CN1355847A - 新型植物基因及其用途 - Google Patents
新型植物基因及其用途 Download PDFInfo
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Abstract
本发明涉及从烟草(Nicotiana tabacum)、番茄(Lycopericon esculentum)、欧洲油菜(Brassica napus)、鼠耳芥(Arabidopsis thaliana)、甜菜(Beta vulgaris)、向日葵(Helianthusannuus)和马铃薯(Solanum tuberosum)中分离与可导致系统获得性抗性(SAR)的信号转导级联相关的鼠耳芥属NIM1基因同源物。本发明进一步涉及用于在转基因植物中表达NIM1同源物以便增加SAR基因表达并提高广谱病害抗性的转化载体和方法。
Description
本发明涉及植物中的广谱病害抗性、包括系统获得性抗性(SAR)的现象。更具体地说,本发明涉及与可导致植物中系统获得性抗性(SAR)的信号转导级联相关的鼠耳芥属NIM1基因同源物的鉴定、分离和表征。
植物经常受到包括病毒、细菌、真菌和线虫在内的各种致病生物体的攻击。农作物植物特别脆弱,因为它们通常生长成遗传均匀的单种栽培物;当病害发作时,损失可能是严重的。然而,大部分植物具有其抵抗致病生物体的先天机制。植物培育者和病理学家已经鉴定了对植物病原体具有抗性的天然变化形式并将其培育成许多农作物植物。这些天然病害抗性基因通常提供了抵抗病原体的高水平抗性或免疫性。
系统获得性抗性(SAR)是一种用于自身抵抗病原体的复合系统植物的组成部分(Hunt和Ryals,1996;Ryals等,1996)。另外参见美国专利号5,614,395。SAR是植物病原体反应的一个特别重要的方面,因为它是病原体诱导的对包括病毒、细菌和真菌在内的广谱感染性病原体的系统抗性。当SAR信号转导途径被阻断时,植物变得对通常导致病害的病原体更为敏感且它们对不经常导致病害的某些感染性病原体也变得敏感(Gaffney等,1997;Delaney等,1994;Delaney等,1997;Bi等,1995;Mauch-Mani和Slusarenko,1996)。这些观察结果表明SAR信号转导途径对维持植物健康是关键的。
在理论上,SAR反应可以被分成两个阶段。在初始阶段中,识别病原体感染且释放通过韧皮部传输到远端组织的信号。这种系统性信号被因SAR基因和病害抗性的表达而起反应的靶细胞觉察。SAR的维持期是指从数周到植物的整个生命期的时间期限,在此过程中该植物是准稳态的且病害抗性得到控制(Ryals等,1996)。
水杨酸(SA)的累积看起来是SAR信号转导所需的。不能因用特异性抑制剂处理,即,苯丙氨酸解氨酶的后生阻抑或特异性降解SA的水杨酸羟化酶的转基因表达而累积SA的植物也不能诱导SAR基因表达或病害抗性(Gaffney等,1993;Delaney等,1994;Mauch-Mani和Slusaerenko,1996;Maher等,1994;Pallas等,1996)。尽管已经建议SA可以用作系统性信号,但是目前这是有争议的;且迄今为止,确切已知所有情况是:如果SA不能累积,那么SAR信号转导将被阻断(Pallas等1996;Shulaev,1995;Vernooij等,1994)。
近来,鼠耳芥属已经表现出可作为研究SAR的模型系统(Uknes等,1992;Uknes等,1993;Cameron等,1994;Mauch-Mani和Slusarenko,1994;Dempsey和Klessig,1995)。已经证实鼠耳芥属中的SAR可以被病原体和诸如SA、2,6-二氯异烟酸(INA)和苯并(1,2,3)噻二唑-7-硫代羧酸S-甲酯(BTH)这样的化学药品所激活(Uknes等,1992;Vernooij等,1995;Lawton等,1996)。在用INA或BTH进行处理或病原体感染后,至少三种与病理相关的(PR)蛋白质基因即PR-1、PR-2和PR-5同时被诱导且伴随抗性发作(Uknes等,1992,1993)。在烟草中,用病原体或免疫化合物处理的最佳特征化物种诱导至少9组基因的表达(Ward等,1991)。通过用不同的SAR基因转化植物已经产生了转基因病害抗性植物(美国专利号5,614,395)。
已经分离出了具有改良的SAR信号转导的大量鼠耳芥属突变体(Delaney,1997)。这些突变体中最重要的是所谓的1sd(病变模拟病害)突变体和acd2(加速的细胞死亡)(Dietrich等,1994;Greenberg等,1994)。这些突变体均在其叶上具有一定程度的自发性坏死病变形成、SA水平升高、SAR基因的mRNA累积和病害抗性显著提高。已经分离并表征了至少7种不同的1sd突变体(Dietrich等;1994;Weymann等,1995)。另一种有意义类型的突变体是cim(组成型免疫)突变体(Lawton等,1993)。另外参见美国专利号5,792,904和国际PCT申请WO 94/16077。类似于1sd突变体和acd2,cim突变体具有升高的SA和SAR基因表达和抗性,不过,与1sd或acd2相反,在其叶上不会表现出可检测到的病变。cpr1(PR基因的组成型表达子)可以是一类cim突变体;然而,因为尚未消除cpr1叶上存在的显微病变,所以cpr1可能是一类1sd突变体(Bowling等,1994)。
已经分离了在SAR信号传输中被阻断的突变体。ndr1(非物种特异性病害抗性)是一种使含有不同无毒力基因的丁香假单胞菌和寄生霜霉的一般无毒性分离物生长的突变体(Century等;1995)。显然这种突变体在SAR信号传输中的早期被阻断。npr1(PR基因的非表达子)是一种不能在INA处理后诱导SAR信号途径表达的突变体(Cao等,1994)。已经根据eds(增强的病害敏感性)突变体在接种低浓度细菌后维持细菌感染的能力分离了它们(Glazebrook等,1996;Parker等,1996)。某些eds突变体在表型上非常类似于npr1且近来已经证实eds5和eds53是npr1的等位基因(Glazebrook等,1996)。nim1(非诱导型免疫)是一种在用INA处理后维持寄生霜霉(即霜霉病致病病原体)生长的突变体(Delaney等,1995;美国专利号5,792,904)。尽管nim1可以在病原体感染后累积SA,但是它不能诱导SAR基因表达或病害抗性,这提示这种突变阻断了SA下游的途径。nim1在其对INA或BTH反应的能力上也受到了削弱,这提示在这些化学药品作用的下游存在阻断作用(Delaney等,1995;Lawton等,1996)。
已经分离并表征了鼠耳芥属的等位基因,它们的突变体分别对nim1和npr1表型负责(Ryals等,1997;Cao等,1997)。野生型NIM1基因产物与导致鼠耳芥属中SAR和基因对基因病害抗性的信号转导级联有关(Ryals等,1997)。Ryals等,1997还报导了nim1的另外5种等位基因的分离方法,这些等位基因显示出在化学诱导的PR-1基因表达和真菌抗性方面从每周削弱到被强力阻断的一系列的表型。将野生型NPR1基因转化成npr1突变体不仅补充了突变,从而恢复了SAR诱导作用对PR-基因表达和病害抗性的反应性,而且在没有SAR诱导作用存在的情况下赋予了所述的转基因植物对丁香假单胞菌导致的感染更高的抗性(Cao等,1997)。WO 98/06748中描述了从鼠耳芥属中分离NPR1和从Nicotiana glutinosa中分离同源物的方法。另外参见WO 97/49822、WO 98/26082和WO 98/29537。
尽管作了大量研究且应用了包括植物的遗传转化在内的完善和深入细致的农作物保护措施,但是每年因病害导致的损失仍保持在10亿美元。因此,存在对开发基于植物中病害抗性的遗传基础不断增加的了解的新的农作物保护措施的持续需求。特别地,存在对鉴定、分离和表征来自其它植物物种的鼠耳芥属NIM1基因的同源物的需求。
在描述本发明的过程中,使用下列术语且这些术语的定义如下所示。
相关/可操作地连接:指的是物理或功能相关的两个DNA序列。例如,认为启动子或调节DNA序列与编码RNA或蛋白质的DNA序列“相关”,条件是这两个序列可操作地连接或定位以使调节DNA序列影响编码或结构DNA序列的表达水平。
嵌合基因:一种重组DNA序列,其中启动子或调节DNA序列可操作地与编码mRNA或表达为蛋白质的DNA序列连接或相关,使得调节DNA序列能够调节相关DNA序列的转录或表达。嵌合基因的调节DNA序列通常不与在自然界中发现的相关的DNA序列可操作地连接。
编码序列:转录成诸如mRNA、rRNA、tRNA、snRNA、有义RNA或反义RNA这样的RNA的核酸序列。优选随后在生物体中翻译所述的RNA以便产生蛋白质。
互补:指的是包括反平行核苷酸序列的两个核苷酸序列,所述的反平行核苷酸序列能够在反平行核苷酸序列中的互补碱基对残基之间形成氢键时彼此配对。
表达:指的是植物中内源性基因或转基因的转录和/或翻译。例如,就反义构建体而言,表达可能指的仅是反义DNA的转录。
表达弹夹:能够指导特定核苷酸序列在合适的宿主细胞中的表达的核酸序列,它包括与感兴趣的核苷酸序列可操作地连接的启动子,所述的感兴趣的核苷酸序列与终止信号可操作地连接。它一般还包括适当翻译所述核苷酸序列所需的序列。包括感兴趣的核苷酸序列的表达弹夹可以是嵌合的,即其成分中的至少一种相对于其其它成分中的至少一种而言是异源的。该表达弹夹还可以是一种天然出现但已经以用于异源表达的重组形式获得的表达弹夹。然而,一般来说,该表达弹夹相对于宿主而言是异源的,即所述表达弹夹的特定核酸序列不会在宿主细胞中天然出现且必须通过转化事件将其引入宿主细胞或宿主细胞的祖先。表达弹夹中核苷酸序列的表达可以在组成型启动子或仅当宿主细胞接触某些特定的外部刺激物时启动转录的诱导型启动子的控制下进行。就诸如植物这样的多细胞生物体而言,所述的启动子对特定的组织或器官或发育阶段也具有特异性。
基因:位于基因组内且除上述编码核酸序列外还包括对控制编码部分的表达、即转录和翻译来说是必不可少的其它主要调节核酸序列的确定区。基因还可以包括其它的5’和3’未翻译序列和终止序列。可以存在的其它元件例如是内含子。
异源DNA序列:本文所用的术语“异源DNA序列”、“外源DNA片段”或“异源核酸”各自指的是来源于对特定宿主细胞而言是外来的来源的序列或如果来自相同来源那么是根据其原始形式修饰的序列。因此,宿主细胞中的异源基因包括对特定宿主细胞而言是内源性的但例如已经通过应用DNA改组修饰的基因。该术语还包括天然出现的DNA序列的非天然出现的多拷贝。因此,该术语指的是一种DNA片段,它对所述细胞而言是外源的或异源的或与所述细胞同源但位于通常未发现所述元件的所述宿主细胞核酸内的某一位置上。表达外源DNA片段以便产生外源性多肽类。
同源DNA序列:天然与引入其中的宿主细胞相关的DNA序列。
同类编码:当核酸序列编码与由参比核酸序列编码的多肽具有相同的氨基酸序列的多肽时该核酸序列与参比核酸序列一起同类编码。
分离的:在本发明的上下文中,分离的核酸分子或分离的酶是一种脱离其天然环境存在且因此不是天然产物的受人控制的核酸分子或酶。分离的核酸分子或酶可以以纯化形式存在或可以存在于诸如例如重组宿主细胞这样的非天然环境中。
最小启动子:启动子元件、特别是TATA元件,它们是无活性的或在没有上游激活的情况下具有显著降低的启动子活性。在有合适的转录因子存在的情况下,最小启动子的功能是启动转录。
天然的:指的是存在于未转化细胞基因组中的基因。
天然出现的:术语“天然出现的”用于描述与由人通过人工方式生产不同的能在自然界中发现的物体。例如,存在于生物体(包括病毒)中的蛋白质或核苷酸序列是天然出现的,它们可以分离自天然来源且没有被人在实验室中进行过人为修饰。
NIM1:Ryals等,1997中描述的基因,它与SAR信号转导级联有关。
NIM1:由NIM1基因编码的蛋白质。
核酸:术语“核酸”指的是脱氧核苷酸或核糖核苷酸及其单或双链形式的聚合物。除非另有限定,该术语包括含有天然核苷酸的已知类似物的核酸,它们与参比核酸具有相似的结合特性且以与天然出现的核苷酸相似的方式被代谢。除非另有说明,特定的核酸序列无疑还包括其保守修饰的变化形式(例如简并密码子取代)和互补序列以及明确指定的序列。具体地,可以通过生成其中一种或多种选择的(或全部)密码子的第三位被混合的碱基和/或脱氧肌苷残基取代的序列来实现简并密码子取代(Batzer等,《核酸研究》(Nucleic Acid Res.)19:5081(1991);Ohtsuka等,《生化杂志》(J.Biol.Chem.)260:2605-2608(1985);Rossolini等,《分子细胞探针》(Mol.Cell.Probes)8:91-98(1994))。也可以将术语“核酸”或“核酸序列”与基因、cDNA和由基因编码的mRNA互换使用。在本发明的上下文中,核酸分子优选是DNA片段。核苷酸由下列标准缩写的其碱基来表示:腺嘌呤(A)、胞嘧啶(C)、胸腺嘧啶(T)和鸟嘌呤(G)。
ORF:可读框。
植物:任何完整的植物。
植物细胞:植物的结构和生理单位,它包括原生质体和细胞壁。植物细胞可以是分离的单细胞或培养的细胞形式或作为诸如例如植物组织、植物器官或完整植物这样的高级机体构造单位的一部分。
植物细胞培养物:植物单位培养物,诸如:例如原生质体、细胞培养物细胞、植物组织中的细胞、花粉、花粉管、胚珠、胚囊、合子和不同发育阶段的胚。
植物材料:指的是叶、茎、根、花或花的部分、果实、花粉、卵细胞、合子、种子、插条、细胞或组织培养物、或植物的任何其它部分或产物。
植物器官:植物明显的和可见的结构和分化部分,诸如根、茎、叶、花芽或胚。
植物组织:组织成结构和功能单位的一组植物细胞。包括植物或培养物中的植物的任何组织。本术语包括但不限于完整植物、植物器官、植物种子、组织培养物和组织成结构和/或功能单位的任何组的植物细胞。该术语连同如上所述的或由本定义包括的任何特殊类型的植物组织一起使用或在没有如上所述的或由本定义包括的任何特殊类型的植物组织的情况下使用并不排除任何其它类型的植物组织。
启动子:含有RNA聚合酶II的结合位点并启动DNA转录的编码区上游的未翻译的DNA序列。启动子区还可以包括起基因表达调节物作用的其它元件。
原生质体:不含细胞壁或仅含部分细胞壁的分离的植物细胞。
纯化的:术语“纯化的”在用于核酸或蛋白质时指的是基本上不含其它在天然状态下与其相关的细胞成分的核酸或蛋白质。优选它是在同源状态下,不过,它可以处于无水的或含水的溶液中。一般使用诸如聚丙烯酰胺凝胶电泳或高效液相层析这样的分析型化学技术来测定纯度和均匀性。属于存在于制品中的主要物质的蛋白质基本上是纯化的。术语“纯化的”指的是核酸或蛋白质在电泳凝胶中主要产生一条带。特别地,它指的是核酸或蛋白质的纯度至少约为50%、更优选纯度至少约为85%且最优选纯度至少约为99%。
重组DNA分子:使用重组DNA技术彼此连接的DNA分子的组合。
调节元件:在控制核苷酸序列表达中所涉及的序列。调节元件包括可操纵地与感兴趣的核苷酸序列和终止信号连接的启动子。它们一般还包括正确翻译所述核苷酸序列所需的序列。
选择性标记基因:其在植物细胞中的表达使细胞产生选择优势的基因。用选择性标记基因转化的细胞所具有的选择优势可能是由于与未转化细胞的生长相比它们在有诸如抗生素或除草剂这样的负选择剂存在情况下的生长能力所造成的。与未转化的细胞相比,转化细胞所具有的选择优势也可能是由于它们将添加的化合物用作营养物、生长因子或能源的增强的或新的能力所造成的。选择性标记基因还指其在植物细胞中的表达使细胞产生负的和正的选择优势的基因或基因组合。
显著提高:大于测定技术中固有的误差限度的酶活性的提高,优选在有抑制剂存在的情况下使野生型酶的活性提高约2倍或2倍以上、更优选提高约5倍或5倍以上且最优选提高约10倍或10倍以上。
就两种或多种核酸或蛋白质序列而言所用的术语“相同”或“同一性”百分比指的是当进行最大相应性比较和序列对比时相同或具有规定的氨基酸残基或核苷酸百分比的两种或多种序列或亚序列,正如使用下列序列对比算法或通过目测检查所测得。
基本上相同:就两种核酸或蛋白质序列而言所用的术语“基本上相同”指的是当进行最大相应性比较和序列对比时具有至少60%、优选80%、更优选90-95%且最优选99%的核苷酸或氨基酸残基同一性的两种或多种序列或亚序列,正如使用下列序列对比算法或通过目测检查所测得的。在至少约50个残基长的序列区内、更优选在约100个残基区内存在基本的同一性且最优选在约150个残基内序列基本上相同。在一个最优选的实施方案中,序列在全长编码区内基本上相同。此外,基本上相同的核酸或蛋白质序列具有基本上相同的功能。
为了进行序列比较,一般一个序列起参比序列的作用,将测试序列与其进行比较。当使用序列比较算法时,将测试和参比序列输入计算机,如果需要指定亚序列等同物并指定序列算法程序参数。然后序列比较算法以指定的程序参数为基准计算出测试序列与参比序列相比较的序列同一性百分比。
例如,可以通过下列方法进行用于比较的序列最佳排列:Smith&Waterman的局部同源性算法《数学应用发展》(Adv.Appl.Math.)2:482(1981);Needleman&Wunsch的同源性序列对比算法《分子生物学杂志》(J.Mol.Biol.)443(1970);Pearson&Lipman的相似性方法检索《美国国家科学院院报》(Proc.Nat’l.Acad.Sci.USA)85:2444(1988);这些算法的计算机化应用(Wisconsin GeneticsSofware Package中的GAP,BESTFIT,FASTA和TFASTA,GeneticsComputer Group,575 Science Dr.,Madison,WI);或目测检查(通常参见Ausubel等,参见下文)。
适合于确定序列同一性和序列相似性百分比的算法的一个实例是BLAST算法,它描述在Altschul等的《分子生物学杂志》(J.Mol.Biol.)215:403-410(1990)中。公众可以通过国家生物技术信息中心 (National Center for Biotechnology Information)(http://www.ncbi.nlm.nih.gov/)进行BLAST分析的软件。该算法包括:首先通过确定查询序列中长度W的短词来确定主要划线的序列对(HSPs),当用数据库序列中相同长度的词进行序列对比时,它们与某些正值阈值得分T相适应或满足它们。T指的是邻近词得分阈值(Altschul等,1990)。命中这些起始的邻近词为启动检索程序打下基础以便找到含有它们的更长的HSPs。然后沿各序列的两个方向扩展命中的词,直到累积的序列对比得分得到增加为止。就核苷酸序列而言,使用参数M(配对残基对的有利结果得分;总>0)和N(错配残基的不利结果得分;总<0)来计算累积得分。就氨基酸序列而言,将得分矩阵用于计算累积得分。当累积序列对比得分从其获得的最大值下降到X量时停止在各方向上扩展命中词,累积得分达到0或0以下,这是因为累积了一种或多种负得分的残基序列对比或达到各序列的末端所导致的。BLAST算法的参数W、T和X确定了序列对比的敏感性和速度。BLATN程序(就核苷酸序列而言)使用词长度(W)为11、期望值(E)为10、截止值为100、M=5、N=4和两链比较作为默认值。就氨基酸序列而言,BLASTP程序使用词长度(W)为3、期望值(E)为10和BLOSUM62得分矩阵作为默认值(参见Henikoff&Henikoff《美国国家科学院院报》(Proc.Natl.Acad.Sci.USA)89:10915(1989)。
除计算序列同一性百分比外,BLAST算法还进行了两种序列之间相似性的统计分析(参见,例如Karlin&Altschul《美国国家科学院院报》(Proc.Natl.Acad.Sci.USA)90:5873-5787(1993))。由BLAST算法提供的相似性的一个测定值是最小的和数概率(P(N)),它产生了一个概率的示数,通过该示数也许会出现两个核苷酸或氨基酸序列之间的配对。例如,如果在测试核酸序列与参比核酸序列进行比较过程中的最小和数概率约小于0.1、更优选约小于0.01且最优选约小于0.001,那么认为测试核酸序列与参比序列相似。
两个核酸序列基本上相同的另一种表示方式是两个分子在严格条件下彼此杂交。术语“特异性地与…杂交”指的是分子仅与特定的核苷酸序列在严格条件下的结合、双重组合或杂交,此时所述的序列存在于复合的混合物(例如总细胞)DNA或RNA中。“基本上结合”指的是探针核酸与靶核酸之间的互补杂交且包括可以通过降低杂交培养基的严格性而容纳的最小错配以便实现对靶核酸序列进行所需检测。
就诸如DNA杂交和Northern杂交这样的核酸杂交实验而言所用的“严格杂交条件”和“严格杂交洗涤条件”具有序列依赖性且在不同的环境参数下是不同的。较长的序列特别在较高的温度下杂交。关于核酸杂交的广泛指导原则可以在下列文献中找到:Tijssen(1993)《生化实验室技术和分子生物学-使用核酸探针的杂交》(LaboratoryTechniques in Biochemistry and Molecular Biology-Hybridizationwith Nucleic Acid Probes)第1部分第2章“杂交原理和核酸探针测定策略的综述”(Overview of principles of hybridization and thestrategy of nucleic acid probe assays),Elsevier,New York。一般来说,就在规定的离子强度和pH下的特定序列而言,将高度严格杂交和洗涤条件选择为低于热熔点(Tm)约5℃。一般来说,在“严格条件”下探针与其靶亚序列杂交而不与其它序列杂交。
Tm是50%的靶序列与完好配对探针杂交的温度(在指定的离子强度和pH下)。将非常严格的条件选择为等于特定探针的Tm。用于在DNA或RNA印迹中的滤膜上杂交具有100个以上互补残基的互补核酸的严格杂交条件的一个实例是在42℃下50%甲酰胺与1mg肝素杂交,其中将杂交过程进行过夜。高度严格的洗涤条件的一个实例是在72℃下用0.15M NaCl洗涤约15分钟。一个洗涤条件的一个实例是在65℃下用0.2×SSC洗涤15分钟(参见下文Sambrook对SSC缓冲液的描述)。通常在高度严格性洗涤前是低度严格性洗涤以便除去背景探针信号。就双链体、例如100个以上核苷酸而言,中度严格性洗涤的一个实例是在45℃下用1×SSC洗涤15分钟。就双链体、例如100个以上核苷酸而言,低度严格性洗涤的一个实例是在40℃下用4-6×SSC洗涤15分钟。就短探针(例如约10-50个核苷酸)而言,严格条件一般包括在pH 7.0-8.3下盐浓度低于约1.0M Na离子、一般约为0.01-1.0M Na离子浓度(或其它盐)且温度一般至少约为30℃。还可以通过添加诸如甲酰胺这样的去稳定剂来实现严格条件。一般来说,在特定杂交测定试验中对不相关探针所观察到的2倍(或2倍以上)的信噪比表明了检测特异性杂交。如果核酸编码的蛋白质基本上相同,那么在严格条件下无法彼此杂交的核酸仍基本上相同。例如,这种情况在使用遗传密码所允许的最大密码简并产生核酸拷贝时发生。
下面是可以用于克隆基本上与本发明参比核苷酸序列相同的同源核苷酸序列的杂交/洗涤条件组的实例:参比核苷酸序列与参比核苷酸序列优选在50℃下的7%十二烷基硫酸钠(SDS)、0.5M NaPO4、1mM EDTA中杂交、其中在50℃下用2×SSC洗涤;更理想的是在50℃下的7%十二烷基硫酸钠(SDS)、0.5M NaPO4、1mM EDTA中杂交、其中在50℃下用1×SSC、0.1%SDS洗涤;更理想的是在50℃下的7%十二烷基硫酸钠(SDS)、0.5M NaPO4、1mM EDTA中杂交、其中在50℃下用0.5×SSC、0.1%SDS洗涤;优选在50℃下的7%十二烷基硫酸钠(SDS)、0.5M NaPO4、1mM EDTA中杂交、其中在50℃下用0.1×SSC、0.1%SDS洗涤;更优选在50℃下的7%十二烷基硫酸钠(SDS)、0.5M NaPO4、1mM EDTA中杂交、其中在65℃下用0.1×SSC、0.1%SDS洗涤。
两种核酸序列或蛋白质基本上相同的进一步证明方式是由第一种核酸编码的蛋白质与由第二种核酸编码的蛋白质发生交叉免疫反应或特异性地与之结合。因此,一种蛋白质通常与第二种蛋白质基本上相同,例如,其中两种蛋白质的区别仅在于保守取代。
术语“特异性地(或选择性地)与抗体结合”或“特异性地(或选择性地)与…发生免疫反应”在涉及蛋白质或肽时指的是结合反应是在有蛋白质和其它生物制品的异源群体存在的情况下蛋白质存在的决定因素。因此,在指定的免疫测定条件下,特异性抗体结合特定的蛋白质而不以显著量结合存在于样品中的其它蛋白质。在这类条件下与抗体的特异性结合可能需要根据其对特定蛋白质的特异性来选择的抗体。例如,可以选择针对具有由本发明任何核酸序列编码的氨基酸序列的蛋白质产生的抗体以便获得与该蛋白质发生特异性免疫反应而不与除多态变体外的其它蛋白质发生免疫反应的抗体。可以将多种免疫测定方式用于选择与特定蛋白质发生特异性免疫反应的抗体。例如,通常将固相ELISA免疫测定法、蛋白质印迹或免疫组织化学法用于选择与蛋白质发生特异性免疫反应的单克隆抗体。参见Harlow和Lane((1988)《抗体-实验室手册》(Antibodies,A LaboratoryManual),Cold Spring Harbor Publications,New York“Harlow和Lane”)对可以用于测定特异性免疫反应性的免疫测定方式和条件的描述。一般来说,特异性或选择性反应将是背景信号或噪声的至少2倍且更常见的是背景的10-100倍以上。
特定核酸序列的“保守修饰的变化形式”指的是编码相同或基本上相同的氨基酸序列或其中核酸序列不编码氨基酸序列的那些核酸序列。由于遗传密码的简并性,所以大量功能相同的核酸编码任何给定的多肽。例如,密码子CGT、CGC、CGA、CGG、AGA和AGG均编码氨基酸精氨酸。因此,在用密码子表示的精氨酸的每一位上,可以将密码子改变成任何相应的所述密码子而不会改变所编码的蛋白质。这类核酸变化形式是“沉默变化形式”,它们是“保守修饰变化形式”的一个种类。除非另有说明,编码蛋白质的本文所述的每个核酸序列也描述了每一种可能的沉默变化形式。本领域技术人员认为可以修饰核酸中的各密码子(ATG除外,它通常仅是甲硫氨酸的密码子)以便通过标准技术得到功能相同的分子。因此,编码蛋白质的核酸的各“沉默变化形式”在各所述序列中是隐含的。
此外,本领域技术人员认识到在所编码的序列中改变、添加或缺失单个氨基酸或较小百分比的氨基酸(一般低于5%、更常见的是低于1%)的各个取代、缺失或添加是“保守修饰的变化形式”,其中这种改变导致氨基酸被化学上相似的氨基酸所取代。提供功能相似氨基酸的保守取代表在本领域中是众所周知的。下列5组中各包含属于相互保守取代的氨基酸:脂族:甘氨酸(G)、丙氨酸(A)、缬氨酸(V)、亮氨酸(L)、异亮氨酸(I);芳香族:苯丙氨酸(F)、酪氨酸(Y)、色氨酸(W);含硫的:甲硫氨酸(M)、半胱氨酸(C);碱性的:精氨酸(R)、赖氨酸(K)、组氨酸(H);酸性的:天冬氨酸(D)、谷氨酸(E)、天冬酰胺(N)、谷氨酰胺(Q)。另外参见Creighton(1984)《蛋白质》(Proteins),W.H.Freeman和Company。此外,在所编码的序列中改变、添加或缺失单个氨基酸或较小百分比的氨基酸的各个取代、缺失或添加也是“保守修饰的变化形式”。
“亚序列”指的是分别包括核酸或氨基酸的较长序列(例如蛋白质)的一部分的核酸或氨基酸序列。
当将其它核苷酸(或其它类似分子)引入核酸时核酸是“延长的”。最常见的是使用聚合酶(例如DNA聚合酶)、例如在核酸3’末端添加序列的聚合酶进行该过程。
当将来自两种核酸中每一种的序列在子代核酸中组合时两种核酸被“重组”。当这两种核酸是用于重组的底物时可“直接”重组两条序列。当使用诸如交换寡核苷酸这样的中间体重组所述序列时可“间接重组”两条序列。就间接重组而言,仅仅一条序列是用于重组的真正底物且在某些情况下两条序列都不是用于重组的底物。
例如配体和受体这样的两种分子之间的“特异性结合亲和力”指的是在分子混合物中一种分子对另一种分子的优先结合。如果结合亲和力约为1×104M-1-约1×106M-1或1×106M-1以上,那么可以将该分子的结合看作是特异性的。
转化:一种用于将异源DNA引入宿主细胞或生物体的过程。
“转化的”、“转基因的”和“重组的”指的是诸如细菌或植物这样已经引入了异源核酸分子的宿主生物体。核酸分子可以被稳定地整合入宿主的基因组或也可以作为染色体外分子存在。这类染色体外分子可以是自体复制的。认为转化的细胞、组织或植物不仅包括转化过程的终产物而且包括其转基因子代。“非转化的”、“非转基因的”或“非重组的”宿主指的是例如细菌或植物这样不含异源核酸分子的野生型生物体。
本发明通过提供几种来自另外的植物物种的鼠耳芥属NIM1基因同源物而满足了上述需求。特别地,本发明涉及NIM1基因的烟草(Nicotiana tabacum)、番茄(Lycopericon esculentum)、欧洲油菜(Brassica napus)、鼠耳芥(Arabidopsis thaliana)、甜菜(Beta vulgaris)、向日葵(Helianthus annuus)和马铃薯(Solanumtuberosum)的同源物的分离,这些同源物编码认为与信号转导级联有关的蛋白质,所述的信号转导级联对可导致植物中系统获得性抗性的生物和化学诱导物敏感。
因此,本发明涉及一种分离的核酸分子,它包括编码SEQ ID NO:2、4、6、8、16、18、20、30、32、34、36、38、40、42、44、46、48、50、52、54、56、58、62、64、66、68、70、72或74的核苷酸序列。
在另一个实施方案中,本发明涉及一种包括SEQ ID NO:1、3、5、7、15、17、19、29、31、33、35、37、39、41、43、45、47、49、51、53、55、57、61、63、65、67、69、71或73的分离的核酸分子。
在另一个实施方案中,本发明涉及一种包括核苷酸序列的分离的核酸分子,所述的核苷酸序列在序列中包括与SEQ ID NO:1、3、5、7、15、17、19、29、31、33、35、37、39、41、43、45、47、49、51、53、55、57、61、63、65、67、69、71或73的至少20、25、30、35、40、45或50个(优选20个)连续碱基对部分相同的至少20、25、30、35、40、45或50个(优选20个)连续的碱基对部分。
在另一个实施方案中,本发明涉及一种包括核苷酸序列的分离的核酸分子,可以使用应用表示为SEQ ID NO:9和10、SEQ ID NO:21和24、SEQ ID NO:22和24、SEQ ID NO:25和28、SEQ ID NO:26和28或SEQ ID NO:59和60的引物对的聚合酶链反应从番茄DNA文库扩增所述的核苷酸序列;
在另一个实施方案中,本发明涉及一种包括核苷酸序列的分离的核酸分子,可以使用应用表示为SEQ ID NO:22和24或SEQ ID NO:26和28的引物对的聚合酶链反应从甜菜DNA文库扩增所述的核苷酸序列;
在另一个实施方案中,本发明涉及一种包括核苷酸序列的分离的核酸分子,可以使用应用表示为SEQ ID NO:26和28的引物对的聚合酶链反应从向日葵DNA文库扩增所述的核苷酸序列;
在另一个实施方案中,本发明涉及一种包括核苷酸序列的分离的核酸分子,可以使用应用表示为SEQ ID NO:21和24、SEQ ID NO:21和23、SEQ ID NO:22和24、SEQ ID NO:25和28或SEQ ID NO:26和28的引物对的聚合酶链反应从马铃薯DNA文库扩增所述的核苷酸序列;
在另一个实施方案中,本发明涉及一种包括核苷酸序列的分离的核酸分子,可以使用应用表示为SEQ ID NO:9和10或SEQ ID NO:26和28的引物对的聚合酶链反应从欧洲油菜DNA文库扩增所述的核苷酸序列;
在另一个实施方案中,本发明涉及一种包括核苷酸序列的分离的核酸分子,可以使用应用表示为SEQ ID NO:13和14、SEQ ID NO:21和24或SEQ ID NO:22和24的引物对的聚合酶链反应从鼠耳芥DNA文库扩增所述的核苷酸序列;
在另一个实施方案中,本发明涉及一种包括核苷酸序列的分离的核酸分子,可以使用应用表示为SEQ ID NO:9和10、SEQ ID NO:11和12、SEQ ID NO:21和24、SEQ ID NO:22和24、SEQ ID NO:25和28或SEQ ID NO:26和28的引物对的聚合酶链反应从烟草DNA文库扩增所述的核苷酸序列;或
在另一个实施方案中,本发明涉及一种包括核苷酸序列的分离的核酸分子,可以使用应用引物对的聚合酶链反应从植物DNA文库扩增所述的核苷酸序列,所述的引物对包括SEQ ID NO:1、3、5、7、15、17、19、29、31、33、35、37、39、41、43、45、47、49、51、53、55、57、61、63、65、67、69、71或73编码序列(CDS)的前20个核苷酸和最后20个核苷酸的反向补体。
本发明还包括一种嵌合基因,它包括在植物中具有活性的与本发明NIM1同源物编码序列可操作地连接的启动子;本发明还包括含有这类嵌合基因的重组载体,其中所述的载体能够稳定地转化入宿主且可以用这类载体稳定地转化宿主。优选所述的宿主是诸如下列农学上重要的农作物之一的植物:水稻、小麦、大麦、黑麦、玉米、马铃薯、canola、向日葵、胡萝卜、甘薯、甜菜、菜豆、豌豆、菊苣、莴苣、甘蓝、花椰菜、嫩茎花椰菜、芜菁、萝卜、菠菜、芦笋、洋葱、大蒜、茄子、胡椒、芹菜、南瓜、南瓜、黄瓜、苹果、梨、温柏、甜瓜、洋李、樱桃、桃、油桃、杏、草莓、葡萄、覆盆子、黑莓、凤梨、鳄梨、番瓜、芒果、香蕉、大豆、烟草、番茄、高粱和糖甘蔗。本发明还包括来自本发明植物的种子。
此外,本发明涉及一种增加植物中SAR基因表达的方法,该方法通过在所述植物中表达自身含有在植物中具有活性的与本发明NIM1同源物编码序列可操作地连接的启动子的嵌合基因来进行,其中在转化的植物中以高于野生型植物的水平表达所编码的蛋白质。
此外,本发明涉及一种提高植物中病害抗性的方法,该方法通过在所述植物中表达自身含有在植物中具有活性的与本发明NIM1同源物编码序列可操作地连接的启动子的嵌合基因来进行,其中在转化的植物中以高于野生型植物的水平表达所编码的蛋白质。
此外,本发明涉及一种选自SEQ ID NO:9-14、21-28、59和60组成的组的PCR引物。
本发明还包括一种用于分离与可导致植物中系统获得性抗性的信号转导级联相关的NIM1同源物的方法,该方法包括使用应用引物对的聚合酶链反应从植物DNA文库扩增DNA分子,其中所述的引物对相当于SEQ ID NO:1、3、5、7、15、17、19、29、31、33、35、37、39、41、43、45、47、49、51、53、55、57、61、63、65、67、69、71或73编码序列(CDS)的前20个核苷酸和最后20个核苷酸的反向补体;或所述的引物对表示为SEQ ID NO:9和10、SEQ ID NO:11和12、SEQID NO:13和14、SEQ ID NO:21和24、SEQ ID NO:22和24、SEQ ID NO:21和23、SEQ ID NO:25和28、SEQ ID NO:26和28或SEQ ID NO:59和60。在一个优选的实施方案中,所述的植物DNA文库是烟草(Nicotianatabacum)、番茄(Lycopericon esculentum)、欧洲油菜(Brassicanapus)、鼠耳芥(Arabidopsis thaliana)、甜菜(Beta vulgaris)、向日葵(Helianthus annuus)和马铃薯(Solanum tuberosum)的DNA文库。
有关本文所述的几种NIM1同源物的Northern数据表示了组成型表达或BTH-诱导性。据推定本文所述的NIM1基因同源物编码对生物和化学诱导物敏感的信号转导级联中所涉及的蛋白质,所述的信号转导级联可在植物中产生系统获得性抗性。本发明还涉及植物中这类NIM1同源物的转基因表达以便增加SAR基因表达并增强病害抗性。
