gatccgggaa gtgaaagtga tcaccctgaa gtccaagctg 2940

gtgtccgatt tccggaagga tttccagttt tacaaagtgc gcgagatcaa caactaccac 3000

cacgcccacg acgcctacct aaacgccgtc gtgggaaccg ccctgatcaa aaagtaccct 3060

aagctggaaa gcgagttcgt gtacggcgac tacaaggtgt acgacgtgcg gaagatgatc 3120

gccaagagcg agcaggaaat cggcaaggct accgccaagt acttcttcta ctctggcagc 3180

gagacaccag gaacaagcga gtcagcaaca ccagagagcg gcagcgaggc atctccagca 3240

agcggaccaa ggcacctgat ggacccccac atcttcacct ctaactttaa caatggcatc 3300

ggcaggcaca agacatacct gtgctatgag gtggagcgcc tggacaatgg caccagcgtg 3360

aagatggatc agcacagagg cttcctgcac aaccaggcca agaatctgct gtgcggcttc 3420

tacggccggc acgcagagct gagatttctg gacctggtgc ctagcctgca gctggatcca 3480

gcccagatct atagggtgac ctggttcatc agctggtccc catgcttttc ctggggatgt 3540

gcaggagagg tgcgcgcctt tctgcaggag aacacacacg tgcggctgag aatcttcgcc 3600

gcccggatct ttgactacga tcctctgtat aaggaggccc tgcagatgct gagagacgca 3660

ggagcccagg tgtccatcat gacctacgat gagttcaagc actgctggga cacatttgtg 3720

gatcaccagg gctgtccctt ccagccttgg gacggactgg atgagcactc ccaggccctg 3780

tctggcaggc tgagggccat cctgcagaac cagggcaata gcggaggatc cggaggatct 3840

ggaggcagca tggccgctgc taaagccccc ggcgctgctg aattcgtccc cgctgatgcc 3900

gacctggata cactgcggac cgccgtgcag ggctgcagag gctgtgaact gtacagaggc 3960

gccacccagg ccgtgttcgg cgagggccct gctcacgccc ctgtgtttgt ggtcggcgag 4020

cagcctggcg accgggaaga tgtggccggc caccccttcg tgggccccgc cggcagactg 4080

ctggacaagg ccctgacaga ggccgacatc gacagggaag ccgtgtacct gaccaacgcc 4140

gttaagcact tcaagttcga ggaaagagga aaaagaagaa tccacaagca acctggcaga 4200

accgaggtgg tggcatgcag cccttggctg accgccgagc tggacgccgt gcggcctcag 4260

ctggtggtgt gcctgggcgc cgtggccgcc aaggccgtgc tgggaccatc ttttaaggtg 4320

tccgagcggc ggggcgaagt ggtcgaggcc ggagagcaca gagtgatcgc cacagtgcac 4380

cctagcagcg tgctgagagc cccagaccgc gccgctgcct acgccgactt cctggccgat 4440

ctgagaaagg tgcggaccgc cgctggagag ctccatagag cctctggagg atctagcgga 4500

ggatcctctg gcagcgagac accaggaaca agcgagtcag caacaccaga gagcagtggc 4560

ggcagcagcg gcggcagcag caacatcatg aactttttca agaccgagat taccctggcc 4620

aacggcgaga tccggaagcg gcctctgatc gagacaaacg gcgaaaccgg ggagatcgtg 4680

tgggataagg gccgggattt tgccaccgtg cggaaagtgc tgagcatgcc ccaagtgaat 4740

atcgtgaaaa agaccgaggt gcagacaggc ggcttcagca aagagtctat cctgcccaag 4800

aggaacagcg ataagctgat cgccagaaag aaggactggg accctaagaa gtacggcggc 4860

ttcgacagcc ccaccgtggc ctattctgtg ctggtggtgg ccaaagtgga aaagggcaag 4920

tccaagaaac tgaagagtgt gaaagagctg ctggggatca ccatcatgga aagaagcagc 4980

ttcgagaaga atcccatcga ctttctggaa gccaagggct acaaagaagt gaaaaaggac 5040

ctgatcatca agctgcctaa gtactccctg ttcgagctgg aaaacggccg gaagagaatg 5100

ctggcctctg ccggcgaact gcagaaggga aacgaactgg ccctgccctc caaatatgtg 5160

aacttcctgt acctggccag ccactatgag aagctgaagg gctcccccga ggataatgag 5220

cagaaacagc tgtttgtgga acagcacaag cactacctgg acgagatcat cgagcagatc 5280

agcgagttct ccaagagagt gatcctggcc gacgctaatc tggacaaagt gctgtccgcc 5340

tacaacaagc accgggataa gcccatcaga gagcaggccg agaatatcat ccacctgttt 5400

accctgacca atctgggagc ccctgccgcc ttcaagtact ttgacaccac catcgaccgg 5460

aagaggtaca ccagcaccaa agaggtgctg gacgccaccc tgatccacca gagcatcacc 5520

ggcctgtacg agacacggat cgacctgtct cagctgggag gtgactctgg cggctcaaaa 5580

agaaccgccg acggcagcga attcgagccc aagaagaaga ggaaagtcta a 5631

Claims (25)