可以使用下面的实施例中所述的技术或通过使用作为构建PCR引物基础的序列表中所示的序列的PCR来分离本发明的DNA序列。例如,可以将具有SEQ ID NO:7的大约前和最后20-25个连续核苷酸的序列的寡核苷酸用作PCR引物以便从来自植物来源(鼠耳芥)的cDNA文库直接扩增cDNA序列(SEQ ID NO:7)。同样可以使用作为PCR引物基础的序列表中所示的DNA序列末端、通过PCR从相应植物的cDNA或基因组DNA文库中扩增本发明的其它DNA序列。
据推定在植物中转基因表达本发明的NIM1同源物可使一系列广泛的植物病原体产生免疫性,所述的植物病原体包括但不限于:病毒或类病毒,例如烟草或黄瓜花叶病毒、环斑病毒或坏死病毒、天竺葵叶卷曲病毒、红色三叶草环斑病毒、番茄丛矮病毒等病毒;真菌,例如卵菌诸如寄生疫霉和烟草霜霉;细菌,例如丁香假单胞菌和烟草野火病假单胞菌;昆虫、诸如蚜虫,例如桃短尾蚜;和鳞翘目,例如Heliothus的种类;以及线虫,例如南方根结线虫。本发明的载体和方法对大量玉米病害生物体来说是有用的、包括但不限于霜霉:诸如;大孢指疫霉、Sclerophthora rayissiae、禾生指梗霉、Pernosclerospora sorghi、Peronosclerospora philippinensis、Peronosclerospora sacchari和Peronosclerospora maydis;锈菌诸如高粱柄锈菌、多堆柄锈菌和Physopella zeae;其它真菌诸如Cercospora zeae-maydis、禾生刺盘孢、串珠镰孢、玉蜀黍赤霉、Exerohilum turcicum、玉蜀黍球梗孢、禾白粉菌、壳针孢属和Bipolaris maydis;和细菌诸如斯获氏欧文氏杆菌。
本发明的方法可以用于将病害抗性赋予各种植物,包括裸子植物、单子叶植物和双子叶植物。尽管可以使属于这些宽类别的任何植物产生病害抗性,但是它特别适用于农学上重要的农作物植物,诸如水稻、小麦、大麦、黑麦、油菜、玉米、马铃薯、胡萝卜、甘薯、甜菜、菜豆、豌豆、菊苣、莴苣、甘蓝、花椰菜、嫩茎花椰菜、芜菁、萝卜、菠菜、芦笋、洋葱、大蒜、茄子、胡椒、芹菜、胡萝卜、南瓜、南瓜、绿皮密生西葫芦、黄瓜、苹果、梨、温柏、甜瓜、杧果、李子、樱桃、桃、蜜桃、杏、草莓、葡萄、覆盆子、黑莓、凤梨、鳄梨、番瓜、杧果、香蕉、大豆、烟草、番茄、高粱和甘蔗。
可以将本发明的NIM1同源物编码序列插入为植物设计的表达弹夹以便使用标准遗传工程技术构建本发明的嵌合基因。对适合于实现所选植物宿主中所需模式和水平的表达的诸如启动子、信号序列、5’和3’未翻译序列和增强子这样的特异性调节序列的选择属于本领域普通技术人员的水平范围内。可以将含有在合适读框中连接的各个元件的所得分子插入能够转化至宿主植物细胞的载体中。
能够在植物或植物细胞中起作用的启动子的实例(即那些能够启动相关编码序列表达的启动子、诸如那些编码植物细胞中NIM1同源物的启动子)包括:鼠耳芥属和玉米遍在蛋白质启动子;花椰菜花叶病毒(CaMV)19S或35S启动子和CaMV双启动子;水稻肌动蛋白启动子;来自烟草、鼠耳芥属或玉米的PR-1启动子;胭脂氨酸合酶启动子;核酮糖-1,5-二磷酸羧化酶小亚单位(ssuRUBISCO)启动子等。特别优选的是鼠耳芥属遍在蛋白质启动子。可以修饰启动子自身以便控制启动子强度,从而按照本领域已知的方法提高相关编码序列的表达。优选用于本发明的启动子是那些产生高水平组成型表达的启动子。
可以使信号或转运肽类与本发明的嵌合DNA构建体中的NIM1同源物编码序列融合以便使所表达的蛋白质定向转运至所需的作用部位。信号肽类的实例包括那些与植物发病机理相关蛋白质天然连接的信号肽,例如PR-1、PR-2等。参见,例如Payne等,1988。转运肽类的实例包括诸如那些在Von Heijne等(1991)、Mazur等(1987)和Vorst等(1988)中所述的叶绿体转运肽类;以及诸如那些在Boutry等(1987)中所述的线粒体转运肽类。另外还包括使所编码的蛋白质集中于诸如液泡这样的各种细胞区室中的序列。例如,参见Neuhaus等(1991)和Chrispeels(1991)。
本发明的嵌合DNA构建体可以含有本发明的多拷贝的启动子或多拷贝的NIM1同源物编码序列。此外,该构建体可以包括标记物的编码序列和诸如信号或转运肽类这样的其它肽类的编码序列,它们各自在带有DNA分子中的其它功能元件的合适的读框中。这类构建体的制备方法属于本领域普通技术人员的水平范围内。
有用的标记物包括提供除草剂、抗生素或药物抗性的肽类,诸如:例如耐原卟啉原氧化酶抑制剂、潮霉素、卡那霉素、G418、庆大霉素、洁霉素、氨甲蝶呤、草甘膦、膦丝菌素等。可以将这些标记物用于从来转化的细胞中筛选用本发明嵌合DNA构建体转化的细胞。其它有用的标记物是可以通过例如颜色反应这样的可肉眼观察的反应方便检测的肽酶,例如荧光素酶、β-葡糖醛酸糖苷酶或β-半乳糖苷酶。
可以用多种已知方法将诸如本文所述的那些用于植物表达的嵌合基因引入植物细胞。本领域技术人员会理解对方法的选择可能取决于为转化而靶向的植物(即单子叶植物或双子叶植物)和/或细胞器(即细胞核、叶绿体、线粒体)的类型。转化植物细胞的适宜方法包括微量注射(Crossway等,1986)、电穿孔(Riggs等,1986)、农杆菌属介导的转化(Hinchee等,1988;Ishida,1996)、定向基因转移(Paszkowski等,1984;Hayashimoto等,1990)和使用获自Agracetus,Inc.,Madison,Wisconsin and Dupont,Inc.,Wilmington,Delaware的装置进行的冲击粒子加速(参见,例如美国专利4,945,050和McCabe等,1988)。另外参见:Weissinger等(1988);Sanford等(1987)(洋葱);Christou等(1988)(大豆);McCabe等(1988)(大豆);Datta等(1990)(水稻);Klein等(1988)(玉米);Klein等(1988)(玉米);Klein等(1988)(玉米);Fromm等(1990);和Gordon-Kamm等(1990)(玉米);Svab等(1990)(烟草叶绿体);Gordon-Kamm等(1993)(玉米);Shimamoto等(1989)(水稻);Christou等(1991)(水稻);Datta等(1990)(水稻);欧洲专利申请EP 0 332581(鸭茅属和其它Pooideae);Vasil(1993)(小麦);Weeks等(1993)(小麦);Wan等(1994)(大麦);Jahne等(1994)(大麦);Umbeck等(1987)(棉花);Casas等(1993)(高粱);Somers等(1992)(燕麦);Torbert等(1995)(燕麦);Weeks等(1993)(小麦);WO 94/13822(小麦);和Nehra等(1994)(小麦)。在Koziel等(1993)、Hill等(1995)和Koziel等(1996)中可以找到通过微粒轰击将重组DNA分子引入玉米的一组特别优选的实施方案。另一个优选的实施方案是公开在EP 0 292 435中的玉米的原生质体转化方法。
一旦将包括NIM1同源物编码序列的嵌合基因转化入特定的植物物种,则可以使用传统的育种技术使之在该物种中繁殖或将其移入相同物种的其它的变种、特别包括商品变种中。本发明特别优选的植物包括上面所列的农学上重要的农作物。通过有性繁殖来传递上述基因工程的转基因种子和植物的遗传特性且这些特性由此可以在子代植物中得到维持和传播。
实施例
通过下列具体的步骤、制备方法和实施例来进一步具体地解释本发明。这些实施例仅用于解释目的而非用来限定本发明的范围。本文所用的标准重组DNA和分子克隆技术在本领域中是众所周知的,并且在Sambrook等人1989;T.J.Silhavy,M.L.Berman;L.W.Enquist,1984和Ausubel,F.M.等人1987中有描述。
I.鼠耳芥属NIMI基因同源物的分离
实施例1:来自烟草的NIM1同源物的分离
将来自从烟草cDNA文库的噬菌体切下的团块的质粒DNA用作使用下列引物对进行的PCR的模板:5’-AGATTATTGTCAAGTCTAATG-3’(SEQ IDNO:9)+5’-TTCCATGTACCTTTGCTTC-3’(SEQ ID NO:10)和5’-GCGGATCCATGGATAATAGTAGG-3’(SEQ ID NO:11)+5’-GCGGATCCTATTTCCTAAAAGGG-3’(SEQ ID NO:12)。循环条件优选94℃下1分钟、40℃下1分钟和72℃下1.5分钟,并且该反应优选进行40个循环。将琼脂糖凝胶上的PCR产物放出、切下并克隆入pCRII-TOPO(Invitrogen)。
这种烟草NIM1同源物的全长cDNA序列表示为SEQ ID NO:1,并且由该cDNA序列编码的蛋白质表示为SEQ ID NO:2。已经在1998年8月17日将包含SEQ IDNO:1的烟草NIM1同源物在NRRhL(AgriculturalResearch Service,Patent Culture Collection,Northern RegionalResearch Center,1815 North University Street,Peoria,lllinois61604,U.S.A.)以pNOV1206保藏,并给予其保藏号为NRRL B-30051。
实施例2:来自番茄的NIM1同源物的分离
使用来自Stratagene的方案从番茄的λZAPII cDNA文库中切下噬菌粒。将噬菌粒(质粒)大量转化入各含有约80,000个克隆的10个库内的大肠杆菌XL1-Blue中,并从这些库中提取DNA。通过使用下列引物进行的PCR筛选所述库中存在的NIM1同源物:5’-AGATTATTGTCAAGTCTAATG-3’(SEQ ID NO:9)和5’-TTCCATGTACCTTTGCTTC-3’(SEQ ID NO:10)。
通过克隆PCR-扩增的DNA片段并测序证实了从所述库中扩增的序列含有NIM1同源物。连续制备较小的库并通过使用上述相同引物的PCR来筛选存在的NIM1同源物,直到获得含有该同源物的单克隆为止。在cDNA克隆含有缺少5’端的部分基因的情况中,采用5’RACE(cDNA末端的快速扩增)获得所述基因的全长序列。
这种番茄NIM1同源物的全长cDNA序列表示为SEQ ID NO:3,并且由该cDNA序列编码的蛋白质表示为SEQ ID NO:4。已经在1998年8月17日将包含SEQ IDNO:3的番茄NIM1同源物在NRRL(AgriculturalResearch Service,Patent Culture Collection,Northern RegionalResearch Center,1815 North University Street,Peoria,lllinois61604,U.S.A.)以pNOV1204保藏,并给予其保藏号为NRRL B-30050。实施例3:来自欧洲油菜的NIM1同源物的分离
使用来自Stratagene的方案从欧洲油菜的λZAPII cDNA文库中切下噬菌粒。将噬菌粒(质粒)大量转化入各含有约80,000个克隆的10个库内的大肠杆菌XL1-Blue中,并从这些库中提取DNA。通过PCR筛选所述库中通过使用序列引物进行的PCR而存在的NIM1同源物:5’-AGATTATTGTCAAGTCTAATG-3’(SEQ ID NO:9)和5’-TTCCATGTACCTTTGCTTC-3’(SEQ ID NO:10)。
通过克隆PCR-扩增的DNA片段并测序证实了从所述库中扩增的序列含有NIM1同源物。连续制备较小的库并通过使用上述相同引物的PCR来筛选存在的NIM1同源物,直到获得含有该同源物的单克隆为止。在cDNA克隆含有缺少5’端的部分基因的情况中,采用5’RACE(cDNA末端的快速扩增)获得所述基因的全长序列。
这种欧洲油菜NIM1同源物的部分cDNA序列表示为SEQ ID NO:5,并且由该cDNA序列编码的蛋白质表示为SEQ ID NO:6。已经在1998年8月17日将包含SEQ IDNO:5的欧洲油菜NIM1同源物在NRRL(Agricultural Research Service,Patent Culture Collection,Northern Regional Research Center,1815 North University Street,Peoria,lllinois 61604,U.S.A.)以pNOV1203保藏,并给予其保藏号为NRRL B-30049。
实施例4:来自鼠耳芥的NIM1同源物的分离
BLAST在查询检测GenBank条目B26306时使用鼠耳芥属或番茄NIM1氨基酸序列检索,B26306含有来自细菌人工染色体(BacterialArtifical Chromosome)(BAC)F18D8的鼠耳芥属基因组序列。预测部分BAC序列编码与NIM1具有显著相似性(47%氨基酸同一性)的蛋白质。将下列引物用于F18D8序列区:5’-TCAAGGCCTTGGATTCAGATG-3’(SEQ ID NO:13)和5’-ATTAACTGCGCTACGTCCGTC-3’(SEQ ID NO:14)。
将所述引物用于与作为模板的来自以pFL61-为基础的鼠耳芥属cDNA文库的DNA的PCR反应。循环条件优选94℃下30秒、53℃下30秒、72℃下30秒。该反应优选进行40个循环。检测预测大小(290个碱基对)的PCR产物并通过使逐渐减小量的文库经过大肠杆菌的连续纯化和重新鉴别通过PCR存在的克隆而从cDNA文库中纯化相应于F18D8引物的cDNA克隆。最终获得单一阳性克隆并测序。该克隆的序列证实了存在与NIM1具有显著同源性的可读框。
这种鼠耳芥NIM1同源物的全长cDNA序列表示为SEQ ID NO:7,并且由该cDNA序列编码的蛋白质表示为SEQ ID NO:8。已经在1999年5月25日将包含SEQ IDNO:7的鼠耳芥NIM1同源物在NRRL(Agricultural Research Service,Patent Culture Collection,Northern Regional Research Center,1815 North University Street,Peoria,lllinois 61604,U.S.A.)以AtNMLc5保藏,并给予其保藏号为NRRL B-30139。
实施例5:简并引物的设计
除NIM1基因(Ryals等人,1997)和上述实施例4中所述的NIM-样基因(AtNMLc5-SEQ ID NO:7)外,鼠耳芥还含有三种其它NIM-样(NML)基因组序列:AtNMLc2(SEQ ID NO:15)、AtNMLc4-1(SEQ IDNO:17)和AtNMLc4-2(SEQ ID NO:19),其中c[#]代表特定NML基因位于其上的染色体号。使用GCG Seqweb多重序列对比程序(Pretty,Wisconsin Genetics Computer Group),将来自鼠耳芥(Ryals等,1997)、烟草(实施例1-SEQ ID NO:1)和番茄(实施例2-SEQ ID NO:3)的NIM1序列以及来自鼠耳芥的NML序列(SEQ ID NO:7、15、17和19)进行序列对比。以这种序列对比为基础,三个区显示出充分的保守性以便设计对来自包括甜菜、向日葵、马铃薯和canola在内的其它农作物物种的NIM1同源物进行PCR扩增所用的简并PCR引物。将根据这些保守区设计的引物列于下面的表1中。使用仅带有来自鼠耳芥(Ryals等,1997)、烟草(实施例1-SEQ ID NO:1)和番茄(实施例2-SEQID NO:3)的NIM1序列的序列组来设计NIM1(A-D)引物。使用除来自鼠耳芥的四种NML序列(SEQ ID NO:7、15、17和19)之外还带有上述三种序列的序列组来设计NIM2(A-D)引物。优选由GenosysBiotechnologies,Inc.(The Woodlands,Texas)来合成引物。根据寡核苷酸中单一位点上一种以上碱基的标志将简并位置列在表1中。“取向”指的是所述引物是否定向于cDNA的3’端(下游)或5’端(上游)。表1:简并引物
引物 | 序列(5’-3’) | SEQ ID NO: | 取向 |
NIM 1A | GAGATTATTGTCAAGTCTAATGTAGATAT T | SEQ ID NO:21 | 下游 |
NIM 1B | ACTGGACTCGGATGATATTGAATTAT T T T G G | SEQ ID NO:22 | 下游 |
NIM 1C | TAACTCAACATCATCAGAATCAAATGCT T C G C G | SEQ ID NO:23 | 上游 |
NIM 1D | GTTGAGCAAGAGCAACTCTATTTTCAAGT C CCGT | SEQ ID NO:24 | 上游 |
NIM 2A | TGCATAGAAATAATTGTGAAGTCTAATGTAGAT G TG C G T | SEQ ID NO:25 | 下游 |
NIM 2B | GGCACTGGACTCAGATGATGTTGAACTT T T GT | SEQ ID NO:26 | 下游 |
NIM 2C | AACTCAACATCATCAGAATCCAATGCCGT T G G | SEQ ID NO:27 | 上游 |
NIM 2D | AGTTGAGCAAGGCCAACTCGATTTTCAAAATT C A T GGT | SEQ ID NO:28 | 上游 |
实施例6:对来自农作物物种NIM-样DNA片段的PCR扩增
使用基因组DNA或cDNA作为模板从鼠耳芥属、番茄、烟草、甜菜、向日葵、马铃薯和canola扩增NIM-样DNA片段。将所用的引物组合以及预计的片段大小列在下面的表2中。表2:引物组合和DNA片段大小
左引物 | 右引物 | 片段大小(bp) |
NIM 1A | NIM 1D | 669 |
NIM 1A | NIM 1C | 195 |
NIM 1B | NIM 1D | 499 |
NIM 2A | NIM 2D | 676 |
NIM 2A | NIM 2C | 200 |
NIM 2B | NIM 2D | 503 |
优选在GeneAmp PCR System 9700(PE Applied Biosystems,Foster City,CA)中使用Ready-To-Go PCR Beads(Amersham,Piscataway,NJ)进行简并引物的PCR。在各反应中使用20-40ng的基因组DNA或5-10ng的cDNA,其中各引物的终浓度为0.8μM。优选的循环参数如下:94℃1分钟;3个循环的[94℃30秒;37℃30秒;72℃2分钟];35个循环的[94℃30秒;60℃30秒;72℃2分钟];72℃7分钟;4℃保持。在2%琼脂糖凝胶上分析反应产物并切下适当大小的DNA片段。例如使用Geneclean III试剂盒(BIO 101,Inc.,Carlsbad,CA)从琼脂糖带上分离DNA片段,例如使用TOPO TA克隆试剂盒(Invitrogen Corporation,Carlsbad,CA)对其进行克隆。例如,使用CONCERT快速质粒小量制备系统(Life Technologies,Inc.,Rockyille,MD)分离质粒并通过标准方案测序。
NIM-样DNA片段获自所尝试的所有植物物种且在许多情况中分离了多种单一NIM-样序列。表3和附图2具体描述了所分离的NIM-样片段。表3:NIM-样PCR片段
物种 | 成功的引物对 | PCR模板 | 单一克隆 | SEQ ID NO: |
鼠耳芥属 | 1A/1D;1B/1D | 基因组DNA | 1 | |
烟草 | 1A/1D;1B/1D;2A/2D;2B/2D | cDNA | 4 | SEQ ID NO:29、31、33和35 |
番茄 | 1A/1D;1B/1D;2A/2D;2B/2D | 基因组DNA,cDNA | 1 | SEQ ID NO:37 |
甜菜 | 1B/1D;2B/2D | 基因组DNA,cDNA | 1 | SEQ ID NO:39 |
向日葵 | 2B/2D | cDNA | 2 | SEQ ID NO:41和43 |
马铃薯 | 1A/1D;1A/1C;1B/1D;2A/2D;2B/2D | cDNA | 3 | SEQ ID NO:45、47和49 |
Canola | 2B/2D | cDNA | 4 | SEQ ID NO:51、53、55和57 |
以这些结果为基础,上述简并引物PCR可以从各种植物物种扩增NIM-样片段。特别地,使用来自所尝试的所有物种的cDNA作为模板的NIM 2B/NIM 2D的引物组合是成功的。特别优选使用Ready-To-Go PCRBeads来获得产物。此外,对除鼠耳芥属、番茄和甜菜外的所有样品来说优选使用cDNA作为模板,其中基因组DNA是充分的。
实施例7:其它简并引物
以4种烟草片段(SEQ ID NO:29、31、33和35)和用于测定番茄是否还含有不使用表1中所列简并引物扩增的相似NIM-样序列的番茄序列(SEQ ID NO:37)的序列对比为基础设计新简并引物对。将由这些片段设计的引物列在下面的表3中且优选由GenosysBiotechnologies,Inc.(The Woodlands,Texas)合成。根据寡核苷酸中单一位点上一种以上碱基的标志将简并位置列在表3中。“取向”指的是所述引物是否定向于cDNA的3’端(下游)或5’端(上游)。表4:其它简并引物
引物 | 序列(5’-3’) | SEQ ID NO: | 取向 |
NIM 3A | TAGATGAAGCATACGCTCTCCACTATGCTGTT C T T T | SEQ ID NO:59 | 下游 |
NIM 3B | GGCTCCTTACGCATGGCAGCAACATGAAGGACT C T TG C | SEQ ID NO:60 | 上游 |
如上所述使用番茄cDNA进行简并引物的PCR并克隆和测序可能的产物。序列分析显示了两类NIM-样片段:第一种与用SEQ ID NO:37表示的番茄序列相同,而第二种在番茄中是唯一的,并且与用SEQ IDNO:31和33表示的烟草序列具有88%的同一性。这种新型番茄序列的序列用SEQ ID NO:61表示。
实施例8:全长NIM-样cDNA’s
优选通过使用SMART RACE cDNA扩增试剂盒(Clontech,Palo Alto,CA)的RACE PCR从NIM-样PCR片段中获得相应的cDNA序列上游和下游。优选对至少三种独立的RACE产物的每个5’-或3’-末端测序,以便消除PCR误差。分别将所得的符合用SEQ ID NO:39、43和31表示的NIM-样PCR片段的甜菜、向日葵B和烟草B NIM1同源物的全长cDNA序列表示为SEQ ID NO:63、65和73。
优选通过RT-PCR克隆符合NIM-样基因组序列AtNMLc2(SEQ IDNO:15)、AtNMLc4-1(SEQ ID NO:17)和AtNMLc4-2(SEQ ID NO:19)的NIM-样鼠耳芥cDNA’s。使用寡脱氧胸苷酸引物反转录来自鼠耳芥的总RNA。使用以各基因组序列(SEQ ID NO:15、17和19)编码区的5’和3’末端为基础设计的特异性有义和反义寡核苷酸引物、通过PCR来扩增所得的第一链cDNA。将预测大小的PCR片段克隆入载体并测序以便证实这些cDNA克隆与NIM-样基因组序列一致。将与NIM-样基因组序列AtNMLc2(SEQ ID NO:15)一致的cDNA序列表示为SEQ ID NO:67;将与NIM-样基因组序列AtNMLc4-1(SEQ ID NO:17)一致的全长cDNA序列表示为SEQ ID NO:69;并将与NIM-样基因组序列AtNMLc4-2(SEQ ID NO:19)一致的全长cDNA序列表示为SEQ ID NO:71。
实施例9:Northern分析
Northern数据表明在用100μM或300μM BTH(苯并(1,2,3)噻二唑-7-硫代羧酸S-甲酯)处理后甜菜NIM-样克隆(SEQID NO:39和63)的表达提高了3-7倍。此外,Northern数据表明向日葵A NIM-样克隆(SEQ ID NO:41)的表达是组成型的。此外,Northern数据表明在用100μM或300μM BTH处理后向日葵B NIM-样克隆(SEQ ID NO:43和65)的表达提高了2倍。
II.植物中本发明基因序列的表达
可以使用常规的重组DNA技术将本发明的NIM1同源物引入植物细胞。一般来说,这一过程包括使用本领域中公知的标准克隆步骤将本发明的编码序列插入与该编码序列是异源的(即正常情况下不存在的)表达系统。所述载体含有转录和翻译所插入编码蛋白质的序列所必不可少的元件。可以使用本领域中公知的大量载体系统,诸如质粒、噬菌体病毒和其它修饰的病毒。合适的载体包括但不限于:病毒载体,诸如λ载体系统λgtl1、λgtl0和卡隆4;质粒载体,诸如pBI121、pBR322、pACYC177、pACYC184、pAR系列、pKK223-3、pUC8、pUC9、pUC18、pUC19、pLG339、pRK290、pKC37、pKC101、pCDNAII和其它相似系统。还可以修饰所述表达系统的成分以便增加表达。例如,可以使用截短的序列、核苷酸取代或其它修饰方法。在合适的条件下可以采用本文所述的表达系统转化实际上任何的农作物植物细胞。可以使转化的细胞再生成完整植物,使得NIM1同源物增加SAR基因表达并提高转基因植物中的病害抗性。
实施例10:植物表达弹夹的构建
首先在表达弹夹中将用于转基因植物中表达的编码序列装配在植物中可表达的合适启动子之后。该表达弹夹还可以包括表达转基因所需或筛选的任何其它序列。这类序列包括但不限于:转录终止子;诸如内含子这样促进表达的外部序列;活性序列和用于将基因产物靶向至特定细胞器和细胞区室的序列。然后便利地将这些表达弹夹转入下述植物转化载体。下面是典型表达弹夹中各种成分的描述。
1.启动子
对表达弹夹中所用启动子的选择决定转基因植物中转基因的空间和时间分布型。所选择的启动子表达特殊细胞类型(诸如叶表皮细胞、叶肉细胞、根皮层细胞)或特殊组织或器官(例如,根、叶或花)中的转基因,并且这种选择反映出累积基因产物所需的位置。另一方面,所选择的启动子可以在不同诱导条件下启动基因的表达。启动子在其强度即促进转录的能力上是可变的。根据所用宿主细胞系统的不同,可以使用包括基因的天然启动子在内的许多合适启动子中的任何一种。下面是可以用于表达弹夹中的启动子的非限制性实例。
a.组成型表达、遍在蛋白质启动子:
遍在蛋白质是一种已知累积在许多细胞类型中的基因产物且已经从用于转基因植物的几个物种中克隆了其启动子(例如向日葵-Binet等人,1991;玉米-Christensen等人,1989;和鼠耳芥属-Norris等人,1993)。已经在转基因单子叶植物系统中研发了玉米遍在蛋白质启动子且为单子叶植物转化而构建的其序列和载体公开在专利申请EP 0342 926(Lubrizol)中。Taylor等人(1993)描述了包括玉米遍在蛋白质启动子和第一种内含子的载体(pAHC25)及其在通过微射轰击引入时大量单子叶植物的细胞混悬液中的高活性。特别优选将鼠耳芥属遍在蛋白质启动子用于本发明的NIM1同源物中。这种遍在蛋白质启动子适合于包括单子叶植物和双子叶植物在内的转基因植物中的基因表达。合适的载体是pAHC25的衍生物或在该申请中记载的通过引入合适的遍在蛋白质启动子和/或内含子序列而修饰的任何转化载体。
b.组成型表达,CaMV35S启动子:
质粒pCGN1761的构建记载在公开的专利申请EP 0392 225(实施例23)中。pCGN1761含有“双”CaMV35S启动子和tml转录终止子(在该启动子和该终止子之间具有单一EcoRI位点)并带有pUC-型主链。构建带有修饰的多接头的pCGN1761衍生物,所述的多接头除包括存在的EcoRI位点外还包括Notl和Xhol位点。将这种衍生物命名为pCGN1761ENX。pCGN1761ENX用于克隆其多接头内的cDNA序列或编码序列(包括微生物ORF序列)以便在转基因植物中在35S启动子控制下表达所述的序列。可以用与启动子5’连接的Hindlll、Sphl、Sall和Xbal位点和与终止子3’连接的Xbal、BamHl和Bgll位点切割如此构建的完整35S启动子-编码序列-tml终止子弹夹以便转入诸如下述的转化载体。此外,可以通过用Hindlll、Sphl、Sall、Xbal或Pstl进行5’切割并用任何的多接头限制位点(EcoRI、Notl或Xhol)进行3’切割来除去双35S启动子片段,以便用另一启动子进行取代。如果需要,通过引入可以促进翻译的序列来进行克隆位点周围的修饰。这在需要超表达时特别有用。例如,可以如美国专利号5,639,949的实施例37中所述通过使翻译起始位点最佳化来修饰pCGN1761ENX。
c.组成型表达,肌动蛋白启动子
已知在大多数细胞类型中表达了几种肌动蛋白的同种型且由此肌动蛋白启动子对组成型启动子来说是一种良好的选择。特别地,已经克隆和表征了来自水稻Actl基因的启动子(McElroy等人,1990)。发现1.3kb的该启动子片段含有水稻原生质体中表达所需的全部调节元件。此外,已经特异性构建了用于单子叶植物的基于Actl启动子的大量表达载体(McElroy等人,1991)。它们结合了Actl-内含子1、Adhl 5’侧翼序列和Adhl-内含子1(来自玉米醇脱氢酶基因)和来自CaMV35S启动子的序列。显示出最高程度表达的载体是35S和Actl内含子或Actl 5’侧翼序列和Actl内含子的融合体。使起始ATG(GUS报道基因的)周围的序列最佳化也会促进表达。可以方便地修饰由McElroy等人(1991)所述的启动子表达弹夹以用于基因表达,并且这些启动子表达弹夹特别适用于单子叶植物宿主。例如,可以从McElroy构建体中除去含启动子的片段并将其用于取代pCGN1761ENX中的双35S启动子,然后将它用于插入特异性基因序列。接着可以将由此构建的融合基因转入合适的转化载体。在一个独立的报导中,还发现带有第一种内含子的水稻Actl启动子指导经培养的大麦细胞中的高度表达(Chibbar等人,1993)。
d.诱导型表达,PR-1启动子:
可以用产生适宜的高度表达水平的所选择的任何其它启动子来取代pCGN1761ENX中的双35S启动子。根据实例,美国专利号5,614,395中所述的化学调节启动子之一可以取代双35S启动子。优选用限制酶从所选择的启动子来源中切割选择的启动子,不过,可以另外选择使用携带合适终止限制位点的引物来对所选择的启动子进行PCR扩增。应进行PCR扩增,然后应对所述启动子重新进行测序以便检查在靶载体中克隆扩增的启动子后的扩增误差。从质粒pCIB1004中切割化学/病原体调节烟草PR-1a启动子(用于构建,参见:例如EP 0332 104中的实施例21)并将其转入质粒pCGN1761ENX(Uknes等人,1992)。用Ncol切割pCIB1004并通过用T4 DNA聚合酶处理使所得线性化片段的3’突出端钝端化。然后用Hindlll切割该片段并将所得的含PR-1a的启动子片段进行凝胶纯化,并克隆入已经除去了双35S启动子的pCGN1761ENX中。该过程通过下列步骤来进行:用Xhol切割和用T4聚合酶钝端化处理;随后用Hindlll切割并分离含有pCIB1004启动子片段被克隆入的片段的较大载体-终止子。这一过程产生一种pCGN1761ENX衍生物,它带有PR-1a启动子和tml终止子以及带有单一EcoRI和Notl位点的间插多接头。可以将所选择的编码序列插入该载体,随后可以将融合产物(即启动子-基因-终止子)转入包括下文所述的那些载体在内的任何选择的转化载体。可以采用各种化学调节物来诱导所选择的编码序列在按照本发明转化的植物中的表达,这些化学调节物包括美国专利号5,523,311和5,614,395中公开的苯并噻二唑、异烟酸和水杨酸化合物。
e.诱导型表达,一种乙醇-诱导型启动子:
还可以将诸如乙醇这样的某些醇类或酮类诱导的启动子用于使本发明的编码序列产生诱导型表达。这类启动子例如是来自构巢曲霉的alcA基因启动子(Caddick等人,1998)。在构巢曲霉中,alcA基因编码醇脱氢酶I,其表达在有化学诱导物存在的情况下由AlcR转录因子来调节。为了本发明的目的,用本发明的编码序列来取代包括与最小35S启动子(Caddick等人,1998)融合的alcA基因启动子序列的质粒palcA:CAT中的CAT编码序列以便形成带有在alcA基因启动子控制下的编码序列的表达弹夹。可以使用本领域中众所周知的方法来进行该过程。
f.诱导型表达、一种糖皮质激素诱导型启动子:
使用基于类固醇激素的系统诱导本发明NIM1同源物的表达也是所关注的。例如,可以使用糖皮质激素介导的诱导系统(Aoyama和Chua,1997)并且通过应用糖皮质激素诱导基因表达,例如一种合成的糖皮质激素,优选地塞米松,优选的浓度范围在0.1mM-1mM、更优选10mM-100mM。为了本发明的目的,用编码NIM1同源物的基因序列取代荧光素酶基因序列以便在与35S最小启动子融合的GAL4上游活化序列6个拷贝的控制下形成带有编码NIM1同源物的基因序列的表达弹夹。使用本领域中众所周知的方法来进行该过程。反式作用因子包括GAL4 DNA的结合结构域(Keegan等人,1986),该结构域与疱疹病毒蛋白VP16(Triezenberg等人,1988)的反式激活域融合,这种反式激活域与大鼠糖皮质激素受体(Picard等人,1988)的激素结合结构域融合。融合蛋白的表达由适合于本领域中公知或本文所述植物中的表达的任何启动子来控制。这种表达弹夹还包括在含有编码与6xGAL4/最小启动子融合的NIM1同源物的基因序列的植物中。因此,获得融合蛋白的组织或器官特异性可导致NIM1同源物的诱导型组织或器官特异性。
g.根特异性表达:
基因表达的另一种模式是根表达。合适的根启动子由de Framond(1991)描述并且也记载于公开的专利申请EP 0452 269中。将这种启动子转入诸如pCGN1761ENX这样合适的载体以便插入所选择的基因,随后将完整的启动子-基因-终止子弹夹转入有意义的转化载体中。
h.创伤诱导型启动子:
创伤诱导型启动子也适合于基因表达。已经描述了大量这类启动子(例如Xu等人,1993);Logemann等人,1989;Rohrmeier&Lehle,1993;Firek等人,1993;Warner等人,1993),它们均适用于本发明。Logemann等人描述了双子叶植物马铃薯wunl基因的5’上游序列。Xu等人证实来自双子叶植物马铃薯的创伤诱导型启动子(pin2)在单子叶植物水稻中具有活性。此外,Rohrmeier&Lehle描述了玉米WiplcDNA的克隆,它是创伤诱导型的且可以将其用于使用标准技术分离关连启动子。类似地,Firek等人和Warner等人已经描述了来自单子叶植物石刁柏的创伤诱导型基因,将它在局部创伤和病原体侵害部位表达。使用本领域中众所周知的克隆技术可以将这些启动子转入与涉及本发明基因融合的合适载体,并用于在植物创伤部位上表达这些基因。
i.髓优选的表达:
专利申请WO 93/07278中描述了优选在髓细胞中表达的玉米trpA基因的分离。列出了从转录起点扩展至-1726bp的基因序列和启动子。使用标准分子生物学技术可以将这种启动子或其部分转入诸如pCGN1761这样的载体,其中它可以取代35S启动子,并可用于以髓优选方式启动外源基因的表达。实际上,可以将含有髓优选的启动子或其部分的片段转入任何载体并修饰以便在转基因植物中的应用。
j.叶特异性表达:
Hudspeth&Grula(1989)已经描述了编码磷酸烯醇羧酶(PEPC)的玉米基因。使用标准分子生物学技术可以将用于该基因的启动子用于以叶特异性方式启动转基因植物中任何基因的表达。
k.花粉特异性表达:
WO 93/07278中描述了在花粉细胞中表达的玉米钙依赖性蛋白激酶(CDPK)的分离。基因序列和启动子从转录起点扩展至1400bp。使用标准分子生物学技术可以将这种启动子或其部分转入诸如pCGN1761这样的载体,其中它可以取代35S启动子,并可用于以花粉特异性方式启动本发明NIM1同源物的表达。
2.转录终止子
各种转录终止子可以应用于表达弹夹。它们负责转基因及其正确聚腺苷酸化后的转录终止。合适的转录终止子是那些已知在植物中起作用的终止子且包括CaMV 35S终止子、tml终止子、胭脂氨酸合酶终止子和豌豆rbcS E9终止子。它们可用于单子叶植物和双子叶植物。此外,可以使用一种基因的天然转录终止子。
3.用于表达的增强或调节的序列
已经发现了大量来自转录单位内的可促进基因表达的序列,并且可以将这些序列与本发明的基因联合使用以便增加其在转基因植物中的表达。
已经证实不同的内含子序列可促进表达、特别是在单子叶植物细胞中。例如,已经发现玉米Adhl基因的内含子可显著促进野生型基因在引入玉米细胞时在其关连启动子控制下的表达。发现内含子1特别有效且在与氯霉素乙酰基转移酶基因的融合构建体中的表达得到提高(Callis等,1987)。在相同的实验系统中,来自玉米bronzel基因的内含子在促进表达方面具有相似的作用。通常将内含子序列引入植物转化载体、一般是未翻译的前导区内。
已知许多来源于病毒的未翻译前导序列也可促进表达,并且它们在双子叶植物细胞中特别有效。特别地,已经证实来自烟草花叶病毒(TMV,“W-序列”)、玉米褪绿斑驳病(MCMV)和苜蓿花叶病毒(AMV)的前导序列可有效促进表达(例如Gallie等人,1987;Skuzeski等人,1990)。
4.细胞内基因产物的靶向
已知在植物中存在靶向基因产物的不同机理,并且已经以一定的具体方式表征了控制这些机理起作用的序列。例如,由在进入叶绿体以产生成熟蛋白质的过程中被切割的不同蛋白质氨基酸末端发现的信号序列控制将基因产物靶向至叶绿体(例如Comao等人,1988)。这些信号序列可以与异源基因产物融合以便影响异源产物引入叶绿体(van den Broeck等人,1985)。编码合适的信号序列的DNA可以分离自编码RUBISCO蛋白质、CAB蛋白质、EPSP合酶、GS2蛋白质和已知是叶绿体定位的许多其它蛋白质的5’末端。另外参见美国专利号5,639,949的实施例37中的小标题“使用叶绿体靶向的表达”。
其它基因产物定位于其它诸如线粒体和过氧化物酶体这样的细胞器(例如Unger等人,1989)。还可以操纵编码这些产物的cDNAs以便使异源基因产物靶向至这些细胞器。这类序列的实例是核编码的腺苷三磷酸酶和特异性线粒体用的天冬氨酸转氨酶同种型。Rogers等人(1985)已经描述了靶向细胞蛋白质体。
此外,已经表征了可使基因产物靶向至其它细胞区室的序列。氨基末端序列负责靶向至ER、质外体和从糊粉细胞进行胞外分泌(Koehler&Ho,1990)。另外,氨基末端序列与羧基末端序列负责基因产物的液泡靶向(Shinshi等人,1990)。
通过将上述合适的靶向序列与有意义的转基因序列融合,有可能使得转基因产物定向于任何细胞器或细胞区室。例如,为了进行叶绿体靶向,使来自RUBISCO基因、CAB基因、EPSP合酶基因或GS2基因的叶绿体信号序列与转基因的氨基末端ATG进行框内融合。所选择的信号序列应包括已知的切割位点,并且所构建的融合体应考虑到切割所需的切割位点后的任何氨基酸。在某些情况中,通过在切割位点与转基因ATG之间添加少量氨基酸或在转基因序列中取代某些氨基酸可以满足这种要求。通过体外翻译体外转录的构建体、随后使用由Bartlett等人(1982)和Wasmann等人(1986)所述的技术,通过叶绿体体外摄取来测试为引入叶绿体所构建的融合体的叶绿体摄入功效。这些构建技术在本领域中是众所周知的且同样可用于线粒体和过氧化物酶体。
上述细胞靶向机理不仅可以与其关连启动子共同使用,而且可以与异源启动子一起使用,从而在与靶向信号来源的启动子不同的表达模式的启动子转录调节下实现特异性靶向细胞的目的。
实施例11:植物转化载体的构建
大量可以应用于植物转化的转化载体对于植物转化领域的普通技术人员来说是公知的,并且可以将涉及本发明的基因与任何这类载体一起使用。对载体的选择将取决于优选的转化技术和用于转化的靶物种。就某些靶物种而言,可以优选不同的抗生素或除草剂选择标记。在转化中通常使用的选择标记包括产生卡那霉素和相关抗生素抗性的nptll基因(Messing&Vierra,1982;Bevan等人,1983)、产生除草剂膦丝菌素抗性的bar基因(White等人,1990;Spencer等人,1990)、产生抗生素潮霉素抗性的hph基因(Blochinger&Diggelmann)和产生氨甲蝶呤抗性的dhfr基因(Bourouis等人,1983)以及产生草甘膦抗性的EPSPS基因(美国专利号4,940,935和5,188,642)。
1.适合于农杆菌属转化的载体
许多载体可以应用于使用根癌土壤杆菌的转化。它们一般携带至少一种T-DNA边缘序列且包括诸如pBIN19(Bevan,《核酸研究》(Nucl.Acids Res.)(1984))和pXYZ这样的载体。下面描述了适合于农杆菌属转化的两种典型载体的构建。
a.pCIB200和pCIB2001:
二元载体pCIB200和pCIB2001用于构建适用于农杆菌属的重组载体,且以下列方式构建它们。通过Narl消化pTJS75而生成pTJS75kan(Schmidhauser&Helinski,1985),从而切下四环素抗性基因,随后插入来自携带NPTII的pUC4K的Accl片段(Messing&Vierra,1982;Bevan等人,1983;McBride等人,1990)。将Xhol接头与含有左和右T-DNA边缘、植物选择性nos/nptll嵌合基因和pUC多接头的PCIB7的EcoRV片段连接(Rothstein等人,1987),并将Xhol-消化的片段克隆入Sall-消化的pTJS75kan中,以便生成pCIB200(还可参见EP 0 332 104,实施例19)。pCIB200含有下列单一多接头限制位点:EcoRI、Sstl、Kpnl、Bglll、Xbal和Sall。pCIB2001是通过插入其它限制位点的多接头而生成的pCIB200的衍生物。在pCIB2001的多接头中的单一限制位点有:EcoRI、Sstl、Kpnl、Bglll、Xbal、Sall、Mlul、Bcll、Avrll、Apal、Hpal和Stul。pCIB2001除含有这些单一限制位点外还具有植物和细菌卡那霉素选择性、用于农杆菌属介导的转化的左和右T-DNA边缘、用于大肠杆菌与其它宿主之间转移的RK2-衍生的trfA功能和同样来自RK2的OriT和OriV功能。pCIB2001多接头适合于克隆含有其自身调节信号的植物表达弹夹。
b.pCIBi0及其潮霉素选择衍生物:
二元载体pCIB10含有编码植物中筛选用的卡那霉素抗性的基因和T-DNA右和左边缘序列并引入来自广泛宿主范围质粒pRK252的序列,使它在大肠杆菌和农杆菌属中复制。Rothstein等人(1987)描述了其构建。构建了引入潮霉素B磷酸转移酶的基因(Gritz等人,1983)的pCIB10的各种衍生物。这些衍生物能够选择仅针对潮霉素(pCIB743)或针对潮霉素和卡那霉素(pCIB715、pCIB717)的转基因植物细胞。
2.适合于非农杆菌属转化的载体
不使用根癌土壤杆菌的转化避开了在选择的转化载体中对T-DNA序列的需求,结果除了诸如上述含有T-DNA序列的载体之外,还可以使用缺乏这些序列的载体。不依赖于农杆菌属的转化技术包括通过粒子轰击、原生质体吸收(例如PEG和电穿孔)和微注射的转化。对载体的选择主要取决于对所转化物种的优先选择。下面描述了适合于非农杆菌属转化的典型载体的构建。
a.pCIB3064:
pCIB3064是一种适合于与通过除草剂basta(或膦丝菌素)选择联用的定向基因转移技术的pUC-衍生的载体。质粒pCIB246包括可操作地与大肠杆菌GUS基因融合的CaMV 35S启动子和CaMV 35S转录终止子,在PCT公开申请WO 93/07278中有描述。这种载体的35S启动子含有两个起始位点的ATG序列5’。使用标准PCR技术使这些位点发生突变,以这样一种方式以便除去ATGs并生成限制位点Sspl和Pvull。新的限制位点是离单一Sall位点96和37bp和离实际起始位点101和42bp。将所得的pCIB246衍生物命名为pCIB3025。然后通过用Sall和Sacl消化从pCIB3025上切下GUS基因,使末端钝端化并重新连接而生成质粒pCIB3060。质粒pJIT82获自John Innes Centre,Norwich,切下含有来自产绿色链霉菌的bar基因的400bp Saml片段并将其插入pCIB3060的Hpal位点(Thompson等人,1987)。该过程生成pCIB3064,它包括在除草剂筛选用的CaMV 35S启动子和终止子控制下的bar基因、氨苄青霉素抗性基因(用于大肠杆菌中的筛选)和带有单一位点Sphl、Pstl、Hindlll和BamHl的多接头。这种载体适合于克隆含有其自身调节信号的植物表达弹夹。
b.pSOG19和pSOG35:
pSOG35是一种使用大肠杆菌基因二氢叶酸还原酶(DFR)作为产生氨甲蝶呤抗性的选择标记的转化载体。将PCR用于扩增35S启动子(-800bp)、来自玉米Adh1基因的内含子6(-550bp)和18bp来自pSOG10的GUS未翻译前导序列。另外通过PCR扩增编码大肠杆菌二氢叶酸还原酶II型基因的250-bp片段,并给这两种PCR片段装配来自包括pUC19载体主链和胭脂氨酸合酶终止子的pB1221(Clontech)的Sacl-Pstl片段。装配这些片段产生含有与内含子6序列融合的35S启动子、GUS前导区、DHFR基因和胭脂氨酸合酶终止子的pSOG19。来自玉米褪绿斑驳病毒(MCMV)的前导序列对pSOG19中的GUS前导区取代的产生了pSOG35载体。pSOG19和pSOG35携带用于氨苄青霉素抗性的pUC基因并具有用于克隆外来物质的Hindlll、Sphl、Pstl和EcoRl位点。
实施例12:转化
一旦将有意义的基因序列克隆入表达系统,则将它转化入植物细胞。用于植物转化和再生的方法在本领域中是众所周知的。例如,已经将Ti质粒载体用于转运外源DNA以及直接DNA吸收、脂质体、电穿孔、微注射和微粒。此外,可以将来自农杆菌属的细菌用于转化植物细胞。下面是用于转化双子叶植物和单子叶植物的有代表性技术的描述。
1.双子叶植物的转化
用于双子叶植物的转化技术在本领域中是众所周知的且包括基于农杆菌属的技术和不需要农杆菌属的技术。非农杆菌属技术包括由原生质体或细胞直接摄入外源性遗传物质。该过程可以通过PEG或电穿孔介导的吸收、粒子轰击介导的转运或微注射来完成。这些技术的实例由Paszkowski等人,1984;Potrykus等人,1985;Reich等人,1986和Klein等人,1987描述。在每种情况中,使用本领域中公知的标准技术使转化的细胞再生成完整植物。
农杆菌属介导的转化是一种用于转化双子叶植物的优选技术,这是由于其高转化效率及其在许多不同物种的广泛应用所导致的。农杆菌属转化一般包括将携带有意义的外源DNA的二元载体(例如pCIB200或pCIB2001)转入合适的农杆菌属菌株的步骤,这种合适的农杆菌属菌株可以依赖于共存Ti质粒上由宿主农杆菌属菌株携带的vir基因补体或染色体的存在方式(例如pCIB200和pCIB2001所用的菌株CIB542)(Uknes等人,1993)。将重组二元载体转入农杆菌属的过程通过使用携带重组二元载体的大肠杆菌进行的三亲交配步骤来完成,所述的大肠杆菌是一种携带诸如pRK2013且能够使重组二元载体达到靶农杆菌属菌株的质粒的辅助大肠杆菌菌株。另一方面,可以通过DNA转化将重组二元载体转入农杆菌属(H_fgen&Willmitzer,1988)。
通过重组农杆菌属转化靶植物物种通常包括农杆菌属与来自植物的外植体的共培养,并按照本领域中众所周知的方案进行。在携带二元质粒T-DNA边缘之间存在的抗生素或除草剂抗性标记的选择培养基上使转化的组织再生。
另一种使用基因转化植物细胞的手段包括使惰性或生物活性颗粒推进到植物组织和细胞上。这项技术公开在美国专利号4,945,050、5,036,006和5,100,792中。一般来说,该过程包括在有效透过细胞外表面并在其内部产生结合的条件下将惰性或生物活性颗粒推进到细胞中。当使用惰性颗粒时,可以通过给颗粒包裹含有所需基因的载体而将所述载体引入细胞。另一方面,靶细胞可被载体包围,使得该载体尾随颗粒进入细胞。也可以将生物活性颗粒(例如各自含有试图引入的DNA的干燥的酵母细胞、干燥的细菌或噬菌体)推进到植物细胞组织中。
2.单子叶植物的转化
目前大多数单子叶植物物种的转化已经成为常规的。优选的技术包括使用PEG或电穿孔技术将基因直接转入原生质体和粒子轰击入愈伤组织。可以使用单DNA物种或多DNA物种进行转化(即共转化)且这两项技术均适用于本发明。共转化可以具有避免完全载体构建和生成带有感兴趣基因的未连锁的基因座和选择性标记的转基因植物的优点,从而能够在随后的生产过程中除去所述的选择性标记,应将这一过程看作是理想的。然而,使用共转化的缺点在于整合入基因组的单一DNA种类的频率低于100%(Schocher等人,1986)。
专利申请EP 0292 435、EP 0392 225和WO 93/07278中描述了用于来自玉米原种近交系的愈伤组织和原生质体的制备、使用PEG或电穿孔转化原生质体和由转化的原生质体再生玉米植物的技术。Grodon-Kamm等人(1990)和Fromm等人(1990)已经公开了使用粒子轰击转化A188-衍生的玉米品系的技术。此外,WO 93/07278和Koziel等人(1993)中描述了通过粒子轰击转化玉米原种近交系的技术。这项技术使用传粉后14-15天从玉米穗上切下的1.5-2.5mm长的未成熟胚和轰击用PDS-1000He Biolstics装置。
还可以通过使用原生质体或粒子轰击的定向基因转移技术来进行水稻的转化。已经描述了针对日本型和印度型的原生质体介导的转化(Zhang等人,1988;Shimamoto等人,1989;Datta等人,1990)。两种类型也通常是使用粒子轰击转化的(Christou等,1991)。此外,WO 93/21335中描述了通过电穿孔转化水稻的技术。
专利申请EP 0 332 581中描述了Pooideae原生质体的生成、转化和再生。这些技术用于转化鸭茅属和小麦。此外,Vasil等人(1992)已经描述了使用粒子轰击入C型长期再生性愈伤组织的细胞进行小麦的转化,Vasil等人(1993)和Weeks等人(1993)也描述了使用粒子轰击未成熟胚和成熟胚衍生的愈伤组织来进行小麦的转化。然而,优选的小麦转化技术涉及通过粒子轰击未成熟胚转化小麦并包括在基因递送之前的高蔗糖或高麦芽糖步骤。在轰击前,将任何数量的胚(0.75-1mm长)平板固定在含有3%蔗糖(Murashiga&Skoog,1962)和3mg/l 2,4-D的MS培养基上以便诱导体细胞胚,使该步骤在黑暗中进行。在选择进行轰击的当天,从诱导培养基中取出胚并置于渗压剂(即含有一般以15%的所需浓度添加的蔗糖或麦芽糖的诱导培养基)上。使胚进行质壁分离2-3小时,然后进行轰击。虽然并不关键,但通常是每个靶平板上有20个胚。应用标准程序使一种合适的携带基因的质粒(诸如pCIB3064或pSG35)沉淀在微米大小的金粒子上。用使用~1000psi的破裂压力并使用标准80目筛的DuPont Biolistics_氦装置向各平板的胚发射。在轰击后,将胚放回黑暗处以便恢复约24小时(仍然在渗压剂上)。24小时后,从渗压剂上取出胚并放回诱导培养基上,在此它们在再生前稳定约1个月。约1个月后,将带有发育中的胚发生愈伤组织的胚外植体转入再生培养基(MS+1mg/升NAA、5mg/升GA),该培养基进一步含有合适的选择剂(就pCIB3064而言是10mg/l basta,就pSOG35而言是2mg/l氨甲蝶呤)。约1个月后,将发育的枝条转入称作“GA7s”的含有半数浓度的MS、2%蔗糖和相同浓度的选择剂的较大无菌容器中。
还描述了使用农杆菌属转化单子叶植物的技术方案。参见,WO94/00977和美国专利号5,591,616。
III.培育和种子生产
实施例13:培育
通过用本发明基因转化获得的植物可以是任何广泛的植物物种,包括那些单子叶植物和双子叶植物;然而,本发明方法中所用的植物优选自上文所述的农学上重要的靶农作物。可以通过培育将本发明基因的表达与对生产和质量来说重要的其它特征一起引入植物品系。培育手段和技术在本领域中是公知的。例如,参见Welsh J.R.(1981);Wood D.R.(编辑)(1983);Mayo O.(1987);Singh,D.P.(1986);和Wricke和Weber(1986)。
通过有性繁殖或营养生长传递上述基因工程的转基因种子和植物的遗传特性,由此可以在子代植物中维持并传播这些特性。通常所述的维持和传播充分利用了为适合特殊目的发展的农业方法,诸如耕种、播种或收获。还可以应用诸如水培或温室技术这样的特殊方法。当生长的农作物易受到由昆虫或感染以及杂草植物竞争所导致的损害和破坏时,实施某些措施以便控制杂草、植物病害、昆虫、线虫和其它不良条件以改善产量。它们包括耕作土壤或除去杂草和感染植物这样的机械措施以及施用诸如除草剂、杀真菌剂、杀配子剂、杀线虫剂、生长调节剂、催熟剂和杀虫剂这样的农用化学品。
在植物培育中可以进一步应用本发明的有利遗传特性的转基因植物和种子,其目的在于开发具有改善特性的植物,所述的改善特性诸如害虫、除草剂或应激耐受性、改善的营养价、产量提高或使倒伏或脱落损失减小的改进结构。不同培育步骤的特征在于充分确定的人的干预诸如选择杂交品系、亲本品系的定向传粉或选择合适的子代植物。根据所需特性的不同,制定不同的培育措施。相关技术在本领域中是众所周知的,而且包括但不限于杂交、近交、回交培育、多线培育、变种混合、种间杂交、非整倍体技术等。杂交技术还包括植物不育化以便通过机械、化学或生化方式产生雄性或雌性不育植物。带有不同品系花粉的雄性不育植物的交叉传粉确保了雄性不育的基因组,而雌性不育植物均匀地获得了两亲本品系的特性。因此,可以将本发明的转基因种子和植物用于改善的植物品系的培育,例如,这种培育方法提高了诸如除草剂或杀虫剂(pestidice)处理这样的常规方法的有效性,或因其改进的遗传特性而不再需要所述的方法。另一方面,可以获得具有改善的应激特性的新型农作物,由于它们具有最佳化遗传“设备”,从而使收获产品的质量优于不能耐受类似的不良发育条件的产品。
实施例14:种子的生产
在种子的生产过程中,发芽的质量和种子的均匀度是主要的产品特征,而由农场主收获和销售的发芽的质量和种子的均匀度并不重要。当难以保持农作物不含其它农作物和杂草种子来控制种子生长病害和生产发芽良好的种子时,在生长、调节和销售纯种子领域中富有经验的种子生产者已经进行了相当广泛而明确的种子生产实践。因此,对农场主而言通常的做法是购买满足特定质量标准的保质种子而不是使用从其拥有的农作物中收获的种子。通常将用作种子的繁殖物质用包括除草剂、杀昆虫剂、杀真菌剂、杀细菌剂、杀线虫剂、杀软体动物剂或其混合物的保护剂包裹材料进行处理。通常使用的保护剂包裹材料包括诸如克菌丹、萎锈灵、福美双(TMTD_)、methalaxyl(Apron_)、虫螨磷(Actellic_)这样的化合物。如果需要,可以将这些化合物与通常在制剂领域使用的其它载体、表面活性剂或促进施用的佐剂一起配制以便产生抵抗由细菌、真菌或动物昆虫导致的破坏的保护作用。可以通过用液体制剂浸渗繁殖物质或通过用组合的湿或干制剂包裹来施用所述的保护剂包裹材料。诸如定向于芽或果实的处理这样的其它施用方法也是可行的。
本发明的再一个方面是提供诸如上述典型方法这样的新型农用方法,其特征在于使用本发明的转基因植物、转基因植物物质或转基因种子。
可以将种子装入由合适的包装材料构成的包、容器或管中,所述的包、容器或管能够密封以包含种子。可以将包、容器或管设计成种子的短期或长期包装或两者均有的包装。合适的包装材料的实例包括诸如牛皮纸这样的纸、硬质塑料或柔顺塑料或其它聚合材料、玻璃或金属。理想的情况是所述的包、容器或管由多层相同或不同类型的包装材料构成。在一个优选的实施方案中,提供所述的包、容器或管以便防止或限制水和湿气接触种子。在一个实例中,将所述的包、容器或管密封、例如加热密封以便防止水或湿气进入。在另一个实施方案中,将吸水物质置于包装材料层之间或接近包装材料层。在另一个实施方案中,将包、容器或管或构成的包装材料进行处理以便限制、抑制或防止种子储存或运输过程中的病害、污染或其它不利影响。这类处理方法的一个实例是例如通过化学方式或通过接触射线进行不育化。本发明包括的是一种商品包,它装有包含本发明基因的转基因植物的种子、合适的载体及其用于使植物产生广谱病害抗性的标记说明书,其中所述转基因植物种子中包含本发明基因,其在所述转化植物中的表达水平高于在野生型植物中的表达水平。
IV.病害抗性评价
用本领域中公知的方法抗性病害抗性评价。参见:Uknes等人(1993);G_rlach等人(1996);Alexander等人(1993)。例如,几种有代表性的病害抗性测定如下所述。
实施例15:寄生疫霉(黑胫病)抗性测定
对如Alexander等人(1993)中所述生长的6周龄植物进行黑胫病致病生物体寄生疫霉抗性测定。给植物浇水、排干水且然后通过给土壤施用10ml孢子囊混悬液(300孢子囊/ml)进行接种。将接种的植物保存在维持在日温度23-25℃和夜温度20-22℃的温室内。本试验用枯萎指数如下:0=无症状;1=无症状;1=有一定枯萎征兆、膨胀度下降;2=明显的枯萎症状、但无腐烂或矮化;3=明显的枯萎症状、伴有矮化,但无明显的茎腐烂;4=严重枯萎、肉眼可见茎腐烂和一定的对根系的损害;5=如4所述,但植物接近死亡或已经死亡并伴有根系的严重减少。在对以随机设计方式排列的植物全盲的情况下取得全部测定得分。实施例16:丁香假单胞菌抗性测定
给几种6-7周龄植物的两个下部的叶子注入浓度为106或3×106/ml水溶液的丁香假单胞菌变种烟草野火病假单胞菌(Pseuodomonas syringae pv.tabaci)菌株#551。在各个时间点评价6株单独的植物。给烟草野火病假单胞菌感染的植物定为5点病害严重等级:5=100%死亡的组织;0=无症状。对每天的评价结果进行T-检验(LSD)并在平均病害等级值后显示分型。在评价当天后面跟着相同字母的数值在统计学上没有显著差异。
实施例17:烟草尾孢抗性测定
将烟草尾孢(ATCC#18366)的孢子混悬液(100,000-150,000个孢子/ml)喷洒在叶表面的紧急溢放口上。将植物维持在100%湿度下5天。此后给植物喷水雾、每天5-10次。在各时间点处评价6株单独的植物。给烟草尾孢定级为显示病害主要症状的叶面积百分比a%。对每天的评价结果进行T-检验(LSD)并在平均病害等级值后显示分型。在评价当天后面跟着相同字母的数值在统计学上没有显著差异。
实施例18:寄生霜霉抗性测定
对如Uknes等人(1993)中所述的植物进行寄生霜霉抗性测定。通过喷洒分生孢子混悬液(大约5×104个孢子/毫升)给植物接种寄生霜霉的相容分离物。在17℃下的生长室内的湿度下通过14小时-日/10小时-夜循环培养接种植物。在有分生孢子梗存在的情况下接种后的3-14天、优选7-12天时检查植物。此外,随机选择经各种处理的几种植物并用乳酚-锥虫蓝染色(Keogh等人,1980)以便于镜检。
上述公开的实施方案是解释性的。对本领域技术人员来说本发明的该公开内容仅是本发明许多变化形式的一部分。所有这类显而易见和可预测的变化形式均包括在权利要求中。
序列表中序列的简要描述SEQ ID NO:1-来自烟草的NIM1同源物的全长cDNA序列。SEQ ID NO:2-由SEQ ID NO:1编码的烟草NIM1同源物的蛋白质序列。SEQ ID NO:3-来自番茄的NIM1同源物的全长cDNA序列。SEQ ID NO:4-由SEQ ID NO:3编码的番茄NIM1同源物的蛋白质序列。SEQ ID NO:5-来自欧洲油菜的NIM1同源物的部分cDNA序列。SEQ ID NO:6-由SEQ ID NO:5编码的欧洲油菜NIM1同源物的部分蛋白质序列。SEQ ID NO:7-来自鼠耳芥的NIM1同源物(AtNMLc5)的全长cDNA序列。SEQ ID NO:8-由SEQ ID NO:7编码的鼠耳芥NIM1同源物AtNMLc5的全长蛋白质序列。SEQID NO:9-14-实施例1-4中使用的寡核苷酸引物。SEQ ID NO:15-来自鼠耳芥的NIM1同源物(AtNMLc2)的基因组DNA序列。SEQ ID NO:16-由SEQ ID NO:15编码的鼠耳芥NIM1同源物AtNMLc2的蛋白质序列。SEQ ID NO:17-来自鼠耳芥的NIM1同源物(AtNMLc4-1)的基因组DNA序列。SEQ ID NO:18-由SEQ ID NO:17编码的鼠耳芥NIM1同源物AtNMLc4-1的蛋白质序列。SEQID NO:19-来自鼠耳芥的NIM1同源物(AtNMLc4-2)的基因组DNA序列。SEQ ID NO:20-由SEQ ID NO:19编码的鼠耳芥NIM1同源物AtNMLc4-2的蛋白质序列。SEQ ID NO:21-PCR引物NIM 1A。SEQ ID NO:22-PCR引物NIM 1B。SEQ ID NO:23-PCR引物NIM 1C。SEQ ID NO:24-PCR引物NIM 1D。SEQ ID NO:25-PCR引物NIM 2A。SEQ ID NO:26-PCR引物NIM 2B。SEQ ID NO:27-PCR引物NIM 2C。SEQ ID NO:28-PCR引物NIM 2D。SEQ ID NO:29-由烟草(烟草A)扩增的659bp NIM-样DNA片段,它是36个序列的共有区且与鼠耳芥NIM1基因序列具有67%的序列同一性。SEQ ID NO:30-由SEQ ID NO:29编码的蛋白质序列。SEQ ID NO:31-由烟草(烟草B)扩增的498bp NIM-样DNA片段,它是2个序列的共有区且与鼠耳芥NIM1基因序列具有62%的序列同一性。SEQ ID NO:32-由SEQ ID NO:31编码的蛋白质序列。SEQ ID NO:33-由烟草(烟草C)扩增的498bp NIN-样DNA片段,它是3个序列的共有区且与鼠耳芥NIM1基因序列具有63%的序列同一性。SEQ ID NO:34-由SEQ ID NO:33编码的蛋白质序列。SEQ ID NO:35-由烟草(烟草D)扩增的399bp NIM-样DNA片段,它与鼠耳芥NIM1基因序列具有59%的序列同一性。SEQ ID NO:36-由SEQ ID NO:35编码的蛋白质序列。SEQ ID NO:37-由番茄(番茄A)扩增的498bp NIM-样DNA片段,它是8个序列的共有区且与鼠耳芥NIM1基因序列具有63%的序列同一性。SEQ ID NO:38-由SEQ ID NO:37编码的蛋白质序列。SEQ ID NO:39-由甜菜(Beta vulgaris)扩增的498bp NIM-样DNA片段,它是24个序列的共有区且与鼠耳芥NIM1基因序列具有66%的序列同一性。SEQ ID NO:40-由SEQ ID NO:39编码的蛋白质序列。SEQ ID NO:41-由向日葵(向日葵A)扩增的498bp NIM-样DNA片段,它是9个序列的共有区且与鼠耳芥NIM1基因序列具有61%的序列同一性。SEQ ID NO:42-由SEQ ID NO:41编码的蛋白质序列。SEQ ID NO:43-由向日葵(向日葵B)扩增的498bp NIM-样DNA片段,它是10个序列的共有区且与鼠耳芥NIM1基因序列具有59%的序列同一性。SEQ ID NO:44-由SEQ ID NO:43编码的蛋白质序列。SEQ ID NO:45-由马铃薯(马铃薯A)扩增的653bp NIM-样DNA片段,它是15个序列的共有区且与鼠耳芥NIM1基因序列具有68%的序列同一性。SEQ ID NO:46-由SEQ ID NO:45编码的蛋白质序列。SEQ ID NO:47-由马铃薯(马铃薯B)扩增的498bp NIM-样DNA片段,它是3个序列的共有区且与鼠耳芥NIM1基因序列具有61%的序列同一性。SEQ ID NO:48-由SEQ ID NO:47编码的蛋白质序列。SEQ ID NO:49-由马铃薯(马铃薯C)扩增的477bp NIM-样DNA片段,它是2个序列的共有区且与鼠耳芥NIM1基因序列具有62%的序列同一性。SEQ ID NO:50-由SEQ ID NO:49编码的蛋白质序列。SEQ ID NO:51-由欧洲油菜(Canola A)扩增的501bp NIM-样DNA片段,它是5个序列的共有区且与鼠耳芥NIM1基因序列具有59%的序列同一性。SEQ ID NO:52-由SEQ ID NO:51编码的蛋白质序列。SEQ ID NO:53-由欧洲油菜(Canola B)扩增的501bp NIM-样DNA片段,它是5个序列的共有区且与鼠耳芥NIM1基因序列具有58%的序列同一性。SEQ ID NO:54-由SEQ ID NO:53编码的蛋白质序列。SEQ ID NO:55-由欧洲油菜(Canola C)扩增的498bp NIM-样DNA片段,它与鼠耳芥NIM1基因序列具有56%的序列同一性。SEQ ID NO:56-由SEQ ID NO:55编码的蛋白质序列。SEQ ID NO:57-由欧洲油菜(Canola D)扩增的498bp NIM-样DNA片段,它与鼠耳芥NIM1基因序列具有73%的序列同一性。SEQ ID NO:58-由SEQ ID NO:57编码的蛋白质序列。SEQ ID NO:59-PCR引物NIM 3A。SEQ ID NO:60-PCR引物NIM 3B。SEQ ID NO:61-由番茄(番茄B)扩增的148bp NIM-样DNA片段,它是3个序列的共有区且与鼠耳芥NIM1基因序列具有72%的序列同一性。SEQ ID NO:62-由SEQ ID NO:61编码的蛋白质序列。SEQ ID NO:63-来自甜菜(Beta vulgaris)的NIM1同源物的全长cDNA序列,它与SEQ ID NO:39的PCR片段一致。SEQ ID NO:64-由SEQ ID NO:62编码的甜菜NIM1同源物的蛋白质序列。SEQ ID NO:65-来自向日葵(向日葵B)的NIM1同源物的全长cDNA序列,它与SEQ ID NO:43的PCR片段一致。SEQ ID NO:66-由SEQ ID NO:65编码的向日葵NIM1同源物的蛋白质序列。SEQ ID NO:67-与鼠耳芥NIM-样基因组序列AtNMLc2(SEQ ID NO:15)一致的cDNA序列。SEQ ID NO:68-由SEQ ID NO:67编码的蛋白质序列。SEQ ID NO:69-与鼠耳芥NIM-样基因组序列AtNMLc4-1(SEQ ID NO:17)一致的cDNA序列。SEQ ID NO:70-由SEQ ID NO:69编码的蛋白质序列。SEQ ID NO:71-与鼠耳芥NIM-样基因组序列AtNMLc4-2(SEQ ID NO:19)一致的cDNA序列。SEQ ID NO:72-由SEQ ID NO:71编码的蛋白质序列。SEQ ID NO:73-来自烟草(烟草B)的NIM1同源物的全长cDNA序列,它与SEQ ID NO:31的PCR片段一致。SEQ ID NO:74-由SEQ ID NO:73编码的烟草NIM1同源物的蛋白质序列。寄存
根据有关国际公认的用于专利程序目的的微生物寄存布达佩斯条约,已经将下列材料寄存在Agricultural Research Service,PatentCulture Collection(NRRL),1815 North University Street,Peoria,lllinois 61604,USA。对所寄存材料的可得到性的所有限制在专利授权后最终将被取消。
克隆 保藏号 寄存日
pNOV1203 NRRL B-30049 1998年8月17日pNOV1204 NRRL B-30050 1998年8月17日pNOV1206 NRRL B-30051 1998年8月17日AtNMLc5 NRRL B-30139 1999年5月25日参考文献
本文引入的参考文献表示本领域中目前的状况。将下列各文献引入本公开说明书中作为参考。
美国专利号4,940,935
美国专利号4,945,050
美国专利号5,036,006
美国专利号5,100,792
美国专利号5,188,642
美国专利号5,523,311
美国专利号5,591,616
美国专利号5,614,395
美国专利号5,639,949
美国专利号5,792,904
EP 0 292 435
EP 0 332 104
EP 0 332 581
EP 0 342 926
EP 0 392 225
EP 0 452 269
国际PCT申请WO 93/07278
国际PCT申请WO 93/21335
国际PCT申请WO 94/00977
国际PCT申请WO 94/13822
国际PCT申请WO 94/16077
国际PCT申请WO 97/49822
国际PCT申请WO 98/06748
国际PCT申请WO 98/26082
国际PCT申请WO 98/29537
Alexander等《美国国家科学院院报》(Proc.Natl.Acad.Sci.USA)90:7327-7331(1993)
Aoyama和Chua《植物杂志》(The Plant Journal)11:605-612(1997)
Ausubel,F.M.等《分子生物学最新方案》(Current Protocols inMolecular Biology),Greene Publishing Assoc.和Wiley-Interscience(1987)出版
Bartlett等、Edelmann等(编辑)《叶绿体分子生物学方法》(Methods in Chloroplast Molecular Biology),Elsevierpp:1081-1091(1982)
Bevan等《自然》(Nature)304:184-187(1983)
Bevan等《核酸研究》(Nucl.Acids Res.)(1984)
Bi等《植物杂志》(Plant J.)8:235-245(1995)
Binet等《植物科学》(Plant Science)79:87-94(1991)
Blochinger&Diggelmann《细胞分子生物学》(Mol.Cell.Biol.)4:2929-2931
Bourouis等《欧洲分子生物学杂志》(EMBO J.)2(7):1099-1104(1983)
Boutry等《自然》(Nature)328:340-342(1987)
Caddick等《天然生物技术》(Nat.Biotechnol.)16:177-180(1998)
Callis等《基因开发》(Genes Develop.)1:1183-1200(1987)
Cameron等《植物杂志》(Plant J.)5:715-725(1994)
Cao等《植物细胞》(Plant Cell)6:1583-1592(1994)
Cao等《细胞》(Cell)88:57-63(1997)
Casas等《美国国家科学院院报》(Proc.Natl.Acad.Sci.USA)90:11212-11216(1993)
Century等《美国国家科学院院报》(Proc.Natl.Acad.Sci.USA)92:6597-6601(1995)
Chibbar等《植物细胞繁殖》(Plant Cell Rep.)12:506-509(1993)
Chrispeels《植物生理学和植物分子生物学年鉴》(Ann.Rev.Plant Physiol.Plant Mol.Biol.)42:21-53(1991)
Christou等《植物生理学》(Plant Physiol.)87:671-674(1988)
Christou等《生物技术》(Biotechnology)9:957-962(1991)
Christensen等《植物分子生物学》(Plant Molec.Biol.)12:619-632(1989)
Comai等《生物化学杂志》《J.Biol.Chem.》263:15104-15109(1988)
Crossway等《生物技术》(BioTechniques)4:320-334(1986)
Datta等《生物技术》(Biotechnology)8:736-740(1990)
de Framond FEBS 290:103-106(1991)