1.一种融合蛋白，所述融合蛋白自N端至C端依次包括第一Cas9切口酶片段、嵌合插入片段、第二Cas9切口酶片段，所述嵌合插入片段包含脱氨酶片段和尿嘧啶DNA结合蛋白片段。

2.如权利要求1所述的融合蛋白，其特征在于，所述第一Cas9切口酶片段的氨基酸序列包括如SEQ ID NO.34～36任一所示的片段或其变体；和/或，所述第二Cas9切口酶片段的氨基酸序列包括如SEQ ID NO.37～39任一所示的片段或其变体。

3.如权利要求1所述的融合蛋白，其特征在于，所述脱氨酶选自胞嘧啶脱氨酶或其变体，所述胞嘧啶脱氨酶选自：APOBEC1、APOBEC2、APOBEC3A、APOBEC3B、APOBEC3C、APOBEC3D、APOBEC3F、APOBEC3G、APOBEC3H、APOBEC4、激活诱导的脱氨酶(AID)和pmCDA1；APOBEC家族；优选地，所述胞嘧啶脱氨酶选自ancAPOBEC1、Apobec3A。

4.如权利要求1所述的融合蛋白，其特征在于，所述尿嘧啶DNA结合蛋白选自来自微生物的Udgx家族的酶或其变体；优选地，所述微生物选自类胞内分枝杆菌、少动鞘氨醇单胞菌、球形红假单胞菌、柑桔溃疡病菌、苜蓿中华根瘤菌、皮诺卡氏菌、盖尔森基兴奴卡氏菌、哥伦布分枝杆菌、贪铜菌、阿氟曼链霉菌、欺诈伯克霍尔德菌、梅洛尼鞘氨醇单胞菌、沼泽红假单胞菌、耻垢分枝杆菌、胞内分枝杆菌、多噬伯克霍尔德菌、慢生型大豆根瘤菌、高效固氮慢生根瘤菌、草莓角斑病菌；进一步优选地，所述微生物为皮诺卡氏菌。

5.如权利要求1所述的融合蛋白，其特征在于，所述融合蛋白还包括核定位信号片段，包括以下1)～2)至少任一项：

l)所述核定位信号片段位于融合蛋白的N端和/或C端；

2)所述核定位信号片段优选为BPNLS或其变体，所述变体与BPNLS具有80％以上序列相似性且具有BPNLS的功能。

6.如权利要求5所述的融合蛋白，其特征在于，包括至少任一项：

1)所述融合蛋白还包括柔性连接肽片段；优选地，所述柔性连接肽片段的氨基酸序列包括SEQ ID NO.40：SGSETPGTSESATPESGS、SEQ ID NO.41：SGSGSETPGTSESATPES；

2)所述融合蛋白自N端到C端依次是核定位信号片段、第一Cas9切口酶片段、胞嘧啶脱氨酶片段、尿嘧啶DNA结合蛋白、第二Cas9切口酶片段和核定位信号片段；或，

3)所述融合蛋白自N端到C端依次是核定位信号片段、第一Cas9切口酶片段、尿嘧啶DNA结合蛋白、胞嘧啶脱氨酶片段、第二Cas9切口酶片段和核定位信号片段。

7.如权利要求1所述的融合蛋白，其特征在于，所述融合蛋白的氨基酸序列如SEQ IDNo.11～13至少任一所示。

8.一种分离的多核苷酸，其特征在于，其编码如权利要求1～7任一权利要求所述的融合蛋白。

9.一种构建体，其特征在于，所述构建体含有如权利要求8所述的分离的多核苷酸。

10.一种表达系统，其特征在于，所述表达系统含有如权利要求9所述的构建体或基因组中整合有外源的如权利要求8所述的多核苷酸。

11.如权利要求10所述的表达系统，其特征在于，所述表达系统的宿主细胞选自真核细胞或原核细胞；优选地，所述宿主细胞选自小鼠细胞、人细胞。

12.一种碱基编辑系统，其包括如权利要求1～7任一权利要求所述的融合蛋白或其编码多核苷酸。

13.如权利要求12所述的碱基编辑系统，其特征在于，所述碱基编辑系统还包括引导RNA；包括以下至少任一项：

1)所述碱基编辑系统包括：a)所述融合蛋白或其编码多核苷酸，b)引导RNA核苷酸序列或其编码多核苷酸；和/或，

2)所述引导RNA将所述融合蛋白靶向靶标序列中的C碱基。

14.如权利要求13所述的碱基编辑系统，其特征在于，包括以下至少任一项：

1)所述碱基编辑系统包含一个或多个载体；所述一个或多个载体包含(i)第一调控元件，所述第一调控元件可操作地连接至所述融合蛋白的编码多核苷酸；以及(ii)第二调控元件，所述第二调控元件可操作地连接至所述引导RNA核苷酸序列的编码多核苷酸；