Delaney等《科学》(Science)266:1247-1250(1994)
Delaney等《美国国家科学院院报》(Proc.Natl.Acad.Sci.USA)92:6602-6606(1995)
Dempsey和Klessig,Bulletin de L’Institut Pasteur 93:167-186(1995)
Dietrich等《细胞》(Cell)77:565-577(1994)
Friek等《植物分子生物学》(Pant Molec.Biol.)22:129-142(1993)
Fromm等《生物技术》(Biotechnology)8:833-839(1990)
Gaffney等《科学》(Science)261:754-756(1993)
Gallie等《核酸研究》(Nucl.Acids Res.)15:8693-8711(1987)
Glazebrook等《遗传学》(Genetics)143:973-982(1996)
Gordon-Kamm等《植物细胞》(Plant Cell)2:603-618(1990)
Gordon-Kamm等《转基因植物》(Transgenic Plants)第2卷pp.21-33,Academic Press(1993)出版
G_rlach等《植物细胞》(Plant Cell)8:629-643(1996)
Greenberg等《细胞》(Cell)77:551-563(1994)
Gritz等《基因》(Gene)25:179-188(1983)
Hayashimoto等《植物生理学》(Plant Physiol.)93:857-863(1990)
Hill等Euphytica 85:119-123(1995)
Hinchee等《生物技术》(Biotechnology)6:915-921(1988)
H_fgen&Willmitzer《核酸研究》(Nucl.Acids Res.)16:9877(1988)
Hudspeth&Grula《植物分子生物学》(Pant Molec.Biol.)12:579-589(1989)
Hunt和Ryals《植物科学评述》(Crit.Rev.in Plant Sci.)15:583-606(1996)
Innis等《PCR方案-方法和应用指南》(PCR Protocols,a Guideto Methods and Applications)编辑,Academic Press(1990)
Ishida等《天然生物技术》(Nature Biotechnology)14:745-750(1996)
Jahne等《应用遗传学原理》(Theor.Appl.Genet.)89:525-533(1994)
Keegan等《科学》(Science)231:699-704(1986)
Keogh等《英国真菌学协会会报》(Trans.Br.Mycol.Soc.)74:329-333(1980)
Klein等《自然》(Nature)327:70-73(1987)
Klein等《美国国家科学院院报》(Proc.Natl.Acad.Sci.USA)85:4305-4309(1988)
Klein等《生物/技术》(Bio/Technology)6:559-563(1988)
Klein等《植物生理学》(Plant Physiol.)91:440-444(1988)
Koziel等《生物技术》(Biotechnology)11:194-200(1993)
Koziel等《纽约科学院年鉴》(Annals of the New York Academyof Science)792:164-171(1996)
Lawton等“系统获得性抗性的分子生物学”(The molecularbiology of systemic aquired resistance)-《植物中的防御反应机理》(Mechanisms of Defence Responses in Plants)B.Fritig和M.Legrand编辑(Dordrecht,The Netherlands:Kluwer AcademicPublishers)pp.422-432(1993)
Lawton等《植物杂志》(Plant J.)10:71-82(1996)
Logemann等《植物细胞》(Plant Cell)1:151-158(1989)
Maher等《美国国家科学院院报》(Proc.Natl.Acad.Sci.USA)91:7802-7806(1994)
Mauch-Mani和Slusarenko《分子植物-微生物相互作用》(Mol.Plant-Microbe Interact.)7:378-383(1994)
Mauch-Mani和Slusarenko《植物细胞》(Plant Cell)8:203-212(1996)
Mayo O.《植物育种原理》(The Theory of Plant Breeding)第2版,Clarendon Press,Oxford(1987)
Mazur等《植物生理学》(Plant Physiol.)85:1110(1987)
McBride等《植物分子生物学》(Plant Molecular Biology)14:266-276(1990)
McCabe等《生物技术》(Biotechnology)6:923-926(1988)
McElroy等《植物细胞》(Plant Cell)2:163-171(1990)
McElroy等《分子基因遗传学》(Mol.Gen.Genet.)231:150-160(1991)
Messing&Vierra《基因》(Gene)19:259-268(1982)
Murashiga&Skoog《植物生理学》(Physiollogia Plantarum)15:473-497(1962)
Nehra等《植物杂志》(The Plant Journal)5:285-297(1994)
Neuhaus等《美国国家科学院院报》(Proc.Natl.Acad.Sci.USA)88:10362-10366(1991)
Norris等《植物分子生物学》(Plant Mol.Biol.)21:895-906(1993)
Pallas等《植物杂志》(Plant J.)10:281-293(1996)
Parker等《植物细胞》(Plant Cell)8:2033-2046(1996)
Paszkowski等《欧洲分子生物学杂志》(EMBO J.)3:2717-2722(1984)
Payne等《植物分子生物学》(Plant Mol.Biol.)11:89-94(1988)
Picard等《细胞》(Cell)54:1073-1080(1988)
Potrykus等《分子基因遗传学》(Mol.Gen.Genet.)199:169-177(1985)
Reich等《生物技术》(Biotechnology)4:1001-1004(1986)
Riggs等《美国国家科学院院报》(Proc.Natl.Acad.Sci.USA)83:5602-5606(1986)
Rogers等《美国国家科学院院报》(Proc.Natl.Acad.Sci.USA)82:6512-6516(1985)
Rohrmeier&Lehle《植物分子生物学》(Plant Molec.Biol.)22:783-792(1993)
Rothstein等《基因》(Gene)53:153-161(1987)
Ryals等《植物细胞》(Plant Cell)8:1809-1819(1996)
Ryals等《植物细胞》(Plant Cell)9:425-439(1997)
Sambrook等《分子克隆》(Molecular Cloning),编辑,Cold SpringHarbor Laboratory Press(1987)
Sanford等《微粒科学与技术》(Particulate Science andTechnology)5:27-37(1987)
Schmidhauser&Helinski《细菌学杂志》(J.Bacteriol.)164:446-455(1985)
Schocher等《生物技术》(Biotechnology)4:1093-1096(1986)
Shimamoto等《自然》(Nature)338:274-277(1989)
Shinshi等《植物分子生物学》(Plant Molec.Biol.)14:357-368(1990)
Shulaev等《植物细胞》(Plant Cell)7:1691-1701(1995)
Silhavy等《使用基因融合体的实验》(Experiments with GeneFusions),编辑,Cold Spring Harbor Laboratory Press(1984)
Singh,D.P.《病害与昆虫害虫抗性的培育》(Breeding forResistance to Diseases and Insect Pests),Springer-Verlag,NY(1986)
Skuzeski等《植物分子生物学》(Plant Molec.Biol.)15:65-79(1990)
Somers等《生物/技术》(Bio/Technology)10:1589-1594(1992)
Spencer等《应用遗传学原理》(Theor.Appl.Genet.)79:625-631(1990)
Stanford等《分子基因遗传学》(Mol.Gen.Genet.)215:200-208(1989)
Svab等《美国国家科学院院报》(Proc.Natl.Acad.Sci.USA)87:8526-8530(1990)
Taylor等《植物细胞繁殖》(Plant Cell Rep.)12:491-495(1993)
Thompson等《欧洲分子生物学杂志》(EMBO J.)6:2519-2523(1987)
Torbert等《基因开发》(Genes Develop.)2:718-729(1988)
Triezenberg等《植物细胞》(Plant Cell)4:645-656(1992)
Uknes等《植物细胞》(Plant Cell)5:159-169(1993)
Uknes等《分子植物微生物相互作用》(Molecular Plant MicrobeInteractions)6:680-685(1993)
Uknes等《分子植物-微生物相互作用》(Mol.Plant-MicrobeInteract.)6:692-698(1993)
Umbeck等《生物/技术》(Bio/Technology)5:263-266(1987)
Unger等《植物分子生物学》(Plant Molec.Biol.)13:411-418(1989)
van den Broeck等《自然》(Nature)313:358-363(1985)
Vasil等《生物技术》(Biotechnology)10:667-674(1992)
Vasil等《生物技术》(Biotechnology)11:1553-1558(1993)
Vernooij等《植物细胞》(Plant Cell)6:959-965(1994)
Vernooij等《分子植物-微生物相互作用》(Mol.Plant-MicrobeInteract.)8:228-234(1995)
Von Heijne等《植物分子生物学报告》(Plant Mol.Biol.Rep.)9:104-126(1991)
Vorst等《基因》(Gene)65:59(1988)
Wan等《植物生理学》(Plant Physiol.)104:37-48(1994)
Ward等《植物细胞》(Plant Cell)3:1085-1094(1991)
Warner等《植物杂志》(Plant J.)3:191-201(1993)
Wasmann等《分子基因遗传学》(Mol.Gen.Genet.)205:446-453(1986)
Weeks等《植物生理学》(Plant Physiol.)102:1077-1084(1993)
Weissinger等《遗传学年鉴》(Annual Rev.Genet.)22:421-477(1988)
Welsh J.R.《植物遗传学和育种基础》(Fundamentals of PlantGnentics and Breeding),John Wiley&Sons,NY(1981)
Weymann等《植物细胞》(Plant Cell)7:2013-2022(1995)
White《核酸研究》(Nucl.Acids Res.)18:1062(1990)
Wood D.R.(编辑)《农作物育种》(Crop Breeding),AmericanSociety of Agronomy Madison,Wisconsin(1983)
Wricek和Weber《定量遗传学和选择植物育种》(QuantitativeGenetics and Selection Plant Breeding),Walter de Gruyter和Co.Berlin(1986)
Xu等《植物分子生物学》(Plant Molec.Biol.)22:573-588(1993)
Zhang等《植物细胞繁殖》(Plant Cell Rep.)7:379-384(1998)
序列表<110>Novartis AG<120>新型植物基因及其用途<130>S-30857A/RTP2095<140><141><160>74<170>PatentIn 2.1版<210>1<211>1767<212>DNA<213>烟草<220><221>CDS<222>(1)..(1764)<223>全长烟草cDNA序列<400>1atg gat aat agt agg act gcg ttt tct gat tcg aat gac atc agc gga 48Met Asp Asn Ser Arg Thr Ala Phe Ser Asp Ser Asn Asp Ile Ser Gly1 5 10 15agc agt agt ata tgc tgc atc ggc ggc ggc atg act gaa ttt ttc tcg 96Ser Ser Ser Ile Cys Cys Ile Gly Gly Gly Met Thr Glu Phe Phe Ser
20 25 30ccg gag act tcg ccg gcg gag atc act tca ctg aaa cgc cta tcg gaa 144Pro Glu Thr Ser Pro Ala Glu Ile Thr Ser Leu Lys Arg Leu Ser Glu
35 40 45aca ctg gaa tct atc ttc gat gcg tct ttg ccg gag ttt gac tac ttc 192Thr Leu Glu Ser Ile Phe Asp Ala Ser Leu Pro Glu Phe Asp Tyr Phe
50 55 60gcc gac gct aag ctt gtg gtt tcc ggc ccg tgt aag gaa att ccg gtg 240Ala Asp Ala Lys Leu Val Val Ser Gly Pro Cys Lys Glu Ile Pro Val65 70 75 80cac cgg tgc att ttg tcg gcg agg agt ccg ttc ttt aag aat ttg ttc 288His Arg Cys Ile Leu Ser Ala Arg Ser Pro Phe Phe Lys Asn Leu Phe
85 90 95tgc ggt aaa aag gag aag aat agt agt aag gtg gaa ttg aag gag gtg 336Cys Gly Lys Lys Glu Lys Asn Ser Ser Lys Val Glu Leu Lys Glu Val
100 105 110atg aaa gag cat gag gtg agc tat gat gct gta atg agt gta ttg gct 384Met Lys Glu His Glu Val Ser Tyr Asp Ala Val Met Ser Val Leu Ala
115 120 125tat ttg tat agt ggt aaa gtt agg cct tca cct aaa gat gtg tgt gtt 432Tyr Leu Tyr Ser Gly Lys Val Arg Pro Ser Pro Lys Asp Val Cys Val
130 135 140tgt gtg gac aat gac tgc tct cat gtg gct tgt agg cca gct gtg gca 480Cys Val Asp Asn Asp Cys Ser His Val Ala Cys Arg Pro Ala Val Ala145 150 155 160ttc ctg gtt gaggtt ttg tac aca tca ttt acc ttt cag atc tct gaa 528Phe Leu Val Glu Val Leu Tyr Thr Ser Phe Thr Phe Gln Ile Ser Glu
165 170 175ttg gtt gac aag ttt cag aga cac cta ctg gat att ctt gac aaa act 576Leu Val Asp Lys Phe Gln Arg His Leu Leu Asp Ile Leu Asp Lys Thr
180 185 190gca gca gac gat gta atg atg gtt tta tct gtt gca aac att tgt ggt 624Ala Ala Asp Asp Val Met Met Val Leu Ser Val Ala Asn Ile Cys Gly
195 200 205aaa gca tgc gag aga ttg ctt tca agc tgc att gag att att gtc aag 672Lys Ala Cys Glu Arg Leu Leu Ser Ser Cys Ile Glu Ile Ile Val Lys
210 215 220tct aat gtt gat atc ata acc ctt gat aaa gcc ttg cct cat gac att 720Ser Asn Val Asp Ile Ile Thr Leu Asp Lys Ala Leu Pro His Asp Ile225 230 235 240gta aaa caa att act gat tca cga gcg gaa ctt ggt cta caa ggg cct 768Val Lys Gln Ile Thr Asp Ser Arg Ala Glu Leu Gly Leu Gln Gly Pro
245 250 255gaa agc aac ggt ttt cct gat aaa cat gtt aag agg ata cat agg gca 816Glu Ser Asn Gly Phe Pro Asp Lys His Val Lys Arg Ile His Arg Ala
260 265 270ttg gat tct gat gat gtt gaa tta cta caa atg ttg cta aga gag ggg 864Leu Asp Ser Asp Asp Val Glu Leu Leu Gln Met Leu Leu Arg Glu Gly
275 280 285cat act acc cta gat gat gca tat gct ctc cat tat gct gta gcg tat 912His Thr Thr Leu Asp Asp Ala Tyr Ala Leu His Tyr Ala Val Ala Tyr
290 295 300tgc gat gca aag act aca gca gaa ctt cta gat ctt gca ctt gct gat 960Cys Asp Ala Lys Thr Thr Ala Glu Leu Leu Asp Leu Ala Leu Ala Asp305 310 315 320att aat cat caa aat tca agg gga tac acg gtg ctg cat gtt gca gcc 1008Ile Asn His Gln Asn Ser Arg Gly Tyr Thr Val Leu His Val Ala Ala
325 330 335atg agg aaa gag cct aaa att gta gtg tcc ctt tta acc aaa gga gct 1056Met Arg Lys Glu Pro Lys Ile Val Val Ser Leu Leu Thr Lys Gly Ala
340 345 350aga cct tct gat ctg aca tcc gat gga aga aaa gca ctt caa atc gcc 1104Arg Pro Ser Asp Leu Thr Ser Asp Gly Arg Lys Ala Leu Gln Ile Ala
355 360 365aag agg ctc act agg ctt gtg gat ttc agt aag tct ccg gag gaa gga 1152Lys Arg Leu Thr Arg Leu Val Asp Phe Ser Lys Ser Pro Glu Glu Gly
370 375 380aaa tct gct tcg aat gat cgg tta tgc att gag att ctg gag caa gca 1200Lys Ser Ala Ser Asn Asp Arg Leu Cys Ile Glu Ile Leu Glu Gln Ala385 390 395 400gaa aga aga gac cct ctg cta gga gaa gct tct gta tct ctt gct atg 1248Glu Arg Arg Asp Pro Leu Leu Gly Glu Ala Ser Val Ser Leu Ala Met
405 410 415gca ggc gat gat ttg cgt atg aag ctg tta tac ctt gaa aat aga gtt 1296Ala Gly Asp Asp Leu Arg Met Lys Leu Leu Tyr Leu Glu Asn Arg Val
420 425 430ggc ctg gct aaa ctc ctt ttt cca atg gaa gct aaa gtt gca atg gac 1344Gly Leu Ala Lys Leu Leu Phe Pro Met Glu Ala Lys Val Ala Met Asp
435 440 445att gct caa gtt gat ggc act tct gag ttc cca ctg gct agc atc ggc 1392Ile Ala Gln Val Asp Gly Thr Ser Glu Phe Pro Leu Ala Ser Ile Gly
450 455 460aaa aag atg gct aat gca cag agg aca aca gta gat ttg aac gag gct 1440Lys Lys Met Ala Asn Ala Gln Arg Thr Thr Val Asp Leu Asn Glu Ala465 470 475 480cct ttc aag ata aaa gag gag cac ttg aat cgg ctt aga gca ctc tct 1488Pro Phe Lys Ile Lys Glu Glu His Leu Asn Arg Leu Arg Ala Leu Ser
485 490 495aga act gta gaa ctt gga aaa cgc ttc ttt cca cgt tgt tca gaa gtt 1536Arg Thr Val Glu Leu Gly Lys Arg Phe Phe Pro Arg Cys Ser Glu Val
500 505 510cta aat aag atc atg gat gct gat gac ttg tct gag ata gct tac atg 1584Leu Asn Lys Ile Met Asp Ala Asp Asp Leu Ser Glu Ile Ala Tyr Met
515 520 525ggg aat gat acg gca gaa gag cgt caa ctg aag aag caa agg tac atg 1632Gly Asn Asp Thr Ala Glu Glu Arg Gln Leu Lys Lys Gln Arg Tyr Met
530 535 540gaa ctt caa gaa att ctg act aaa gca ttc act gag gat aaa gaa gaa 1680Glu Leu Gln Glu Ile Leu Thr Lys Ala Phe Thr Glu Asp Lys Glu Glu545 550 555 560tat gat aag act aac aac atc tcc tca tct tgt tcc tct aca tct aag 1728Tyr Asp Lys Thr Asn Asn Ile Ser Ser Ser Cys Ser Ser Thr Ser Lys
565 570 575gga gta gat aag ccc aat aag ctc cct ttt agg aaa tag 1767Gly Val Asp Lys Pro Asn Lys Leu Pro Phe Arg Lys
580 585<210>2<211>588<212>PRT<213>烟草<400>2Met Asp Asn Ser Arg Thr Ala Phe Ser Asp Ser Asn Asp Ile Ser Gly1 5 10 15Ser Ser Ser Ile Cys Cys Ile Gly Gly Gly Met Thr Glu Phe Phe Ser
20 25 30Pro Glu Thr Ser Pro Ala Glu Ile Thr Ser Leu Lys Arg Leu Ser Glu
35 40 45Thr Leu Glu Ser Ile Phe Asp Ala Ser Leu Pro Glu Phe Asp Tyr Phe
50 55 60Ala Asp Ala Lys Leu Val Val Ser Gly Pro Cys Lys Glu Ile Pro Val65 70 75 80His Arg Cys Ile Leu Ser Ala Arg Ser Pro Phe Phe Lys Asn Leu Phe
85 90 95Cys Gly Lys Lys Glu Lys Asn Ser Ser Lys Val Glu Leu Lys Glu Val
100 105 110Met Lys Glu His Glu Val Ser Tyr Asp Ala Val Met Ser Val Leu Ala
115 120 125Tyr Leu Tyr Ser Gly Lys Val Arg Pro Ser Pro Lys Asp Val Cys Val
130 135 140Cys Val Asp Asn Asp Cys Ser His Val Ala Cys Arg Pro Ala Val Ala145 150 155 160Phe Leu Val Glu Val Leu Tyr Thr Ser Phe Thr Phe Gln Ile Ser Glu
165 170 175Leu Val Asp Lys Phe Gln Arg His Leu Leu Asp Ile Leu Asp Lys Thr
180 185 190Ala Ala Asp Asp Val Met Met Val Leu Ser Val Ala Asn Ile Cys Gly
195 200 205Lys Ala Cys Glu Arg Leu Leu Ser Ser Cys Ile Glu Ile Ile Val Lys
210 215 220Ser Asn Val Asp Ile Ile Thr Leu Asp Lys Ala Leu Pro His Asp Ile225 230 235 240Val Lys Gln Ile Thr Asp Ser Arg Ala Glu Leu Gly Leu Gln Gly Pro
245 250 255Glu Ser Asn Gly Phe Pro Asp Lys His Val Lys Arg Ile His Arg Ala
260 265 270Leu Asp Ser Asp Asp Val Glu Leu Leu Gln Met Leu Leu Arg Glu Gly
275 280 285His Thr Thr Leu Asp Asp Ala Tyr Ala Leu His Tyr Ala Val Ala Tyr
290 295 300Cys Asp Ala Lys Thr Thr Ala Glu Leu Leu Asp Leu Ala Leu Ala Asp305 310 315 320Ile Asn His Gln Asn Ser Arg Gly Tyr Thr Val Leu His Val Ala Ala
325 330 335Met Arg Lys Glu Pro Lys Ile Val Val Ser Leu Leu Thr Lys Gly Ala
340 345 350Arg Pro Ser Asp Leu Thr Ser Asp Gly Arg Lys Ala Leu Gln Ile Ala
355 360 365Lys Arg Leu Thr Arg Leu Val Asp Phe Ser Lys Ser Pro Glu Glu Gly
370 375 380Lys Ser Ala Ser Asn Asp Arg Leu Cys Ile Glu Ile Leu Glu Gln Ala385 390 395 400Glu Arg Arg Asp Pro Leu Leu Gly Glu Ala Ser Val Ser Leu Ala Met
405 410 415Ala Gly Asp Asp Leu Arg Met Lys Leu Leu Tyr Leu Glu Asn Arg Val
420 425 430Gly Leu Ala Lys Leu Leu Phe Pro Met Glu Ala Lys Val Ala Met Asp
435 440 445Ile Ala Gln Val Asp Gly Thr Ser Glu Phe Pro Leu Ala Ser Ile Gly
450 455 460Lys Lys Met Ala Asn Ala Gln Arg Thr Thr Val Asp Leu Asn Glu Ala465 470 475 480Pro Phe Lys Ile Lys Glu Glu His Leu Asn Arg Leu Arg Ala Leu Ser
485 490 495Arg Thr Val Glu Leu Gly Lys Arg Phe Phe Pro Arg Cys Ser Glu Val
500 505 510Leu Asn Lys Ile Met Asp Ala Asp Asp Leu Ser Glu Ile Ala Tyr Met
515 520 525Gly Asn Asp Thr Ala Glu Glu Arg Gln Leu Lys Lys Gln Arg Tyr Met
530 535 540Glu Leu Gln Glu Ile Leu Thr Lys Ala Phe Thr Glu Asp Lys Glu Glu545 550 555 560Tyr Asp Lys Thr Asn Asn Ile Ser Ser Ser Cys Ser Ser Thr Ser Lys
565 570 575Gly Val Asp Lys Pro Asn Lys Leu Pro Phe Arg Lys
580 585<210>3<211>1731<212>DNA<213>番茄<220><221>CDS<222>(1)..(1728)<223>全长番茄cDNA序列<400>3atg gat agt aga act gct ttt tcg gat tcc aat gat att agt gga agc 48Met Asp Ser Arg Thr Ala Phe Ser Asp Ser Asn Asp Ile Ser Gly Ser1 5 10 15agt agt ata tgc tgc atg aac gaa tcg gaa act tca ctg gca gac gtc 96Ser Ser Ile Cys Cys Met Asn Glu Ser Glu Thr Ser Leu Ala Asp Val
20 25 30aat tcc ctc aaa cgt cta tca gaa aca cta gag tct atc ttc gat gcg 144Asn Ser Leu Lys Arg Leu Ser Glu Thr Leu Glu Ser Ile Phe Asp Ala
35 40 45tct gcg ccg gat ttc gac ttc ttc gct gat gct aag ctt ctg gct cca 192Ser Ala Pro Asp Phe Asp Phe Phe Ala Asp Ala Lys Leu Leu Ala Pro
50 55 60ggc ggt aag gaa att ccg gtg cat cgg tgc att ttg tcg gcg agg agt 240Gly Gly Lys Glu Ile Pro Val His Arg Cys Ile Leu Ser Ala Arg Ser65 70 75 80cct ttt ttt aag aat gta ttc tgt ggg aaa gat agc agc acg aag ctg 288Pro Phe Phe Lys Asn Val Phe Cys Gly Lys Asp Ser Ser Thr Lys Leu
85 90 95gaa ctc aaa gag ctg atg aaa gag tat gag gtg agt ttt gat gcc gtg 336Glu Leu Lys Glu Leu Met Lys Glu Tyr Glu Val Ser Phe Asp Ala Val
100 105 110gtc agt gtg ctc gcc tat ttg tat agt gga aaa gtt agg cct gca tct 384Val Ser Val Leu Ala Tyr Leu Tyr Ser Gly Lys Val Arg Pro Ala Ser
115 120 125aaa gat gtg tgt gtt tgt gtg gac aat gag tgc ttg cat gta gct tgt 432Lys Asp Val Cys Val Cys Val Asp Asn Glu Cys Leu His Val Ala Cys
130 135 140agg cca gct gtg gcc ttc atg gtt cag gtt ttg tac gca tcc ttt acc 480Arg Pro Ala Val Ala Phe Met Val Gln Val Leu Tyr Ala Ser Phe Thr145 150 155 160ttt cag atc tct caa ttg gtc gac aag ttt cag aga cac cta ttg gat 528Phe Gln Ile Ser Gln Leu Val Asp Lys Phe Gln Arg His Leu Leu Asp
165 170 175att ctt gac aaa gct gta gca gat gat gta atg atg gtt tta tcc gtt 576Ile Leu Asp Lys Ala Val Ala Asp Asp Val Met Met Val Leu Ser Val