所述(i)和(ii)位于相同或不同载体上；

2)所述碱基编辑系统包含(i)融合蛋白，以及(ii)包含所述引导核苷酸序列的编码多核苷酸的载体。

15.如权利要求1～7任一权利要求所述的融合蛋白、如权利要求8所述的分离的多核苷酸、如权利要求9所述的构建体或如权利要求10或11所述的表达系统或如权利要求12～14任一项所述的碱基编辑系统在基因编辑中的用途。

16.如权利要求15所述的用途，其特征在于，包括以下至少任一项：

1)所述基因编辑实现碱基颠换；

2)所述基因编辑实现C至G的颠换或者G至C的颠换；

3)所述基因编辑用于实现致病位点的修正、基因功能研究、增强细胞功能、细胞治疗至少任一。

17.如权利要求16所述的用途，其特征在于，包括以下至少任一项：

1)所述治病位点导致的疾病选自以下至少任一项：自身免疫性疾病、肿瘤、病毒感染性疾病、细菌感染性疾病；

2)所述融合蛋白、分离的多核苷酸、构建体、表达系统或碱基编辑系统与其他药物或试剂联用。

18.一种基因编辑方法，包括：通过如权利要求1～7任一权利要求所述的融合蛋白、如权利要求8所述的分离的多核苷酸、如权利要求9所述的构建体或如权利要求10或11所述的表达系统或如权利要求12～14任一项所述的碱基编辑系统对靶标序列进行碱基编辑。

19.如权利要求18所述的方法，其特征在于，包括以下至少任一项：

1)所述方法在体外实施；优选地，所述方法在培养的细胞中实施；

2)所述方法在体内实施；优选地，所述方法在哺乳动物中实施；进一步优选地，所述哺乳动物是啮齿动物；更进一步优选地，所述哺乳动物是人。

20.一种碱基编辑的细胞，其特征在于，所述细胞包含C至G或G至C的突变。

21.根据权利要求20所述的细胞，其特征在于，其根据权利要求18或19所述的方法制备得到。

22.一种报告系统，其特征在于，所述报告系统包含如SEQ ID NO.31所示的核苷酸序列。

23.如权利要求22所述的报告系统，其特征在于，包括以下至少任一项：

1)使用时，当所述报告系统中的SEQ ID NO.31所示的核苷酸序列突变为SEQ ID NO.30时，所述报告系统呈现蓝色荧光；

当所述报告系统中的SEQ ID NO.31所示的核苷酸序列突变为SEQ ID NO.32时，所述报告系统呈现绿色荧光；

2)所述报告系统包含质粒，所述质粒包含如SEQ ID NO.31所示的核苷酸序列，所述质粒的核苷酸序列如SEQ ID NO.2所示。

24.如权利要求23所述的报告系统的用途，其特征在于，所述报告系统用于检测如权利要求1～7任一权利要求所述的融合蛋白、如权利要求8所述的分离的多核苷酸、如权利要求9所述的构建体或如权利要求10或11所述的表达系统或如权利要求12～14任一项所述的碱基编辑系统的C-G编辑效率。

25.一种尿嘧啶DNA结合蛋白在用于构建碱基编辑蛋白或碱基编辑系统中的用途，其特征在于，所述碱基编辑蛋白或碱基编辑系统用于实现C至G或者G至C的颠换；所述尿嘧啶DNA结合蛋白选自：

1)来自微生物的Udgx家族的酶；优选地，所述微生物选自类胞内分枝杆菌、少动鞘氨醇单胞菌、球形红假单胞菌、柑桔溃疡病菌、苜蓿中华根瘤菌、皮诺卡氏菌、盖尔森基兴奴卡氏菌、哥伦布分枝杆菌、贪铜菌、阿氟曼链霉菌、欺诈伯克霍尔德菌、梅洛尼鞘氨醇单胞菌、沼泽红假单胞菌、耻垢分枝杆菌、胞内分枝杆菌、多噬伯克霍尔德菌、慢生型大豆根瘤菌、高效固氮慢生根瘤菌、草莓角斑病菌；进一步优选地，所述微生物为皮诺卡氏菌；或，

2)与1)具有80％以上序列相似性的氨基酸序列、且具有1)所限定的氨基酸序列的功能，优选为具有胞嘧啶脱氨酶活性。

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