180 185 190gca aac att tgc ggt aaa gca tgt gaa aga tta ctt tca aga tgc att 624Ala Asn Ile Cys Gly Lys Ala Cys Glu Arg Leu Leu Ser Arg Cys Ile
195 200 205gat att att gtc aag tct aat gtt gat atc ata acc ctt gat aag tcc 672Asp Ile Ile Val Lys Ser Asn Val Asp Ile Ile Thr Leu Asp Lys Ser
210 215 220ttg cct cat gac att gta aaa caa atc act gat tca cgt gct gaa ctt 720Leu Pro His Asp Ile Val Lys Gln Ile Thr Asp Ser Arg Ala Glu Leu225 230 235 240ggt ctg caa ggg cct gaa agc aat ggt ttt cct gat aaa cat gtt aag 768Gly Leu Gln Gly Pro Glu Ser Asn Gly Phe Pro Asp Lys His Val Lys
245 250 255agg ata cat aga gca ttg gac tct gat gat gtt gaa tta cta agg atg 816Arg Ile His Arg Ala Leu Asp Ser Asp Asp Val Glu Leu Leu Arg Met
260 265 270ttg ctt aaa gag ggg cat act act ctt gat gat gca tat gct ctc cac 864Leu Leu Lys Glu Gly His Thr Thr Leu Asp Asp Ala Tyr Ala Leu His
275 280 285tat gct gta gca tat tgc gat gca aag act aca gca gaa ctt tta gat 912Tyr Ala Val Ala Tyr Cys Asp Ala Lys Thr Thr Ala Glu Leu Leu Asp
290 295 300ctt tca ctt gct gat gtt aat cat caa aat cct aga gga cac acg gta 960Leu Ser Leu Ala Asp Val Asn His Gln Asn Pro Arg Gly His Thr Val305 310 315 320ctt cat gtt gct gcc atg agg aaa gaa cct aaa att ata gtg tcc ctt 1008Leu His Val Ala Ala Met Arg Lys Glu Pro Lys Ile Ile Val Ser Leu
325 330 335tta acc aaa gga gct aga cct tct gat ctg aca tcc gat ggc aaa aaa 1056Leu Thr Lys Gly Ala Arg Pro Ser Asp Leu Thr Ser Asp Gly Lys Lys
340 345 350gca ctt caa att gct aag agg ctc act agg ctt gta gat ttt acc aag 1104Ala Leu Gln Ile Ala Lys Arg Leu Thr Arg Leu Val Asp Phe Thr Lys
355 360 365tct aca gag gaa gga aaa tct gct cca aag gat cgg tta tgc att gag 1152Ser Thr Glu Glu Gly Lys Ser Ala Pro Lys Asp Arg Leu Cys Ile Glu
370 375 380att ctg gag caa gca gaa aga aga gat cca cta cta gga gaa gct tca 1200Ile Leu Glu Gln Ala Glu Arg Arg Asp Pro Leu Leu Gly Glu Ala Ser385 390 395 400tta tct ctt gct atg gca ggc gat gat ttg cgt atg aag ctg tta tac 1248Leu Ser Leu Ala Met Ala Gly Asp Asp Leu Arg Met Lys Leu Leu Tyr
405 410 415ctt gaa aat aga gtt ggt ctg gct aaa ctc ctt ttt ccc atg gaa gca 1296Leu Glu Asn Arg Val Gly Leu Ala Lys Leu Leu Phe Pro Met Glu Ala
420 425 430aaa gtt gca atg gac att gca caa gtt gat ggc acg tct gaa tta ccc 1344Lys Val Ala Met Asp Ile Ala Gln Val Asp Gly Thr Ser Glu Leu Pro
435 440 445ctg gct agc atg agg aag aag ata gct gat gca cag agg aca aca gtg 1392Leu Ala Ser Met Arg Lys Lys Ile Ala Asp Ala Gln Arg Thr Thr Val
450 455 460gat ttg aac gag gct cct ttc aag atg aaa gag gag cac ttg aat cgg 1440Asp Leu Asn Glu Ala Pro Phe Lys Met Lys Glu Glu His Leu Asn Arg465 470 475 480ctt agg gct ctc tct aga act gtg gaa ctt gga aaa cgg ttc ttt cca 1488Leu Arg Ala Leu Ser Arg Thr Val Glu Leu Gly Lys Arg Phe Phe Pro
485 490 495cgt tgt tca gaa gtt cta aat aag atc atg gat gct gat gac ttg tct 1536Arg Cys Ser Glu Val Leu Asn Lys Ile Met Asp Ala Asp Asp Leu Ser
500 505 510gag ata gct tac atg ggg aat gat aca gta gaa gag cgt caa ctg aag 1584Glu Ile Ala Tyr Met Gly Asn Asp Thr Val Glu Glu Arg Gln Leu Lys
515 520 525aag caa agg tac atg gaa ctt caa gaa att ttg tct aaa gca ttc acg 1632Lys Gln Arg Tyr Met Glu Leu Gln Glu Ile Leu Ser Lys Ala Phe Thr
530 535 540gag gat aaa gaa gaa ttt gct aag act aac atg tcc tca tct tgt tcc 1680Glu Asp Lys Glu Glu Phe Ala Lys Thr Asn Met Ser Ser Ser Cys Ser545 550 555 560tct aca tct aag gga gta gat aag ccc aat aat ctc cca ttt agg aaa 1728Ser Thr Ser Lys Gly Val Asp Lys Pro Asn Asn Leu Pro Phe Arg Lys
565 570 575tag 1731<210>4<211>576<212>PRT<213>番茄<400>4Met Asp Ser Arg Thr Ala Phe Ser Asp Ser Asn Asp Ile Ser Gly Ser1 5 10 15Ser Ser Ile Cys Cys Met Asn Glu Ser Glu Thr Ser Leu Ala Asp Val
20 25 30Asn Ser Leu Lys Arg Leu Ser Glu Thr Leu Glu Ser Ile Phe Asp Ala
35 40 45Ser Ala Pro Asp Phe Asp Phe Phe Ala Asp Ala Lys Leu Leu Ala Pro
50 55 60Gly Gly Lys Glu Ile Pro Val His Arg Cys Ile Leu Ser Ala Arg Ser65 70 75 80Pro Phe Phe Lys Asn Val Phe Cys Gly Lys Asp Ser Ser Thr Lys Leu
85 90 95Glu Leu Lys Glu Leu Met Lys Glu Tyr Glu Val Ser Phe Asp Ala Val
100 105 110Val Ser Val Leu Ala Tyr Leu Tyr Ser Gly Lys Val Arg Pro Ala Ser
115 120 125Lys Asp Val Cys Val Cys Val Asp Asn Glu Cys Leu His Val Ala Cys
130 135 140Arg Pro Ala Val Ala Phe Met Val Gln Val Leu Tyr Ala Ser Phe Thr145 150 155 160Phe Gln Ile Ser Gln Leu Val Asp Lys Phe Gln Arg His Leu Leu Asp
165 170 175Ile Leu Asp Lys Ala Val Ala Asp Asp Val Met Met Val Leu Ser Val
180 185 190Ala Asn Ile Cys Gly Lys Ala Cys Glu Arg Leu Leu Ser Arg Cys Ile
195 200 205Asp Ile Ile Val Lys Ser Asn Val Asp Ile Ile Thr Leu Asp Lys Ser
210 215 220Leu Pro His Asp Ile Val Lys Gln Ile Thr Asp Ser Arg Ala Glu Leu225 230 235 240Gly Leu Gln Gly Pro Glu Ser Asn Gly Phe Pro Asp Lys His Val Lys
245 250 255Arg Ile His Arg Ala Leu Asp Ser Asp Asp Val Glu Leu Leu Arg Met
260 265 270Leu Leu Lys Glu Gly His Thr Thr Leu Asp Asp Ala Tyr Ala Leu His
275 280 285Tyr Ala Val Ala Tyr Cys Asp Ala Lys Thr Thr Ala Glu Leu Leu Asp
290 295 300Leu Ser Leu Ala Asp Val Asn His Gln Asn Pro Arg Gly His Thr Val305 310 315 320Leu His Val Ala Ala Met Arg Lys Glu Pro Lys Ile Ile Val Ser Leu
325 330 335Leu Thr Lys Gly Ala Arg Pro Ser Asp Leu Thr Ser Asp Gly Lys Lys
340 345 350Ala Leu Gln Ile Ala Lys Arg Leu Thr Arg Leu Val Asp Phe Thr Lys
355 360 365Ser Thr Glu Glu Gly Lys Ser Ala Pro Lys Asp Arg Leu Cys Ile Glu
370 375 380Ile Leu Glu Gln Ala Glu Arg Arg Asp Pro Leu Leu Gly Glu Ala Ser385 390 395 400Leu Ser Leu Ala Met Ala Gly Asp Asp Leu Arg Met Lys Leu Leu Tyr
405 410 415Leu Glu Asn Arg Val Gly Leu Ala Lys Leu Leu Phe Pro Met Glu Ala
420 425 430Lys Val Ala Met Asp Ile Ala Gln Val Asp Gly Thr Ser Glu Leu Pro
435 440 445Leu Ala Ser Met Arg Lys Lys Ile Ala Asp Ala Gln Arg Thr Thr Val
450 455 460Asp Leu Asn Glu Ala Pro Phe Lys Met Lys Glu Glu His Leu Asn Arg465 470 475 480Leu Arg Ala Leu Ser Arg Thr Val Glu Leu Gly Lys Arg Phe Phe Pro
485 490 495Arg Cys Ser Glu Val Leu Asn Lys Ile Met Asp Ala Asp Asp Leu Ser
500 505 510Glu Ile Ala Tyr Met Gly Asn Asp Thr Val Glu Glu Arg Gln Leu Lys
515 520 525Lys Gln Arg Tyr Met Glu Leu Gln Glu Ile Leu Ser Lys Ala Phe Thr
530 535 540Glu Asp Lys Glu Glu Phe Ala Lys Thr Asn Met Ser Ser Ser Cys Ser545 550 555 560Ser Thr Ser Lys Gly Val Asp Lys Pro Asn Asn Leu Pro Phe Arg Lys
565 570 575<210>5<211>1740<212>DNA<213>欧洲油菜<220><221>CDS<222>(1)..(1737)<223>Canola cDNA序列<400>5atg gag acc att gct rga ttt gat gat ttc tat gag atc agc agc act 48Met Glu Thr Ile Ala Xaa Phe Asp Asp Phe Tyr Glu Ile Ser Ser Thr1 5 10 15agc ttc cyc gcc gca ccg gcg cca acc gat aac tcc gga tca tcc acc 96Ser Phe Xaa Ala Ala Pro Ala Pro Thr Asp Asn Ser Gly Ser Ser Thr
20 25 30gtc twc ccg acg gag ctt ytc acc aga ccc gag gta tcc gcg ttt caa 144Val Xaa Pro Thr Glu Leu Xaa Thr Arg Pro Glu Val Ser Ala Phe Gln
35 40 45ctc ctc tcc aac agc ctc gag tcc gtc ttc gac tcg ccg gaa gcg ttc 192Leu Leu Ser Asn Ser Leu Glu Ser Val Phe Asp Ser Pro Glu Ala Phe
50 55 60tac agc gac gcc aag ctt gtt ctc tcc gac gac aag gaa gta tcc ttc 240Tyr Ser Asp Ala Lys Leu Val Leu Ser Asp Asp Lys Glu Val Ser Phe65 70 75 80cac cgt tgc att ctc tcg gcg aga agc ctc ttc ttc aag gcc gct ttg 288His Arg Cys Ile Leu Ser Ala Arg Ser Leu Phe Phe Lys Ala Ala Leu
85 90 95rca gcc gcc gag aag gtg cag aag tcc acc ccc gtg aag ctc gag ctg 336Xaa Ala Ala Glu Lys Val Gln Lys Ser Thr Pro Val Lys Leu Glu Leu
100 105 110aag aca ctc gcg gcg gaa tac gac gtc ggg ttc gat tct gtg gtg gct 384Lys Thr Leu Ala Ala Glu Tyr Asp Val Gly Phe Asp Ser Val Val Ala
115 120 125gtt ctg gcg tac gtt tac agc ggc aga gtg agg ccg cct ccg aag gga 432Val Leu Ala Tyr Val Tyr Ser Gly Arg Val Arg Pro Pro Pro Lys Gly
130 135 140gtt tct gaa tgc gca gac gak agc tgc tgc cac gtg gcg tgc cgt ccg 480Val Ser Glu Cys Ala Asp Xaa Ser Cys Cys His Val Ala Cys Arg Pro145 150 155 160gct gtg gat ttc atg gtg gag gtt ctc tac ttg gct ttc gtc ttc cag 528Ala Val Asp Phe Met Val Glu Val Leu Tyr Leu Ala Phe Val Phe Gln
165 170 175att cag gaa ctg gtt acc atg tat cag agg cat tta ctg gat gtt gta 576Ile Gln Glu Leu Val Thr Met Tyr Gln Arg His Leu Leu Asp Val Val
180 185 190gac aaa gtt awc ata gaa gac act ttg gtc gtc ctc aag ctt gct aac 624Asp Lys Val Xaa Ile Glu Asp Thr Leu Val Val Leu Lys Leu Ala Asn
195 200 205atc tgc ggt aaa gcg tgc aag aag cta ttc gat aag tgc aga gag atc 672Ile Cys Gly Lys Ala Cys Lys Lys Leu Phe Asp Lys Cys Arg Glu Ile
210 215 220att gtc aag tct aac gtg gat gtt gtt act cta aag aag tca ttg cct 720Ile Val Lys Ser Asn Val Asp Val Val Thr Leu Lys Lys Ser Leu Pro225 230 235 240gag rac att gcc aag caa gta atc gat atc cgc aaa gag ctc ggc ttg 768Glu Xaa Ile Ala Lys Gln Val Ile Asp Ile Arg Lys Glu Leu Gly Leu
245 250 255gag gta gct gaa cca gag aaa cat gtc tcc aac ata cac aag gcg ctt 816Glu Val Ala Glu Pro Glu Lys His Val Ser Asn Ile His Lys Ala Leu
260 265 270gag tca gac gat ctt gac ctt gtc gtt atg ctt ttg aaa gag ggc cac 864Glu Ser Asp Asp Leu Asp Leu Val Val Met Leu Leu Lys Glu Gly His
275 280 285acg aat cta gac gaa gcg tat gct ctc cat ttt gct gtt gcg tat tgc 912Thr Asn Leu Asp Glu Ala Tyr Ala Leu His Phe Ala Val Ala Tyr Cys
290 295 300gat gag aag aca gcg agg aat ctc ctg gaa ctg ggg ttt gcg gat gtc 960Asp Glu Lys Thr Ala Arg Asn Leu Leu Glu Leu Gly Phe Ala Asp Val305 310 315 320aac cgg aga aac ccg aga ggg tac acg gta att cac gtc gct gcg atg 1008Asn Arg Arg Asn Pro Arg Gly Tyr Thr Val Ile His Val Ala Ala Met
325 330 335agg aaa gag ccg aca ctg ata gca ttg ttg ttg acg aaa ggg gct aat 1056Arg Lys Glu Pro Thr Leu Ile Ala Leu Leu Leu Thr Lys Gly Ala Asn
340 345 350gca tta gaa atg tct ttg gac ggg aga act gct ctg ttg atc gcg aaa 1104Ala Leu Glu Met Ser Leu Asp Gly Arg Thr Ala Leu Leu Ile Ala Lys
355 360 365caa gtc act aag gcg gcc gag tgt tgt att ctg gag aaa ggg aag tta 1152Gln Val Thr Lys Ala Ala Glu Cys Cys Ile Leu Glu Lys Gly Lys Leu
370 375 380gct gcc aaa ggc gga gta tgt gta gag ata ctc aag caa cca gac aac 1200Ala Ala Lys Gly Gly Val Cys Val Glu Ile Leu Lys Gln Pro Asp Asn385 390 395 400aca cga gaa cca ttt cct gaa gat gtt tct ccc tcc ctt gca gtg gct 1248Thr Arg Glu Pro Phe Pro Glu Asp Val Ser Pro Ser Leu Ala Val Ala
405 410 415gct gat caa ttc aag ata agg ttg att gat ctt gaa aac aga gtt caa 1296Ala Asp Gln Phe Lys Ile Arg Leu Ile Asp Leu Glu Asn Arg Val Gln
420 425 430atg gct cga tgt ctc tat cca atg gaa gca caa gtt gca atg gat ttc 1344Met Ala Arg Cys Leu Tyr Pro Met Glu Ala Gln Val Ala Met Asp Phe
435 440 445gcc cga atg aag gga aca cgc gag ttt gtc gtg acg aca gca act gac 1392Ala Arg Met Lys Gly Thr Arg Glu Phe Val Val Thr Thr Ala Thr Asp
450 455 460cta cac atg gaa cct ttc aag ttc gta gaa atg cat cag agt aga cta 1440Leu His Met Glu Pro Phe Lys Phe Val Glu Met His Gln Ser Arg Leu465 470 475 480aca gcg ctt tct aaa act gtg gaa ttc ggg aaa cgc ttc ttc cca cgc 1488Thr Ala Leu Ser Lys Thr Val Glu Phe Gly Lys Arg Phe Phe Pro Arg
485 490 495tgt tcg aaa gtg ctc gat gat att gtg gac tct gag gac ttg act ata 1536Cys Ser Lys Val Leu Asp Asp Ile Val Asp Ser Glu Asp Leu Thr Ile
500 505 510ctg gct ctc gta gaa gaa gac act cct gag caa cga caa caa aag agg 1584Leu Ala Leu Val Glu Glu Asp Thr Pro Glu Gln Arg Gln Gln Lys Arg
515 520 525cag agg ttc atg gaa ata cag gag att gtt caa atg gcg ttt agt aaa 1632Gln Arg Phe Met Glu Ile Gln Glu Ile Val Gln Met Ala Phe Ser Lys
530 535 540gac aag gag gat ctt gga aag tcg tct ctc tca gct tcg tct tct tcc 1680Asp Lys Glu Asp Leu Gly Lys Ser Ser Leu Ser Ala Ser Ser Ser Ser545 550 555 560aca tcc aaa tta act ggt aaa aag agg tct att gct aaa ccc tct cac 1728Thr Ser Lys Leu Thr Gly Lys Lys Arg Ser Ile Ala Lys Pro Ser His
565 570 575cgg cgt cgg tga 1740ArgArg Arg<210>6<211>579<212>PRT<213>欧洲油菜<400>6Met Glu Thr Ile Ala Xaa Phe Asp Asp Phe Tyr Glu Ile Ser Ser Thr1 5 10 15Ser Phe Xaa Ala Ala Pro Ala Pro Thr Asp Asn Ser Gly Ser Ser Thr
20 25 30Val Xaa Pro Thr Glu Leu Xaa Thr Arg Pro Glu Val Ser Ala Phe Gln
35 40 45Leu Leu Ser Asn Ser Leu Glu Ser Val Phe Asp Ser Pro Glu Ala Phe
50 55 60Tyr Ser Asp Ala Lys Leu Val Leu Ser Asp Asp Lys Glu Val Ser Phe65 70 75 80His Arg Cys Ile Leu Ser Ala Arg Ser Leu Phe Phe Lys Ala Ala Leu
85 90 95Xaa Ala Ala Glu Lys Val Gln Lys Ser Thr Pro Val Lys Leu Glu Leu
100 105 110Lys Thr Leu Ala Ala Glu Tyr Asp Val Gly Phe Asp Ser Val Val Ala
115 120 125Val Leu Ala Tyr Val Tyr Ser Gly Arg Val Arg Pro Pro Pro Lys Gly
130 135 140Val Ser Glu Cys Ala Asp Xaa Ser Cys Cys His Val Ala Cys Arg Pro145 150 155 160Ala Val Asp Phe Met Val Glu Val Leu Tyr Leu Ala Phe Val Phe Gln
165 170 175Ile Gln Glu Leu Val Thr Met Tyr Gln Arg His Leu Leu Asp Val Val
180 185 190Asp Lys Val Xaa Ile Glu Asp Thr Leu Val Val Leu Lys Leu Ala Asn
195 200 205Ile Cys Gly Lys Ala Cys Lys Lys Leu Phe Asp Lys Cys Arg Glu Ile
210 215 220Ile Val Lys Ser Asn Val Asp Val Val Thr Leu Lys Lys Ser Leu Pro225 230 235 240Glu Xaa Ile Ala Lys Gln Val Ile Asp Ile Arg Lys Glu Leu Gly Leu
245 250 255Glu Val Ala Glu Pro Glu Lys His Val Ser Asn Ile His Lys Ala Leu
260 265 270Glu Ser Asp Asp Leu Asp Leu Val Val Met Leu Leu Lys Glu Gly His
275 280 285Thr Asn Leu Asp Glu Ala Tyr Ala Leu His Phe Ala Val Ala Tyr Cys
290 295 300Asp Glu Lys Thr Ala Arg Asn Leu Leu Glu Leu Gly Phe Ala Asp Val305 310 315 320Asn Arg Arg Asn Pro Arg Gly Tyr Thr Val Ile His Val Ala Ala Met
325 330 335Arg Lys Glu Pro Thr Leu Ile Ala Leu Leu Leu Thr Lys Gly Ala Asn
340 345 350Ala Leu Glu Met Ser Leu Asp Gly Arg Thr Ala Leu Leu Ile Ala Lys
355 360 365Gln Val Thr Lys Ala Ala Glu Cys Cys Ile Leu Glu Lys Gly Lys Leu
370 375 380Ala Ala Lys Gly Gly Val Cys Val Glu Ile Leu Lys Gln Pro Asp Asn385 390 395 400Thr Arg Glu Pro Phe Pro Glu Asp Val Ser Pro Ser Leu Ala Val Ala
405 410 415Ala Asp Gln Phe Lys Ile Arg Leu Ile Asp Leu Glu Asn Arg Val Gln
420 425 430Met Ala Arg Cys Leu Tyr Pro Met Glu Ala Gln Val Ala Met Asp Phe
435 440 445Ala Arg Met Lys Gly Thr Arg Glu Phe Val Val Thr Thr Ala Thr Asp
450 455 460Leu His Met Glu Pro Phe Lys Phe Val Glu Met His Gln Ser Arg Leu465 470 475 480Thr Ala Leu Ser Lys Thr Val Glu Phe Gly Lys Arg Phe Phe Pro Arg
485 490 495Cys Ser Lys Val Leu Asp Asp Ile Val Asp Ser Glu Asp Leu Thr Ile
500 505 510Leu Ala Leu Val Glu Glu Asp Thr Pro Glu Gln Arg Gln Gln Lys Arg
515 520 525Gln Arg Phe Met Glu Ile Gln Glu Ile Val Gln Met Ala Phe Ser Lys
530 535 540Asp Lys Glu Asp Leu Gly Lys Ser Ser Leu Ser Ala Ser Ser Ser Ser545 550 555 560Thr Ser Lys Leu Thr Gly Lys Lys Arg Ser Ile Ala Lys Pro Ser His
565 570 575Arg Arg Arg<210>7<211>1761<212>DNA<213>鼠耳芥<220><221>CDS<222>(1)..(1758)<223>AtNMLC5 cDNA序列<400>7atg gct act ttg act gag cca tca tca tct ttg agt ttc aca tct tct 48Met Ala Thr Leu Thr Glu Pro Ser Ser Ser Leu Ser Phe Thr Ser Ser1 5 10 15cat ttc tct tat ggt tct att ggg tcc aat cac ttc tca tca agc tca 96His Phe Ser Tyr Gly Ser Ile Gly Ser Asn His Phe Ser Ser Ser Ser
20 25 30gct tct aat cct gaa gtt gtt agt cta acc aaa ctc agc tcc aat ctt 144Ala Ser Asn Pro Glu Val Val Ser Leu Thr Lys Leu Ser Ser Asn Leu
35 40 45gag cag ctt ctt agt aat tca gat tgt gat tac agt gat gca gag atc 192Glu Gln Leu Leu Ser Asn Ser Asp Cys Asp Tyr Ser Asp Ala Glu Ile
50 55 60att gtt gat ggt gtt cca gtt ggt gtt cat aga tgc att tta gct gca 240Ile Val Asp Gly Val Pro Val Gly Val His Arg Cys Ile Leu Ala Ala65 70 75 80aga agt aag ttt ttc caa gat ttg ttt aag aaa gaa aag aaa att tcg 288Arg Ser Lys Phe Phe Gln Asp Leu Phe Lys Lys Glu Lys Lys Ile Ser
85 90 95aaa act gag aaa cca aag tat cag ttg aga gag atg tta cct tat gga 336Lys Thr Glu Lys Pro Lys Tyr Gln Leu Arg Glu Met Leu Pro Tyr Gly
100 105 110gct gtt gct cat gaa gct ttc ttg tat ttc ttg agt tat ata tat act 384Ala Val Ala His Glu Ala Phe Leu Tyr Phe Leu Ser Tyr Ile Tyr Thr
115 120 125ggg aga tta aag cct ttt cca ttg gag gtt tcg act tgt gtt gat cca 432Gly Arg Leu Lys Pro Phe Pro Leu Glu Val Ser Thr Cys Val Asp Pro
130 135 140gtt tgt tct cat gat tgt tgt cga cct gcc att gat ttt gtt gtt caa 480Val Cys Ser His Asp Cys Cys Arg Pro Ala Ile Asp Phe Val Val Gln145 150 155 160ttg atg tat gct tcc tct gtt ctc caa gtg cct gag cta gtt tca tct 528Leu Met Tyr Ala Ser Ser Val Leu Gln Val Pro Glu Leu Val Ser Ser
165 170 175ttt cag cgg cgg ctt tgt aac ttt gtg gag aag acc ctt gtt gag aat 576Phe Gln Arg Arg Leu Cys Asn Phe Val Glu Lys Thr Leu Val Glu Asn
180 185 190gtt ctt ccc att ctt atg gtt gct ttc aat tgt aag ttg act cag ctt 624Val Leu Pro Ile Leu Met Val Ala Phe Asn Cys Lys Leu Thr Gln Leu
195 200 205ctt gat cag tgt att gag aga gtg gcg agg tca gat ctt tac agg ttc 672Leu Asp Gln Cys Ile Glu Arg Val Ala Arg Ser Asp Leu Tyr Arg Phe
210 215 220tgt att gaa aag gaa gtt cct ccc gaa gta gca gag aag att aaa cag 720Cys Ile Glu Lys Glu Val Pro Pro Glu Val Ala Glu Lys Ile Lys Gln225 230 235 240ctt cga ctt ata tcc ccg caa gac gaa gaa acc agt ccc aag att tcg 768Leu Arg Leu Ile Ser Pro Gln Asp Glu Glu Thr Ser Pro Lys Ile Ser
245 250 255gag aaa ttg ctt gaa aga atc ggt aaa att ctc aag gcc ttg gat tca 816Glu Lys Leu Leu Glu Arg Ile Gly Lys Ile Leu Lys Ala Leu Asp Ser
260 265 270gat gat gtt gag ctt gtg aag ctt ctt ttg act gag tca gat atc act 864Asp Asp Val Glu Leu Val Lys Leu Leu Leu Thr Glu Ser Asp Ile Thr
275 280 285cta gat caa gcc aat ggt ctg cat tat tct gtt gtg tat agt gat ccg 912Leu Asp Gln Ala Asn Gly Leu His Tyr Ser Val Val Tyr Ser Asp Pro
290 295 300aaa gtt gtt gcc gag att ctt gct ctg gat atg ggt gat gtg aac tac 960Lys Val Val Ala Glu Ile Leu Ala Leu Asp Met Gly Asp Val Asn Tyr305 310 315 320agg aat tcc cgg ggt tac acg gtt ctt cat ttt gct gcg atg cgt aga 1008Arg Asn Ser Arg Gly Tyr Thr Val Leu His Phe Ala Ala Met Arg Arg
325 330 335gag cca tcg atc att ata tcg ctt atc gat aaa ggc gcc aat gca tct 1056Glu Pro Ser Ile Ile Ile Ser Leu Ile Asp Lys Gly Ala Asn Ala Ser
340 345 350gag ttt aca tct gac gga cgc agc gca gtt aat ata ttg aga aga ctg 1104Glu Phe Thr Ser Asp Gly Arg Ser Ala Val Asn Ile Leu Arg Arg Leu
355 360 365aca aat cca aag gat tat cat acc aaa aca gca aaa ggg cgt gaa tct 1152Thr Asn Pro Lys Asp Tyr His Thr Lys Thr Ala Lys Gly Arg Glu Ser
370 375 380agt aag gcc agg cta tgc atc gat ata ttg gaa aga gaa atc agg aag 1200Ser Lys Ala Arg Leu Cys Ile Asp Ile Leu Glu Arg Glu Ile Arg Lys385 390 395 400aac ccc atg gtt cta gat aca cca atg tgt tcc att tct atg cct gaa 1248Asn Pro Met Val Leu Asp Thr Pro Met Cys Ser Ile Ser Met Pro Glu
405 410 415gat ctc cag atg aga ctg ttg tac cta gaa aag aga gtg ggt ctt gct 1296Asp Leu Gln Met Arg Leu Leu Tyr Leu Glu Lys Arg Val Gly Leu Ala
420 425 430cag ttg ttc ttt cca acg gaa gct aaa gtg gct atg gac att ggt aac 1344Gln Leu Phe Phe Pro Thr Glu Ala Lys Val Ala Met Asp Ile Gly Asn
435 440 445gta gaa ggt aca agt gag ttc aca ggg ttg tca cct cct tca agt ggg 1392Val Glu Gly Thr Ser Glu Phe Thr Gly Leu Ser Pro Pro Ser Ser Gly
450 455 460tta acc gga aac ttg agt cag gtt gat tta aac gaa act cct cat atg 1440Leu Thr Gly Asn Leu Ser Gln Val Asp Leu Asn Glu Thr Pro His Met465 470 475 480caa acc caa aga ctt ctt act cgt atg gtg gct cta atg aaa aca gtt 1488Gln Thr Gln Arg Leu Leu Thr Arg Met Val Ala Leu Met Lys Thr Val
485 490 495gag act ggt cga agg ttt ttt cca tat ggt tca gag gtt cta gat aag 1536Glu Thr Gly Arg Arg Phe Phe Pro Tyr Gly Ser Glu Val Leu Asp Lys
500 505 510tac atg gct gag tat ata gac gac gac atc ctc gac gat ttc cat ttt 1584Tyr Met Ala Glu Tyr Ile Asp Asp Asp Ile Leu Asp Asp Phe His Phe
515 520 525gag aag gga tct aca cat gaa aga aga ttg aaa aga atg aga tat aga 1632Glu Lys Gly Ser Thr His Glu Arg Arg Leu Lys Arg Met Arg Tyr Arg
530 535 540gag ctt aag gat gat gtc caa aag gca tat agc aaa gac aaa gag tct 1680Glu Leu Lys Asp Asp Val Gln Lys Ala Tyr Ser Lys Asp Lys Glu Ser545 550 555 560aag att gcg cgg tct tgt ctt tct gct tca tct tct cct tct tct tct 1728Lys Ile Ala Arg Ser Cys Leu Ser Ala Ser Ser Ser Pro Ser Ser Ser
565 570 575tcc ata aga gat gat ctg cac aac aca aca tga 1761Ser Ile Arg Asp Asp Leu His Asn Thr Thr
580 585<210>8<211>586<212>PRT<213>鼠耳芥<400>8Met Ala Thr Leu Thr Glu Pro Ser Ser Ser Leu Ser Phe Thr Ser Ser1 5 10 15His Phe Ser Tyr Gly Ser Ile Gly Ser Asn His Phe Ser Ser Ser Ser
20 25 30Ala Ser Asn Pro Glu Val Val Ser Leu Thr Lys Leu Ser Ser Asn Leu
35 40 45Glu Gln Leu Leu Ser Asn Ser Asp Cys Asp Tyr Ser Asp Ala Glu Ile
50 55 60Ile Val Asp Gly Val Pro Val Gly Val His Arg Cys Ile Leu Ala Ala65 70 75 80Arg Ser Lys Phe Phe Gln Asp Leu Phe Lys Lys Glu Lys Lys Ile Ser
85 90 95Lys Thr Glu Lys Pro Lys Tyr Gln Leu Arg Glu Met Leu Pro Tyr Gly
100 105 110Ala Val Ala His Glu Ala Phe Leu Tyr Phe Leu Ser Tyr Ile Tyr Thr
115 120 125Gly Arg Leu Lys Pro Phe Pro Leu Glu Val Ser Thr Cys Val Asp Pro
130 135 140Val Cys Ser His Asp Cys Cys Arg Pro Ala Ile Asp Phe Val Val Gln145 150 155 160Leu Met Tyr Ala Ser Ser Val Leu Gln Val Pro Glu Leu Val Ser Ser
165 170 175Phe Gln Arg Arg Leu Cys Asn Phe Val Glu Lys Thr Leu Val Glu Asn
180 185 190Val Leu Pro Ile Leu Met Val Ala Phe Asn Cys Lys Leu Thr Gln Leu
195 200 205Leu Asp Gln Cys Ile Glu Arg Val Ala Arg Ser Asp Leu Tyr Arg Phe
210 215 220Cys Ile Glu Lys Glu Val Pro Pro Glu Val Ala Glu Lys Ile Lys Gln225 230 235 240Leu Arg Leu Ile Ser Pro Gln Asp Glu Glu Thr Ser Pro Lys Ile Ser
245 250 255Glu Lys Leu Leu Glu Arg Ile Gly Lys Ile Leu Lys Ala Leu Asp Ser
260 265 270Asp Asp Val Glu Leu Val Lys Leu Leu Leu Thr Glu Ser Asp Ile Thr
275 280 285Leu Asp Gln Ala Asn Gly Leu His Tyr Ser Val Val Tyr Ser Asp Pro
290 295 300Lys Val Val Ala Glu Ile Leu Ala Leu Asp Met Gly Asp Val Asn Tyr305 310 315 320Arg Asn Ser Arg Gly Tyr Thr Val Leu His Phe Ala Ala Met Arg Arg
325 330 335Glu Pro Ser Ile Ile Ile Ser Leu Ile Asp Lys Gly Ala Asn Ala Ser
340 345 350Glu Phe Thr Ser Asp Gly Arg Ser Ala Val Asn Ile Leu Arg Arg Leu
355 360 365Thr Asn Pro Lys Asp Tyr His Thr Lys Thr Ala Lys Gly Arg Glu Ser
370 375 380Ser Lys Ala Arg Leu Cys Ile Asp Ile Leu Glu Arg Glu Ile Arg Lys385 390 395 400Asn Pro Met Val Leu Asp Thr Pro Met Cys Ser Ile Ser Met Pro Glu
405 410 415Asp Leu Gln Met Arg Leu Leu Tyr Leu Glu Lys Arg Val Gly Leu Ala
420 425 430Gln Leu Phe Phe Pro Thr Glu Ala Lys Val Ala Met Asp Ile Gly Asn
435 440 445Val Glu Gly Thr Ser Glu Phe Thr Gly Leu Ser Pro Pro Ser Ser Gly
450 455 460Leu Thr Gly Asn Leu Ser Gln Val Asp Leu Asn Glu Thr Pro His Met465 470 475 480Gln Thr Gln Arg Leu Leu Thr Arg Met Val Ala Leu Met Lys Thr Val
485 490 495Glu Thr Gly Arg Arg Phe Phe Pro Tyr Gly Ser Glu Val Leu Asp Lys
500 505 510Tyr Met Ala Glu Tyr Ile Asp Asp Asp Ile Leu Asp Asp Phe His Phe
515 520 525Glu Lys Gly Ser Thr His Glu Arg Arg Leu Lys Arg Met Arg Tyr Arg
530 535 540Glu Leu Lys Asp Asp Val Gln Lys Ala Tyr Ser Lys Asp Lys Glu Ser545 550 555 560Lys Ile Ala Arg Ser Cys Leu Ser Ala Ser Ser Ser Pro Ser Ser Ser
565 570 575Ser Ile Arg Asp Asp Leu His Asn Thr Thr
580 585<210>9<211>21<212>DNA<213>人工序列<220><223>人T序列描述:PCR引物<400>9agattattgt caagtctaat g 21<210>10<211>19<212>DNA<213>人工序列<220><223>人工序列描述:PCR引物<400>10ttccatgtac ctttgcttc 19<210>11<211>23<212>DNA<213>人工序列<220><223>人工序列描述:PCR引物<400>11gcggatccat ggataatagt agg 23<210>12<211>23<212>DNA<213>人工序列<220><223>人T序列描述:PCR引物<400>12gcggatccta tttcctaaaa ggg 23<210>13<211>21<212>DNA<213>人工序列<220><223>人工序列描述:PCR引物<400>13tcaaggcctt ggattcagat g 21<210>14<211>21<212>DNA<213>人工序列<220><223>人工序列描述:PCR引物<400>14attaactgcg ctacgtccgt c 21<210>15<211>1477<212>DNA<213>鼠耳芥<220><221>CDS<222>(1)..(1476)<23>AtNMLc2基因组序列<400>15atg agc aat ctt gaa gaa tct ttg aga tct cta tcg ttg gat ttc ctg 48Met Ser Asn Leu Glu Glu Ser Leu Arg Ser Leu Ser Leu Asp Phe Leu1 5 10 15aac cta cta atc aac ggt caa gct ttc tcc gac gtg act ttc agc gtt 96Asn Leu Leu Ile Asn Gly Gln Ala Phe Ser Asp Val Thr Phe Ser Val
20 25 30gaa ggt cgt tta gtc cac gct cac cgt tgt atc ctc gcc gca cgg agt 144Glu Gly Arg Leu Val His Ala His Arg Cys Ile Leu Ala Ala Arg Ser
35 40 45ctt ttc ttc cgc aaa ttc ttt tgt ggg aca gac tca cca caa cct gtc 192Leu Phe Phe Arg Lys Phe Phe Cys Gly Thr Asp Ser Pro Gln Pro Val
50 55 60aca ggt ata gac ccg acc caa cat ggg tcc gta ccc gct agc cca aca 240Thr Gly Ile Asp Pro Thr Gln His Gly Ser Val Pro Ala Ser Pro Thr65 70 75 80aga ggc tcc acg gcc cca gct gga att ata cca gtg aac tca gtc ggt 288Arg Gly Ser Thr Ala Pro Ala Gly Ile Ile Pro Val Asn Ser Val Gly
85 90 95tat gag gtt ttt ctg ttg cta ctt cag ttt ctt tat agc gga caa gtc 336Tyr Glu Val Phe Leu Leu Leu Leu Gln Phe Leu Tyr Ser Gly Gln Val
100 105 110tcc atc gtg ccg cag aaa cac gag cct aga cct aat tgt ggc gag aga 384Ser Ile Val Pro Gln Lys His Glu Pro Arg Pro Asn Cys Gly Glu Arg
115 120 125gga tgt tgg cac act cat tgc tca gcc gcc gtt gat ctt gct ctt gat 432Gly Cys Trp His Thr His Cys Ser Ala Ala Val Asp Leu Ala Leu Asp
130 135 140act ctc gcc gcc tct cgt tac ttc ggc gtc gag cag ctc gca ttg ctc 480Thr Leu Ala Ala Ser Arg Tyr Phe Gly Val Glu Gln Leu Ala Leu Leu145 150 155 160acc cag aaa caa ttg gca agc atg gtg gag aaa gcc tct atc gaa gat 528Thr Gln Lys Gln Leu Ala Ser Met Val Glu Lys Ala Ser Ile Glu Asp
165 170 175gtg atg aaa gtt tta ata gca tca aga aag caa gac atg cat caa tta 576Val Met Lys Val Leu Ile Ala Ser Arg Lys Gln Asp Met His Gln Leu
180 185 190tgg acc acc tgc tct cac tta gtt atg agc aat ctt gaa gaa tct ttg 624Trp Thr Thr Cys Ser His Leu Val Met Ser Asn Leu Glu Glu Ser Leu
195 200 205aga tct cta tcg ttg gat ttc ctg aac cta cta atc aac ggt caa gct 672Arg Ser Leu Ser Leu Asp Phe Leu Asn Leu Leu Ile Asn Gly Gln Ala
210 215 220ttc tcc gac gtg act ttc agc gtt gaa ggt cgt tta gtc cac gct cac 720Phe Ser Asp Val Thr Phe Ser Val Glu Gly Arg Leu Val His Ala His225 230 235 240cgt tgt atc ctc gcc gca cgg agt ctt ttc ttc cgc aaa ttc ttt tgt 768Arg Cys Ile Leu Ala Ala Arg Ser Leu Phe Phe Arg Lys Phe Phe Cys
245 250 255ggg aca gac tca cca caa cct gtc aca ggt ata gac ccg acc caa cat 816Gly Thr Asp Ser Pro Gln Pro Val Thr Gly Ile Asp Pro Thr Gln His
260 265 270ggg tcc gta ccc gct agc cca aca aga ggc tcc acg gcc cca gct gga 864Gly Ser Val Pro Ala Ser Pro Thr Arg Gly Ser Thr Ala Pro Ala Gly
275 280 285att ata cca gtg aac tca gtc ggt tat gag gtt ttt ctg ttg cta ctt 912Ile Ile Pro Val Asn Ser Val Gly Tyr Glu Val Phe Leu Leu Leu Leu
290 295 300cag ttt ctt tat agc gga caa gtc tcc atc gtg ccg cag aaa cac gag 960Gln Phe Leu Tyr Ser Gly Gln Val Ser Ile Val Pro Gln Lys His Glu305 310 315 320cct aga cct aat tgt ggc gag aga gga tgt tgg cac act cat tgc tca 1008Pro Arg Pro Asn Cys Gly Glu Arg Gly Cys Trp His Thr His Cys Ser
325 330 335gcc gcc gtt gat ctt gct ctt gat act ctc gcc gcc tct cgt tac ttc 1056Ala Ala Val Asp Leu Ala Leu Asp Thr Leu Ala Ala Ser Arg Tyr Phe
340 345 350ggc gtc gag cag ctc gca ttg ctc acc cag aaa caa ttg gca agc atg 1104Gly Val Glu Gln Leu Ala Leu Leu Thr Gln Lys Gln Leu Ala Ser Met
355 360 365gtg gag aaa gcc tct atc gaa gat gtg atg aaa gtt tta ata gca tca 1152Val Glu Lys Ala Ser Ile Glu Asp Val Met Lys Val Leu Ile Ala Ser
370 375 380aga aag caa gac atg cat caa tta tgg acc acc tgc tct cac tta gtt 1200Arg Lys Gln Asp Met His Gln Leu Trp Thr Thr Cys Ser His Leu Val385 390 395 400atg agc aat ctt gaa gaa tct ttg aga tct cta tcg ttg gat ttc ctg 1248Met Ser Asn Leu Glu Glu Ser Leu Arg Ser Leu Ser Leu Asp Phe Leu
405 410 415aac cta cta atc aac ggt caa gct ttc tcc gac gtg act ttc agc gtt 1296Asn Leu Leu Ile Asn Gly Gln Ala Phe Ser Asp Val Thr Phe Ser Val
420 425 430gaa ggt cgt tta gtc cac gct cac cgt tgt atc ctc gcc gca cgg agt 1344Glu Gly Arg Leu Val His Ala His Arg Cys Ile Leu Ala Ala Arg Ser
435 440 445ctt ttc ttc cgc aaa ttc ttt tgt ggg aca gac tca cca caa cct gtc 1392Leu Phe Phe Arg Lys Phe Phe Cys Gly Thr Asp Ser Pro Gln Pro Val
450 455 460aca ggt ata gac ccg acc caa cat ggg tcc gta ccc gct agc cca aca 1440Thr Gly Ile Asp Pro Thr Gln His Gly Ser Val Pro Ala Ser Pro Thr465 470 475 480aga ggc tcc acg gcc cca gct gga att ata cca gtg a 1477Arg Gly Ser Thr Ala Pro Ala Gly Ile Ile Pro Val
485 490<210>16<211>492<212>PRT<213>鼠耳芥<400>16Met Ser Asn Leu Glu Glu Ser Leu Arg Ser Leu Ser Leu Asp Phe Leu1 5 10 15Asn Leu Leu Ile Asn Gly Gln Ala Phe Ser Asp Val Thr Phe Ser Val
20 25 30Glu Gly Arg Leu Val His Ala His Arg Cys Ile Leu Ala Ala Arg Ser
35 40 45Leu Phe Phe Arg Lys Phe Phe Cys Gly Thr Asp Ser Pro Gln Pro Val
50 55 60Thr Gly Ile Asp Pro Thr Gln His Gly Ser Val Pro Ala Ser Pro Thr65 70 75 80Arg Gly Ser Thr Ala Pro Ala Gly Ile Ile Pro Val Asn Ser Val Gly
85 90 95Tyr Glu Val Phe Leu Leu Leu Leu Gln Phe Leu Tyr Ser Gly Gln Val
100 105 110Ser Ile Val Pro Gln Lys His Glu Pro Arg Pro Asn Cys Gly Glu Arg
115 120 125Gly Cys Trp His Thr His Cys Ser Ala Ala Val Asp Leu Ala Leu Asp
130 135 140Thr Leu Ala Ala Ser Arg Tyr Phe Gly Val Glu Gln Leu Ala Leu Leu145 150 155 160Thr Gln Lys Gln Leu Ala Ser Met Val Glu Lys Ala Ser Ile Glu Asp
165 170 175Val Met Lys Val Leu Ile Ala Ser Arg Lys Gln Asp Met His Gln Leu
180 185 190Trp Thr Thr Cys Ser His Leu Val Met Ser Ash Leu Glu Glu Ser Leu
195 200 205Arg Ser Leu Ser Leu Asp Phe Leu Asn Leu Leu Ile Asn Gly Gln Ala
210 215 220Phe Ser Asp Val Thr Phe Ser Val Glu Gly Arg Leu Val His Ala His225 230 235 240Arg Cys Ile Leu Ala Ala Arg Ser Leu Phe Phe Arg Lys Phe Phe Cys
245 250 255Gly Thr Asp Ser Pro Gln Pro Val Thr Gly Ile Asp Pro Thr Gln His
260 265 270Gly Ser Val Pro Ala Ser Pro Thr Arg Gly Ser Thr Ala Pro Ala Gly
275 280 285Ile Ile Pro Val Asn Ser Val Gly Tyr Glu Val Phe Leu Leu Leu Leu
290 295 300Gln Phe Leu Tyr Ser Gly Gln Val Ser Ile Val Pro Gln Lys His Glu305 310 315 320Pro Arg Pro Asn Cys Gly Glu Arg Gly Cys Trp His Thr His Cys Ser
325 330 335Ala Ala Val Asp Leu Ala Leu Asp Thr Leu Ala Ala Ser Arg Tyr Phe
340 345 350Gly Val Glu Gln Leu Ala Leu Leu Thr Gln Lys Gln Leu Ala Ser Met
355 360 365Val Glu Lys Ala Ser Ile Glu Asp Val Met Lys Val Leu Ile Ala Ser
370 375 380Arg Lys Gln Asp Met His Gln Leu Trp Thr Thr Cys Ser His Leu Val385 390 395 400Met Ser Asn Leu Glu Glu Ser Leu Arg Ser Leu Ser Leu Asp Phe Leu
405 410 415Asn Leu Leu Ile Asn Gly Gln Ala Phe Ser Asp Val Thr Phe Ser Val
420 425 430Glu Gly Arg Leu Val His Ala His Arg Cys Ile Leu Ala Ala Arg Ser
435 440 445Leu Phe Phe Arg Lys Phe Phe Cys Gly Thr Asp Ser Pro Gln Pro Val
450 455 460Thr Gly Ile Asp Pro Thr Gln His Gly Ser Val Pro Ala Ser Pro Thr465 470 475 480Arg Gly Ser Thr Ala Pro Ala Gly Ile Ile Pro Val
485 490<210>17<211>1804<212>DNA<213>鼠耳芥<220><221>CDS<222>(1)..(1803)<223>AtNMLc 4-1基因组序列<400>17atg gct gca act gca ata gag cca tct tca tct ata agt ttc aca tct 48Met Ala Ala Thr Ala Ile Glu Pro Ser Ser Ser Ile Ser Phe Thr Ser1 5 10 15tct cac tta tca aac cct tct cct gtt gtt act act tat cac tca gct 96Ser His Leu Ser Asn Pro Ser Pro Val Val Thr Thr Tyr His Ser Ala
20 25 30gct aat ctt gaa gag ctc agc tct aac ttg gag cag ctt ctc act aat 144Ala Asn Leu Glu Glu Leu Ser Ser Asn Leu Glu Gln Leu Leu Thr Asn
35 40 45cca gat tgc gat tac act gac gca gag atc atc att gaa gaa gaa gct 192Pro Asp Cys Asp Tyr Thr Asp Ala Glu Ile Ile Ile Glu Glu Glu Ala
50 55 60aac cct gtg agt gtt cat aga tgt gtt tta gct gct agg agc aag ttt 240Asn Pro Val Ser Val His Arg Cys Val Leu Ala Ala Arg Ser Lys Phe65 70 75 80ttt ctt gat ctg ttt aag aaa gat aaa gat agt agt gag aag aaa cct 288Phe Leu Asp Leu Phe Lys Lys Asp Lys Asp Ser Ser Glu Lys Lys Pro
85 90 95aag tat caa atg aaa gat tta tta cca tat gga aat gtg gga cgt gag 336Lys Tyr Gln Met Lys Asp Leu Leu Pro Tyr Gly Asn Val Gly Arg Glu
100 105 110gca ttt ctg cat ttc ttg agc tat atc tac act ggg agg tta aag cct 384Ala Phe Leu His Phe Leu Ser Tyr Ile Tyr Thr Gly Arg Leu Lys Pro
115 120 125ttt cct atc gag gtt tca act tgt gtt gat tca gtt tgt gct cat gat 432Phe Pro Ile Glu Val Ser Thr Cys Val Asp Ser Val Cys Ala His Asp
130 135 140tct tgt aaa ccg gcc att gat ttt gct gtt gag ttg atg tat gct tca 480Ser Cys Lys Pro Ala Ile Asp Phe Ala Val Glu Leu Met Tyr Ala Ser145 150 155 160ttt gtg ttc caa atc ccg gat ctt gtt tcg tca ttt cag cgg aag ctt 528Phe Val Phe Gln Ile Pro Asp Leu Val Ser Ser Phe Gln Arg Lys Leu
165 170 175cgt aac tat gtt gag aag tca cta gta gag aat gtt ctt cct atc ctc 576Arg Asn Tyr Val Glu Lys Ser Leu Val Glu Asn Val Leu Pro Ile Leu
180 185 190tta gtt gcg ttt cat tgt gat ttg aca cag ctt ctt gat caa tgc att 624Leu Val Ala Phe His Cys Asp Leu Thr Gln Leu Leu Asp Gln Cys Ile
195 200 205gag aga gtg gcg aga tca gac tta gac aga ttc tgt atc gaa aag gag 672Glu Arg Val Ala Arg Ser Asp Leu Asp Arg Phe Cys Ile Glu Lys Glu
210 215 220ctt cct tta gaa gta ttg gaa aaa atc aaa cag ctt cga gtt aag tcg 720Leu Pro Leu Glu Val Leu Glu Lys Ile Lys Gln Leu Arg Val Lys Ser225 230 235 240gtg aac ata ccc gag gtg gag gat aaa tcg ata gag aga aca ggg aaa 768Val Asn Ile Pro Glu Val Glu Asp Lys Ser Ile Glu Arg Thr Gly Lys
245 250 255gta ctc aag gca ttg gat tca gat gat gta gaa ctc gtg aag ctt ctt 816Val Leu Lys Ala Leu Asp Ser Asp Asp Val Glu Leu Val Lys Leu Leu
260 265 270ttg act gag tca gat ata act cta gac caa gcc aat ggt cta cat tat 864Leu Thr Glu Ser Asp Ile Thr Leu Asp Gln Ala Asn Gly Leu His Tyr
275 280 285gca gtg gca tac agt gat ccg aaa gtt gtg aca cag gtt ctt gat cta 912Ala Val Ala Tyr Ser Asp Pro Lys Val Val Thr Gln Val Leu Asp Leu
290 295 300gat atg gct gat gtt aat ttc aga aat tcc agg ggg tat acg gtt ctt 960Asp Met Ala Asp Val Asn Phe Arg Asn Ser Arg Gly Tyr Thr Val Leu305 310 315 320cat att gct gct atg cgt aga gag cca aca att atc ata cca ctt att 1008His Ile Ala Ala Met Arg Arg Glu Pro Thr Ile Ile Ile Pro Leu Ile
325 330 335caa aaa gga gct aat gct tca gat ttc acg ttt gat gga cgc agt gcg 1056Gln Lys Gly Ala Asn Ala Ser Asp Phe Thr Phe Asp Gly Arg Ser Ala
340 345 350gta aat ata tgt agg aga ctc act agg ccg aaa gat tat cat acc aaa 1104Val Asn Ile Cys Arg Arg Leu Thr Arg Pro Lys Asp Tyr His Thr Lys
355 360 365acc tca agg aaa gaa cct agt aaa tac cgc tta tgc atc gat atc ttg 1152Thr Ser Arg Lys Glu Pro Ser Lys Tyr Arg Leu Cys Ile Asp Ile Leu
370 375 380gaa agg gaa att aga agg aat cca ttg gtt agt ggg gat aca ccc act 1200Glu Arg Glu Ile Arg Arg Asn Pro Leu Val Ser Gly Asp Thr Pro Thr385 390 395 400tgt tcc cat tcg atg ccc gag gat ctc caa atg agg ttg tta tac tta 1248Cys Ser His Ser Met Pro Glu Asp Leu Gln Met Arg Leu Leu Tyr Leu
405 410 415gaa aag cga tgg gac ttg cgt cag ttg ttc ttc cca gca gaa gcc aat 1296Glu Lys Arg Trp Asp Leu Arg Gln Leu Phe Phe Pro Ala Glu Ala Asn
420 425 430gtg gct atg gac gtt gct aat gtt gaa ggg aca agc gag tgc aca ggt 1344Val Ala Met Asp Val Ala Asn Val Glu Gly Thr Ser Glu Cys Thr Gly
435 440 445ctt cta act cca cct cca tca aat gat aca act gaa aac ttg ggt aaa 1392Leu Leu Thr Pro Pro Pro Ser Asn Asp Thr Thr Glu Asn Leu Gly Lys
450 455 460gtc gat tta aat gaa acg cct tat gtg caa acg aaa aga atg ctt aca 1440Val Asp Leu Asn Glu Thr Pro Tyr Val Gln Thr Lys Arg Met Leu Thr465 470 475 480cgt atg aaa gcc ctc atg aaa aca ggt aaa agc tta agg aaa tgt act 1488Arg Met Lys Ala Leu Met Lys Thr Gly Lys Ser Leu Arg Lys Cys Thr
485 490 495ttc aag ttt tat tct ctg acc aca aga ttg act gat tcg aaa ccg ttc 1536Phe Lys Phe Tyr Ser Leu Thr Thr Arg Leu Thr Asp Ser Lys Pro Phe
500 505 510aac aac gca gtt gag aca ggt cgg aga tac ttc cca tct tgt tat gag 1584Asn Asn Ala Val Glu Thr Gly Arg Arg Tyr Phe Pro Ser Cys Tyr Glu
515 520 525gtt ctg gat aag tac atg gat cag tat atg gac gaa gaa atc cct gat 1632Val Leu Asp Lys Tyr Met Asp Gln Tyr Met Asp Glu Glu Ile Pro Asp
530 535 540atg tcg tat ccc gag aaa ggc act gtg aaa gag aga aga cag aag agg 1680Met Ser Tyr Pro Glu Lys Gly Thr Val Lys Glu Arg Arg Gln Lys Arg545 550 555 560atg aga tat aac gag ctg aag aac gac gtt aaa aaa gca tat agc aaa 1728Met Arg Tyr Asn Glu Leu Lys Asn Asp Val Lys Lys Ala Tyr Ser Lys
565 570 575gac aaa gtc gcg cgg tct tgt ctt tct tct tca tca cca gct tct tct 1776Asp Lys Val Ala Arg Ser Cys Leu Ser Ser Ser Ser Pro Ala Ser Ser
580 585 590ctt aga gaa gcc tta gag aat cca aca t 1804Leu Arg Glu Ala Leu Glu Asn Pro Thr
595 600<210>18<211>601<212>PRT<213>鼠耳芥<400>18Met Ala Ala Thr Ala Ile Glu Pro Ser Ser Ser Ile Ser Phe Thr Ser1 5 10 15Ser His Leu Ser Asn Pro Ser Pro Val Val Thr Thr Tyr His Ser Ala
20 25 30Ala Asn Leu Glu Glu Leu Ser Ser Asn Leu Glu Gln Leu Leu Thr Asn
35 40 45Pro Asp Cys Asp Tyr Thr Asp Ala Glu Ile Ile Ile Glu Glu Glu Ala
50 55 60Asn Pro Val Ser Val His Arg Cys Val Leu Ala Ala Arg Ser Lys Phe65 70 75 80Phe Leu Asp Leu Phe Lys Lys Asp Lys Asp Ser Ser Glu Lys Lys Pro
85 90 95Lys Tyr Gln Met Lys Asp Leu Leu Pro Tyr Gly Asn Val Gly Arg Glu
100 105 110Ala Phe Leu His Phe Leu Ser Tyr Ile Tyr Thr Gly Arg Leu Lys Pro
115 120 125Phe Pro Ile Glu Val Ser Thr Cys Val Asp Ser Val Cys Ala His Asp
130 135 140Ser Cys Lys Pro Ala Ile Asp Phe Ala Val Glu Leu Met Tyr Ala Ser145 150 155 160Phe Val Phe Gln Ile Pro Asp Leu Val Ser Ser Phe Gln Arg Lys Leu
165 170 175Arg Asn Tyr Val Glu Lys Ser Leu Val Glu Asn Val Leu Pro Ile Leu
180 185 190Leu Val Ala Phe His Cys Asp Leu Thr Gln Leu Leu Asp Gln Cys Ile
195 200 205Glu Arg Val Ala Arg Ser Asp Leu Asp Arg Phe Cys Ile Glu Lys Glu
210 215 220Leu Pro Leu Glu Val Leu Glu Lys Ile Lys Gln Leu Arg Val Lys Ser225 230 235 240Val Asn Ile Pro Glu Val Glu Asp Lys Ser Ile Glu Arg Thr Gly Lys
245 250 255Val Leu Lys Ala Leu Asp Ser Asp Asp Val Glu Leu Val Lys Leu Leu
260 265 270Leu Thr Glu Ser Asp Ile Thr Leu Asp Gln Ala Asn Gly Leu His Tyr
275 280 285Ala Val Ala Tyr Ser Asp Pro Lys Val Val Thr Gln Val Leu Asp Leu
290 295 300Asp Met Ala Asp Val Asn Phe Arg Asn Ser Arg Gly Tyr Thr Val Leu305 310 315 320His Ile Ala Ala Met Arg Arg Glu Pro Thr Ile Ile Ile Pro Leu Ile
325 330 335Gln Lys Gly Ala Asn Ala Ser Asp Phe Thr Phe Asp Gly Arg Ser Ala
340 345 350Val Asn Ile Cys Arg Arg Leu Thr Arg Pro Lys Asp Tyr His Thr Lys
355 360 365Thr Ser Arg Lys Glu Pro Ser Lys Tyr Arg Leu Cys Ile Asp Ile Leu
370 375 380Glu Arg Glu Ile Arg Arg Asn Pro Leu Val Ser Gly Asp Thr Pro Thr385 390 395 400Cys Ser His Ser Met Pro Glu Asp Leu Gln Met Arg Leu Leu Tyr Leu
405 410 415Glu Lys Arg Trp Asp Leu Arg Gln Leu Phe Phe Pro Ala Glu Ala Asn
420 425 430Val Ala Met Asp Val Ala Asn Val Glu Gly Thr Ser Glu Cys Thr Gly
435 440 445Leu Leu Thr Pro Pro Pro Ser Asn Asp Thr Thr Glu Asn Leu Gly Lys
450 455 460Val Asp Leu Asn Glu Thr Pro Tyr Val Gln Thr Lys Arg Met Leu Thr465 470 475 480Arg Met Lys Ala Leu Met Lys Thr Gly Lys Ser Leu Arg Lys Cys Thr
485 490 495Phe Lys Phe Tyr Ser Leu Thr Thr Arg Leu Thr Asp Ser Lys Pro Phe
500 505 510Asn Asn Ala Val Glu Thr Gly Arg Arg Tyr Phe Pro Ser Cys Tyr Glu
515 520 525Val Leu Asp Lys Tyr Met Asp Gln Tyr Met Asp Glu Glu Ile Pro Asp
530 535 540Met Ser Tyr Pro Glu Lys Gly Thr Val Lys Glu Arg Arg Gln Lys Arg545 550 555 560Met Arg Tyr Asn Glu Leu Lys Asn Asp Val Lys Lys Ala Tyr Ser Lys
565 570 575Asp Lys Val Ala Arg Ser Cys Leu Ser Ser Ser Ser Pro Ala Ser Ser
580 585 590Leu Arg Glu Ala Leu Glu Asn Pro Thr
595 600<210>19<211>1803<212>DNA<213>鼠耳芥<220><221>CDS<222>(1)..(1803)<223>AtNMLc4-2基因组序列<400>19atg gcc acc acc acc acc acc acc acc gct aga ttc tct gat tca tac 48Met Ala Thr Thr Thr Thr Thr Thr Thr Ala Arg Phe Ser Asp Ser Tyr1 5 10 15gag ttc agc aac aca agc ggc aat agc ttc ttc gcc gcc gag tca tct 96Glu Phe Ser Asn Thr Ser Gly Asn Ser Phe Phe Ala Ala Glu Ser Ser
20 25 30ctt gat tat ccg acg gaa ttt ctc acg cca ccg gag gta tca gct ctt 144Leu Asp Tyr Pro Thr Glu Phe Leu Thr Pro Pro Glu Val Ser Ala Leu
35 40 45aaa ctt ctg tct aac tgc ctc gag tct gtt ttc gac tcg ccg gag acg 192Lys Leu Leu Ser Asn Cys Leu Glu Ser Val Phe Asp Ser Pro Glu Thr
50 55 60ttc tac agc gat gct aag cta gtt ctc gcc ggc ggc cgg gaa gtt tct 240Phe Tyr Ser Asp Ala Lys Leu Val Leu Ala Gly Gly Arg Glu Val Ser65 70 75 80ttt cac cgt tgt att ctt tcc gcg aga att cct gtc ttc aaa agc gct 288Phe His Arg Cys Ile Leu Ser Ala Arg Ile Pro Val Phe Lys Ser Ala
85 90 95tta gcc acc gtg aag gaa caa aaa tcc tcc acc acc gtg aag ctc cag 336Leu Ala Thr Val Lys Glu Gln Lys Ser Ser Thr Thr Val Lys Leu Gln
100 105 110ctg aaa gag atc gcc aga gat tac gaa gtc ggc ttt gac tcg gtt gtg 384Leu Lys Glu Ile Ala Arg Asp Tyr Glu Val Gly Phe Asp Ser Val Val
115 120 125gcg gtt ttg gcg tat gtt tac agc ggc aga gtg agg tcc ccg ccg aag 432Ala Val Leu Ala Tyr Val Tyr Ser Gly Arg Val Arg Ser Pro Pro Lys
130 135 140gga gct tct gct tgc gta gac gac gat tgt tgc cac gtg gct tgc cgg 480Gly Ala Ser Ala Cys Val Asp Asp Asp Cys Cys His Val Ala Cys Arg145 150 155 160tca aag gtg gat ttc atg gtg gag gtt ctt tat ctg tct ttc gtt ttc 528Ser Lys Val Asp Phe Met Val Glu Val Leu Tyr Leu Ser Phe Val Phe
165 170 175cag att caa gaa tta gtt act ctg tat gag agg cag ttc ttg gaa att 576Gln Ile Gln Glu Leu Val Thr Leu Tyr Glu Arg Gln Phe Leu Glu Ile
180 185 190gta gac aaa gtt gta gtc gaa gac atc ttg gtt ata ttc aag ctt gat 624Val Asp Lys Val Val Val Glu Asp Ile Leu Val Ile Phe Lys Leu Asp
195 200 205act cta tgt ggt aca aca tac aag aag ctt ttg gat aga tgc ata gaa 672Thr Leu Cys Gly Thr Thr Tyr Lys Lys Leu Leu Asp Arg Cys Ile Glu
210 215 220att atc gtg aag tct gat ata gaa cta gtt agt ctt gag aag tct tta 720Ile Ile Val Lys Ser Asp Ile Glu Leu Val Ser Leu Glu Lys Ser Leu225 230 235 240cct caa cac att ttc aag caa atc ata gac atc cgc gaa gcg ctc tgt 768Pro Gln His Ile Phe Lys Gln Ile Ile Asp Ile Arg Glu Ala Leu Cys
245 250 255cta gag cca cct aaa cta gaa agg cat gtc aag aac ata tac aag gcg 816Leu Glu Pro Pro Lys Leu Glu Arg His Val Lys Asn Ile Tyr Lys Ala
260 265 270cta gac tca gat gat gtt gag ctt gtc aag atg ctt ttg cta gaa gga 864Leu Asp Ser Asp Asp Val Glu Leu Val Lys Met Leu Leu Leu Glu Gly
275 280 285cac acc aat ctc gat gag gcg tat gct ctt cat ttt gct atc gct cac 912His Thr Asn Leu Asp Glu Ala Tyr Ala Leu His Phe Ala Ile Ala His
290 295 300tgc gct gtg aag acc gcg tat gat ctc ctc gag ctt gag ctt gcg gat 960Cys Ala Val Lys Thr Ala Tyr Asp Leu Leu Glu Leu Glu Leu Ala Asp305 310 315 320gtt aac ctt aga aat ecg agg gga tac act gtg ctt cat gtt gct gcg 1008Val Asn Leu Arg Asn Pro Arg Gly Tyr Thr Val Leu His Val Ala Ala
325 330 335atg cgg aag gag ccg aag ttg ata ata tct ttg tta atg aaa ggg gca 1056Met Arg Lys Glu Pro Lys Leu Ile Ile Ser Leu Leu Met Lys Gly Ala
340 345 350aat att tta gac aca aca ttg gat ggt aga acc gct tta gtg att gta 1104Asn Ile Leu Asp Thr Thr Leu Asp Gly Arg Thr Ala Leu Val Ile Val
355 360 365aaa cga ctc act aaa gcg gat gac tac aaa act agt acg gag gac ggt 1152Lys Arg Leu Thr Lys Ala Asp Asp Tyr Lys Thr Ser Thr Glu Asp Gly
370 375 380acg cct tct ctg aaa ggc gga tta tgc ata gag gta ctt gag cat gaa 1200Thr Pro Ser Leu Lys Gly Gly Leu Cys Ile Glu Val Leu Glu His Glu385 390 395 400caa aaa cta gaa tat ttg tcg cct ata gag gct tca ctt tct ctt cca 1248Gln Lys Leu Glu Tyr Leu Ser Pro Ile Glu Ala Ser Leu Ser Leu Pro
405 410 415gta act cca gag Gag ttg agg atg agg ttg ctc tat tat gaa aac cga 1296Val Thr Pro Glu Glu Leu Arg Met Arg Leu Leu Tyr Tyr Glu Asn Arg
420 425 430gtt gca ctt gct cga ctt ctc ttt cca gtg gaa act gaa act gta cag 1344Val Ala Leu Ala Arg Leu Leu Phe Pro Val Glu Thr Glu Thr Val Gln
435 440 445ggt att gcc aaa ttg gag gaa aca tgc gag ttt aca gct tct agt ctc 1392Gly Ile Ala Lys Leu Glu Glu Thr Cys Glu Phe Thr Ala Ser Ser Leu
450 455 460gag cct gat cat cac att ggt gaa aag cgg aca tca cta gac cta aat 1440Glu Pro Asp His His Ile Gly Glu Lys Arg Thr Ser Leu Asp Leu Asn465 470 475 480atg gcg ccg ttc caa atc cat gag aag cat ttg agt aga cta aga gca 1488Met Ala Pro Phe Gln Ile His Glu Lys His Leu Ser Arg Leu Arg Ala
485 490 495ctt tgt aaa acc gtg gaa ctg ggg aaa cgc tac ttc aaa cga tgt tcg 1536Leu Cys Lys Thr Val Glu Leu Gly Lys Arg Tyr Phe Lys Arg Cys Ser
500 505 510ctt gat cac ttt atg gat act gag gac ttg aat cat ctt gct agc gta 1584Leu Asp His Phe Met Asp Thr Glu Asp Leu Asn His Leu Ala Ser Val
515 520 525gaa gaa gat act cct gag aaa cgg cta caa aag aag caa agg tac atg 1632Glu Glu Asp Thr Pro Glu Lys Arg Leu Gln Lys Lys Gln Arg Tyr Met
530 535 540gaa cta caa gag act ctg atg aag acc ttt agt gag gac aag gag gaa 1680Glu Leu Gln Glu Thr Leu Met Lys Thr Phe Ser Glu Asp Lys Glu Glu545 550 555 560tgt gga aag tct tcc aca ccg aaa cca acc tct gcg gtg agg tct aat 1728Cys Gly Lys Ser Ser Thr Pro Lys Pro Thr Ser Ala Val Arg Ser Asn
565 570 575aga aaa ctc tct cac cgg cgc cta aaa gtg gac aaa cgg gat ttt ttg 1776Arg Lys Leu Ser His Arg Arg Leu Lys Val Asp Lys Arg Asp Phe Leu
580 585 590aaa cga cct tac ggg aac ggg gat taa 1803Lys Arg Pro Tyr Gly Asn Gly Asp
595 600<210>20<211>600<212>PRT<213>鼠耳芥<400>20Met Ala Thr Thr Thr Thr Thr Thr Thr Ala Arg Phe Ser Asp Ser Tyr 1 5 10 15Glu Phe Ser Asn Thr Ser Gly Asn Ser Phe Phe Ala Ala Glu Ser Ser
20 25 30Leu Asp Tyr Pro Thr Glu Phe Leu Thr Pro Pro Glu Val Ser Ala Leu
35 40 45Lys Leu Leu Ser Asn Cys Leu Glu Ser Val Phe Asp Ser Pro Glu Thr
50 55 60Phe Tyr Ser Asp Ala Lys Leu Val Leu Ala Gly Gly Arg Glu Val Ser65 70 75 80Phe His Arg Cys Ile Leu Ser Ala Arg Ile Pro Val Phe Lys Ser Ala
85 90 95Leu Ala Thr Val Lys Glu Gln Lys Ser Ser Thr Thr Val Lys Leu Gln
100 105 110Leu Lys Glu Ile Ala Arg Asp Tyr Glu Val Gly Phe Asp Ser Val Val
115 120 125Ala Val Leu Ala Tyr Val Tyr Ser Gly Arg Val Arg Ser Pro Pro Lys
130 135 140Gly Ala Ser Ala Cys Val Asp Asp Asp Cys Cys His Val Ala Cys Arg145 150 155 160Ser Lys Val Asp Phe Met Val Glu Val Leu Tyr Leu Ser Phe Val Phe
165 170 175Gln Ile Gln Glu Leu Val Thr Leu Tyr Glu Arg Gln Phe Leu Glu Ile
180 185 190Val Asp Lys Val Val Val Glu Asp Ile Leu Val Ile Phe Lys Leu Asp
195 200 205Thr Leu Cys Gly Thr Thr Tyr Lys Lys Leu Leu Asp Arg Cys Ile Glu
210 215 220Ile Ile Val Lys Ser Asp Ile Glu Leu Val Ser Leu Glu Lys Ser Leu225 230 235 240Pro Gln His Ile Phe Lys Gln Ile Ile Asp Ile Arg Glu Ala Leu Cys
245 250 255Leu Glu Pro Pro Lys Leu Glu Arg His Val Lys Asn Ile Tyr Lys Ala
260 265 270Leu Asp Ser Asp Asp Val Glu Leu Val Lys Met Leu Leu Leu Glu Gly
275 280 285His Thr Asn Leu Asp Glu Ala Tyr Ala Leu His Phe Ala Ile Ala His
290 295 300Cys Ala Val Lys Thr Ala Tyr Asp Leu Leu Glu Leu Glu Leu Ala Asp305 310 315 320Val Asn Leu Arg Asn Pro Arg Gly Tyr Thr Val Leu His Val Ala Ala
325 330 335Met Arg Lys Glu Pro Lys Leu Ile Ile Ser Leu Leu Met Lys Gly Ala
340 345 350Asn Ile Leu Asp Thr Thr Leu Asp Gly Arg Thr Ala Leu Val Ile Val
355 360 365Lys Arg Leu Thr Lys Ala Asp Asp Tyr Lys Thr Ser Thr Glu Asp Gly
370 375 380Thr Pro Ser Leu Lys Gly Gly Leu Cys Ile Glu Val Leu Glu His Glu385 390 395 400Gln Lys Leu Glu Tyr Leu Ser Pro Ile Glu Ala Ser Leu Ser Leu Pro
405 410 415Val Thr Pro Glu Glu Leu Arg Met Arg Leu Leu Tyr Tyr Glu Asn Arg
420 425 430Val Ala Leu Ala Arg Leu Leu Phe Pro Val Glu Thr Glu Thr Val Gln
435 440 445Gly Ile Ala Lys Leu Glu Glu Thr Cys Glu Phe Thr Ala Ser Ser Leu
450 455 460Glu Pro Asp His His Ile Gly Glu Lys Arg Thr Ser Leu Asp Leu Asn465 470 475 480Met Ala Pro Phe Gln Ile His Glu Lys His Leu Ser Arg Leu Arg Ala
485 490 495Leu Cys Lys Thr Val Glu Leu Gly Lys Arg Tyr Phe Lys Arg Cys Ser
500 505 510Leu Asp His Phe Met Asp Thr Glu Asp Leu Asn His Leu Ala Ser Val
515 520 525Glu Glu Asp Thr Pro Glu Lys Arg Leu Gln Lys Lys Gln Arg Tyr Met
530 535 540Glu Leu Gln Glu Thr Leu Met Lys Thr Phe Ser Glu Asp Lys Glu Glu545 550 555 560Cys Gly Lys Ser Ser Thr Pro Lys Pro Thr Ser Ala Val Arg Ser Asn
565 570 575Arg Lys Leu Ser His Arg Arg Leu Lys Val Asp Lys Arg Asp Phe Leu
580 585 590Lys Arg Pro Tyr Gly Asn Gly Asp
595 600<210>21<211>28<212>DNA<213>人工序列<220><223>人工序列描述:PCR引物NIM1A<400>21gakattattg tcaagtctaa tgtwgata 28<210>22<211>25<212>DNA<213>人工序列<220><223>人工序列描述:PCR引物NIM1B<400>22aytkgaytck gatgatrttg artta 25<210>23<211>27<212>DNA<213>人工序列<220><223>人工序列描述:PCR引物NIM1C<400>23taaytcaaya tcatcmgart cmartgc 27<210>24<211>28<212>DNA<213>人工序列<223>人工序列描述:PCR引物NIM1D<220><221>misc_特征<222>(1)..(28)<223>n=a、t、c或g<400>24gttkagcmag nscaactcta ttttcaag 28<210>25<211>32<212>DNA<213>人工序列<220><223>人工序列描述:PCR引物NIM2A<400>25tgcatwgara twrttgtsaa gtctratgtw ga 32<210>26<211>27<212>DNA<213>人工序列<220><223>人工序列描述:PCR引物NIM2B<400>26ggcaytggay tcwgatgatg ttgaryt 27<210>27<211>27<212>DNA<213>人工序列<220><223>人工序列描述:PCR引物NIM2C<400>27arytcaacat catcwgartc cartgcc 27<210>28<211>31<212>DNA<213>人工序列<220><223>人工序列描述:PCR引物NIM2D<400>28agttkagcma gdccaactck attttcaarrt 31<210>29<211>659<212>DNA<213>烟草<220><221>CDS<222>(1)..(657)<223>烟草A<400>29tgc atg gag att att gtc aag tct aat gtt gat atc ata acc ctt gat 48Cys Met Glu Ile Ile Val Lys Ser Asn Val Asp Ile Ile Thr Leu Asp1 5 10 15aag gcc ttg cct cat gac att gta aaa caa att acc gat tca cga gca 96Lys Ala Leu Pro His Asp Ile Val Lys Gln Ile Thr Asp Ser Arg Ala
20 25 30gaa ctt ggt cta caa ggg cct gaa agc aat ggt ttt cct gat aaa cat 144Glu Leu Gly Leu Gln Gly Pro Glu Ser Asn Gly Phe Pro Asp Lys His
35 40 45gtt aag agg ata cat agg gca tta gat tct gat gat gtt gaa tta ctg 192Val Lys Arg Ile His Arg Ala Leu Asp Ser Asp Asp Val Glu Leu Leu
50 55 60cag atg ttg cta aga gag ggg cat act act cta gat gat gca tat gct 240Gln Met Leu Leu Arg Glu Gly His Thr Thr Leu Asp Asp Ala Tyr Ala65 70 75 80ctc cac tat gct gta gca tat tgc gat gca aag act aca gca gaa ctt 288Leu His Tyr Ala Val Ala Tyr Cys Asp Ala Lys Thr Thr Ala Glu Leu
85 90 95cta gat ctt gca ctt gct gat gtt aat cat caa aat tca aga gga tac 336Leu Asp Leu Ala Leu Ala Asp Val Asn His Gln Asn Ser Arg Gly Tyr
100 105 110aca gtg ctg cat gtt gca gcc atg agg aaa gag cct aaa att ata gtg 384Thr Val Leu His Val Ala Ala Met Arg Lys Glu Pro Lys Ile Ile Val
115 120 125tcc ctt tta acc aaa gga gct aga cct tct gat ctg aca tcc gat ggc 432Ser Leu Leu Thr Lys Gly Ala Arg Pro Ser Asp Leu Thr Ser Asp Gly
130 135 140aga aaa gca ctt caa att gcc aag agg ctc act agg ctt gtg gat ttc 480Arg Lys Ala Leu Gln Ile Ala Lys Arg Leu Thr Arg Leu Val Asp Phe145 150 155 160agt aag tct cca gag gaa gga aaa tct gct tcg aag gat cgg tta tgc 528Ser Lys Ser Pro Glu Glu Gly Lys Ser Ala Ser Lys Asp Arg Leu Cys
165 170 175att gag att ctg gag caa gca gaa aga aga gat cca ctg cta gga gaa 576Ile Glu Ile Leu Glu Gln Ala Glu Arg Arg Asp Pro Leu Leu Gly Glu
180 185 190gct tct gta tct ctt gct atg gcg ggc gat gat ttg cgt atg aag ctg 624Ala Ser Val Ser Leu Ala Met Ala Gly Asp Asp Leu Arg Met Lys Leu
195 200 205tta tac ctt gaa aat aga gtt ggc ctt gct caa ct 659Leu Tyr Leu Glu Asn Arg Val Gly Leu Ala Gln
210 215<210>30<211>219<212>PRT<213>烟草<400>30Cys Met Glu Ile Ile Val Lys Ser Asn Val Asp Ile Ile Thr Leu Asp1 5 10 15Lys Ala Leu Pro His Asp Ile Val Lys Gln Ile Thr Asp Ser Arg Ala
20 25 30Glu Leu Gly Leu Gln Gly Pro Glu Ser Asn Gly Phe Pro Asp Lys His
35 40 45Val Lys Arg Ile His Arg Ala Leu Asp Ser Asp Asp Val Glu Leu Leu
50 55 60Gln Met Leu Leu Arg Glu Gly His Thr Thr Leu Asp Asp Ala Tyr Ala65 70 75 80Leu His Tyr Ala Val Ala Tyr Cys Asp Ala Lys Thr Thr Ala Glu Leu
85 90 95Leu Asp Leu Ala Leu Ala Asp Val Asn His Gln Asn Ser Arg Gly Tyr
100 105 110Thr Val Leu His Val Ala Ala Met Arg Lys Glu Pro Lys Ile Ile Val
115 120 125Ser Leu Leu Thr Lys Gly Ala Arg Pro Ser Asp Leu Thr Ser Asp Gly
130 135 140Arg Lys Ala Leu Gln Ile Ala Lys Arg Leu Thr Arg Leu Val Asp Phe145 150 155 160Ser Lys Ser Pro Glu Glu Gly Lys Ser Ala Ser Lys Asp Arg Leu Cys
165 170 175Ile Glu Ile Leu Glu Gln Ala Glu Arg Arg Asp Pro Leu Leu Gly Glu
180 185 190Ala Ser Val Ser Leu Ala Met Ala Gly Asp Asp Leu Arg Met Lys Leu
195 200 205Leu Tyr Leu Glu Asn Arg Val Gly Leu Ala Gln
210 215<210>31<211>498<212>DNA<213>烟草<220><221>CDS<222>(2)..(496)<223>烟草B<400>31g gca ctg gat tct gat gat gtt gag ctg gtc aag ctt cta ctc aac gag 49Ala Leu Asp Ser Asp Asp Val Glu Leu Val Lys Leu Leu Leu Asn Glu
1 5 10 15tct gag ata agc tta gat gaa gcc tac gct ctt cat tat gct gtt gca 97Ser Glu Ile Ser Leu Asp Glu Ala Tyr Ala Leu His Tyr Ala Val Ala
20 25 30tat tgt gat ccc aag gtt gtg act gag gtt ctt gga ctg ggt gtt gct 145Tyr Cys Asp Pro Lys Val Val Thr Glu Val Leu Gly Leu Gly Val Ala
35 40 45gat gtc aat cta cgt aat act cgc ggt tac act gtg ctt cac att gct 193Asp Val Asn Leu Arg Asn Thr Arg Gly Tyr Thr Val Leu His Ile Ala
50 55 60gcc atg cgt aag gag cca gca ata att gta tcg ctt ttg act aag gga 241Ala Met Arg Lys Glu Pro Ala Ile Ile Val Ser Leu Leu Thr Lys Gly65 70 75 80gct cat gtg tca gag att aca ttg gat ggg caa agt gct gtt agt atc 289Ala His Val Ser Glu Ile Thr Leu Asp Gly Gln Ser Ala Val Ser Ile
85 90 95tgt agg agg cta act agg cct aag gag tac cat gca aaa aca gaa caa 337Cys Arg Arg Leu Thr Arg Pro Lys Glu Tyr His Ala Lys Thr Glu Gln
100 105 110ggc cag gaa gca aac aaa gat cgg gta tgt att gat gtt ttg gag aga 385Gly Gln Glu Ala Asn Lys Asp Arg Val Cys Ile Asp Val Leu Glu Arg
115 120 125gag atg cgt cgc aac cca atg gct gga gat gca ttg ctt tct tcc caa 433Glu Met Arg Arg Asn Pro Met Ala Gly Asp Ala Leu Leu Ser Ser Gln
130 135 140atg ttg gcc gat gat ctg cac atg aaa ctg cac tat ttt gaa aat cga 481Met Leu Ala Asp Asp Leu His Met Lys Leu His Tyr Phe Glu Asn Arg145 150 155 160gtt gga ctt gct caa ct 498Val Gly Leu Ala Gln
165<210>32<211>165<212>PRT<213>烟草<400>32Ala Leu Asp Ser Asp Asp Val Glu Leu Val Lys Leu Leu Leu Asn Glu1 5 10 15Ser Glu Ile Ser Leu Asp Glu Ala Tyr Ala Leu His Tyr Ala Val Ala
20 25 30Tyr Cys Asp Pro Lys Val Val Thr Glu Val Leu Gly Leu Gly Val Ala
35 40 45Asp Val Asn Leu Arg Asn Thr Arg Gly Tyr Thr Val Leu His Ile Ala
50 55 60Ala Met Arg Lys Glu Pro Ala Ile Ile Val Ser Leu Leu Thr Lys Gly65 70 75 80Ala His Val Ser Glu Ile Thr Leu Asp Gly Gln Ser Ala Val Ser Ile
85 90 95Cys Arg Arg Leu Thr Arg Pro Lys Glu Tyr His Ala Lys Thr Glu Gln
100 105 110Gly Gln Glu Ala Asn Lys Asp Arg Val Cys Ile Asp Val Leu Glu Arg
115 120 125Glu Met Arg Arg Asn Pro Met Ala Gly Asp Ala Leu Leu Ser Ser Gln
130 135 140Met Leu Ala Asp Asp Leu His Met Lys Leu His Tyr Phe Glu Asn Arg145 150 155 160Val Gly Leu Ala Gln
165<210>33<211>498<212>DNA<213>烟草<220><221>CDS<222>(2)..(496)<223>烟草C<400>33g gca ctg gac tcw gat gat gtt gag ttt gtc aag ctt cta ctg agt gag 49Ala Leu Asp Xaa Asp Asp Val Glu Phe Val Lys Leu Leu Leu Ser Glu
1 5 10 15tct aac ata agc tta gat gaa gcc tac gct ctt cat tat gct gtg gca 97Ser Asn Ile Ser Leu Asp Glu Ala Tyr Ala Leu His Tyr Ala Val Ala
20 25 30tat tgt gat ccc aag gtt gtg act gag gtt ctt gga ctg ggt gtt gcg 145TYr Cys Asp Pro Lys Val Val Thr Glu Val Leu Gly Leu Gly Val Ala
35 40 45gat gtc aac cta cgt aat act cgt ggt tac act gtg ctt cac att gct 193Asp Val Asn Leu Arg Asn Thr Arg Gly Tyr Thr Val Leu His Ile Ala
50 55 60tcc atg cgt aag gag cca gca gta att gta tcg ctt ttg act aag gga 241Ser Met Arg Lys Glu Pro Ala Val Ile Val Ser Leu Leu Thr Lys Gly65 70 75 80gct cgt gca tca gag act aca ttg gat ggg cag agt gct gtt agt atc 289Ala Arg Ala Ser Glu Thr Thr Leu Asp Gly Gln Ser Ala Val Ser Ile
85 90 95tgt agg agg ctg act agg cct aag gag tac cat gca aaa aca gaa caa 337Cys Arg Arg Leu Thr Arg Pro Lys Glu Tyr His Ala Lys Thr Glu Gln
100 105 110ggc cag gaa gca aac aaa gat cgg gta tgt att gat gtt ttg gag aga 385Gly Gln Glu Ala Asn Lys Asp Arg Val Cys Ile Asp Val Leu Glu Arg
115 120 125gag atg cgt cgc aac cca atg gct gga gat gca ttg ttt tct tcc cca 433Glu Met Arg Arg Asn Pro Met Ala Gly Asp Ala Leu Phe Ser Ser Pro
130 135 140atg ttg gcc gat gat ctg cac atg aaa ctg cac tac ctt gaa aat aga 481Met Leu Ala Asp Asp Leu His Met Lys Leu His Tyr Leu Glu Asn Arg145 150 155 160gtt ggc ctg gct caa ct 498Val Gly Leu Ala Gln
165<210>34<211>165<212>PRT<213>烟草<400>34Ala Leu Asp Xaa Asp Asp Val Glu Phe Val Lys Leu Leu Leu Ser Glu1 5 10 15Ser Asn Ile Ser Leu Asp Glu Ala Tyr Ala Leu His Tyr Ala Val Ala
20 25 30Tyr Cys Asp Pro Lys Val Val Thr Glu Val Leu Gly Leu Gly Val Ala
35 40 45Asp Val Asn Leu Arg Asn Thr Arg Gly Tyr Thr Val Leu His Ile Ala
50 55 60Ser Met Arg Lys Glu Pro Ala Val Ile Val Ser Leu Leu Thr Lys Gly65 70 75 80Ala Arg Ala Ser Glu Thr Thr Leu Asp Gly Gln Ser Ala Val Ser Ile
85 90 95Cys Arg Arg Leu Thr Arg Pro Lys Glu Tyr His Ala Lys Thr Glu Gln
100 105 110Gly Gln Glu Ala Asn Lys Asp Arg Val Cys Ile Asp Val Leu Glu Arg
115 120 125Glu Met Arg Arg Asn Pro Met Ala Gly Asp Ala Leu Phe Ser Ser Pro
130 135 140Met Leu Ala Asp Asp Leu His Met Lys Leu His Tyr Leu Glu Asn Arg145 150 155 160Val Gly Leu Ala Gln
165<210>35<211>399<212>DNA<213>烟草<220><221>CDS<222>(1)..(399)<223>烟草D<400>35act gat tcg gat gat gtt gag tta ctt aag tta ctt ctt gaa gag tct 48Thr Asp Ser Asp Asp Val Glu Leu Leu Lys Leu Leu Leu Glu Glu Ser1 5 10 15aat gtc act tta gac gat gct tgt gct ctt cat tat gca gct gct tat 96Asn Val Thr Leu Asp Asp Ala Cys Ala Leu His Tyr Ala Ala Ala Tyr
20 25 30tgt aac tcc aag gtt gtg aat gag gtc ctc gag ctg gat tta gct gat 144Cys Asn Ser Lys Val Val Asn Glu Val Leu Glu Leu Asp Leu Ala Asp
35 40 45gtc aat ctt cag aac tcc cga gga tat aac gtc ctt cac gtt gct gct 192Val Asn Leu Gln Asn Ser Arg Gly Tyr Asn Val Leu His Val Ala Ala
50 55 60aga aga aag gag cca tca ata ata atg gga cta ctt gaa aaa gga gca 240Arg Arg Lys Glu Pro Ser Ile Ile Met Gly Leu Leu Glu Lys Gly Ala65 70 75 80tct ttc ttg aat act aca cgg gat gga aac aca gca cta tct atc tgt 288Ser Phe Leu Asn Thr Thr Arg Asp Gly Asn Thr Ala Leu Ser Ile Cys
85 90 95cgg aga ttg act cgg cca aag gat tat aat gag cca aca aag caa ggg 336Arg Arg Leu Thr Arg Pro Lys Asp Tyr Asn Glu Pro Thr Lys Gln Gly
100 105 110aaa gaa act aat aag gac cgc ata tgc att gat att ttg gag aga gag 384Lys Glu Thr Asn Lys Asp Arg Ile Cys Ile Asp Ile Leu Glu Arg Glu
115 120 125acg aat agg aat cct 399Thr Asn Arg Asn Pro
130<210>36<211>133<212>PRT<213>烟草<400>36Thr Asp Ser Asp Asp Val Glu Leu Leu Lys Leu Leu Leu Glu Glu Ser1 5 10 15Asn Val Thr Leu Asp Asp Ala Cys Ala Leu His Tyr Ala Ala Ala Tyr
20 25 30Cys Asn Ser Lys Val Val Asn Glu Val Leu Glu Leu Asp Leu Ala Asp
35 40 45Val Asn Leu Gln Asn Ser Arg Gly Tyr Asn Val Leu His Val Ala Ala
50 55 60Arg Arg Lys Glu Pro Ser Ile Ile Met Gly Leu Leu Glu Lys Gly Ala65 70 75 80Ser Phe Leu Asn Thr Thr Arg Asp Gly Asn Thr Ala Leu Ser Ile Cys
85 90 95Arg Arg Leu Thr Arg Pro Lys Asp Tyr Asn Glu Pro Thr Lys Gln Gly
100 105 110Lys Glu Thr Asn Lys Asp Arg Ile Cys Ile Asp Ile Leu Glu Arg Glu
115 120 125Thr Asn Arg Asn Pro
130<210>37<211>498<212>DNA<213>番茄<220><221>CDS<222>(2)..(496)<223>番茄A<400>37g gca ttg gat tct gat gat gtt gag tta cta agg atg ttg ctt aaa gag 49Ala Leu Asp Ser Asp Asp Val Glu Leu Leu Arg Met Leu Leu Lys Glu
1 5 10 15ggg cat act act ctt gat gat gca tat gct ctc cac tat gct gta gca 97Gly His Thr Thr Leu Asp Asp Ala Tyr Ala Leu His Tyr Ala Val Ala
20 25 30tat tgc gat gca aag act aca gca gaa ctt tta gat ctt tca ctt gct 145Tyr Cys Asp Ala Lys Thr Thr Ala Glu Leu Leu Asp Leu Ser Leu Ala
35 40 45gat gtt aat cat caa aat cct aga gga cac acg gta ctt cat gtt gct 193Asp Val Asn His Gln Asn Pro Arg Gly His Thr Val Leu His Val Ala
50 55 60gcc atg agg aaa gaa cct aaa att ata gtg tcc ctt tta acc aaa gga 241Ala Met Arg Lys Glu Pro Lys Ile Ile Val Ser Leu Leu Thr Lys Gly65 70 75 80gct aga cct tct gat ctg aca tcc gat ggc aaa aaa gca ctt caa att 289Ala Arg Pro Ser Asp Leu Thr Ser Asp Gly Lys Lys Ala Leu Gln Ile
85 90 95gct aag agg ctc act agg ctt gta gat ttt acc aag tct aca gag gaa 337Ala Lys Arg Leu Thr Arg Leu Val Asp Phe Thr Lys Ser Thr Glu Glu
100 105 110gga aaa tct gct cca aag gat cgg tta tgc att gag att ctg gag caa 385Gly Lys Ser Ala Pro Lys Asp Arg Leu Cys Ile Glu Ile Leu Glu Gln
115 120 125gca gaa aga aga gat cca cta cta gga gaa gct tca tta tct ctt gct 433Ala Glu Arg Arg Asp Pro Leu Leu Gly Glu Ala Ser Leu Ser Leu Ala
130 135 140atg gca ggc gat gat ttg cgt atg aag ctg tta tac ctt gaa aat aga 481Met Ala Gly Asp Asp Leu Arg Met Lys Leu Leu Tyr Leu Glu Asn Arg145 150 155 160gtt ggc ctt gct aaa ct 498Val Gly Leu Ala Lys
165<210>38<211>165<212>PRT<213>番茄<400>38Ala Leu Asp Ser Asp Asp Val Glu Leu Leu Arg Met Leu Leu Lys Glu1 5 10 15Gly His Thr Thr Leu Asp Asp Ala Tyr Ala Leu His Tyr Ala Val Ala
20 25 30Tyr Cys Asp Ala Lys Thr Thr Ala Glu Leu Leu Asp Leu Ser Leu Ala
35 40 45Asp Val Asn His Gln Asn Pro Arg Gly His Thr Val Leu His Val Ala
50 55 60Ala Met Arg Lys Glu Pro Lys Ile Ile Val Ser Leu Leu Thr Lys Gly65 70 75 80Ala Arg Pro Ser Asp Leu Thr Ser Asp Gly Lys Lys Ala Leu Gln Ile
85 90 95Ala Lys Arg Leu Thr Arg Leu Val Asp Phe Thr Lys Ser Thr Glu Glu
100 105 110Gly Lys Ser Ala Pro Lys Asp Arg Leu Cys Ile Glu Ile Leu Glu Gln
115 120 125Ala Glu Arg Arg Asp Pro Leu Leu Gly Glu Ala Ser Leu Ser Leu Ala
130 135 140Met Ala Gly Asp Asp Leu Arg Met Lys Leu Leu Tyr Leu Glu Asn Arg145 150 155 160Val Gly Leu Ala Lys
165<210>39<211>498<212>DNA<213>甜菜<220><221>CDS<222>(2)..(496)<223>甜菜<400>39g gca ttg gat tct gat gat gtt gag tta gtc aga atg ctt tta aaa gag 49Ala Leu Asp Ser Asp Asp Val Glu Leu Val Arg Met Leu Leu Lys Glu
1 5 10 15cgc cat aca act cta gat gat gca tat gcc ctt cac tat gct gtg gca 97Arg His Thr Thr Leu Asp Asp Ala Tyr Ala Leu His Tyr Ala Val Ala
20 25 30cat tgt gat gcc aag acc acc acg gag ctt ctt gag ctt ggg ctt gca 145His Cys Asp Ala Lys Thr Thr Thr Glu Leu Leu Glu Leu Gly Leu Ala
35 40 45gat gtt aat ctt aga aat cta agg ggt cac act gtg cta cat gtg gca 193Asp Val Asn Leu Arg Asn Leu Arg Gly His Thr Val Leu His Val Ala
50 55 60gcc atg aga aaa gag cct aag ata att gta tcc ttg tta acc aag gga 241Ala Met Arg Lys Glu Pro Lys Ile Ile Val Ser Leu Leu Thr Lys Gly65 70 75 80gcc cat ccg tct gat ata aca tca gat gat aaa aaa gca ctg cag ata 289Ala His Pro Ser Asp Ile Thr Ser Asp Asp Lys Lys Ala Leu Gln Ile
85 90 95gca aag aga cta aca aaa gct gtg gac ttc tat aaa act aca gaa caa 337Ala Lys Arg Leu Thr Lys Ala Val Asp Phe Tyr Lys Thr Thr Glu Gln
100 105 110gga aaa gat gca cca aag gat cgg ttg tgc att gaa ata ctg gag caa 385Gly Lys Asp Ala Pro Lys Asp Arg Leu Cys Ile Glu Ile Leu Glu Gln
115 120 125gct gaa aga aga gaa cca ttg cta gga gaa ggt tct gtt tct ctt gca 433Ala Glu Arg Arg Glu Pro Leu Leu Gly Glu Gly Ser Val Ser Leu Ala
130 135 140aag gca gga gat gat ctg cgt atg aag cta tta tac ctt gaa aat cga 481Lys Ala Gly Asp Asp Leu Arg Met Lys Leu Leu Tyr Leu Glu Asn Arg145 150 155 160gtt ggc ctt gct caa ct 498Val Gly Leu Ala Gln
165<210>40<211>165<212>PRT<213>甜菜<400>40Ala Leu Asp Ser Asp Asp Val Glu Leu Val Arg Met Leu Leu Lys Glu1 5 10 15Arg His Thr Thr Leu Asp Asp Ala Tyr Ala Leu His Tyr Ala Val Ala
20 25 30His Cys Asp Ala Lys Thr Thr Thr Glu Leu Leu Glu Leu Gly Leu Ala
35 40 45Asp Val Asn Leu Arg Asn Leu Arg Gly His Thr Val Leu His Val Ala
50 55 60Ala Met Arg Lys Glu Pro Lys Ile Ile Val Ser Leu Leu Thr Lys Gly65 70 75 80Ala His Pro Ser Asp Ile Thr Ser Asp Asp Lys Lys Ala Leu Gln Ile
85 90 95Ala Lys Arg Leu Thr Lys Ala Val Asp Phe Tyr Lys Thr Thr Glu Gln
100 105 110Gly Lys Asp Ala Pro Lys Asp Arg Leu Cys Ile Glu Ile Leu Glu Gln
115 120 125Ala Glu Arg Arg Glu Pro Leu Leu Gly Glu Gly Ser Val Ser Leu Ala
130 135 140Lys Ala Gly Asp Asp Leu Arg Met Lys Leu Leu Tyr Leu Glu Asn Arg145 150 155 160Val Gly Leu Ala Gln
165<210>41<211>498<212>DNA<213>向日葵<220><221>CDS<222>(2)..(496)<223>向日葵A<400>41g gca ttg gat tct gat gat gtt gag yta gtc aca atg tta tta cga gaa 49Ala Leu Asp Ser Asp Asp Val Glu Xaa Val Thr Met Leu Leu Arg Glu
1 5 10 15ggt cat act tca tta gac ggt tct tgc gct ctt cat tac gct gtt gcg 97Gly His Thr Ser Leu Asp Gly Ser Cys Ala Leu His Tyr Ala Val Ala
20 25 30tac gca gat gct aaa acg aca acc gaa tta ctg gat tta gca ctt gct 145Tyr Ala Asp Ala Lys Thr Thr Thr Glu Leu Leu Asp Leu Ala Leu Ala
35 40 45gac gta aat cat aaa aac tcg agg ggt ttt acc gta ctt cat gtt gcc 193Asp Val Asn His Lys Asn Ser Arg Gly Phe Thr Val Leu His Val Ala
50 55 60gct atg aga aaa gag ccg agt att atc gtt tcg ctt ctt acg aaa ggg 241Ala Met Arg Lys Glu Pro Ser Ile Ile Val Ser Leu Leu Thr Lys Gly65 70 75 80gcc cga ccc tcg gat ctc acc cct gat ggg aga aaa gca cta cag att 289Ala Arg Pro Ser Asp Leu Thr Pro Asp Gly Arg Lys Ala Leu Gln Ile
85 90 95tcg aag agg ttg acc aga gcg gtt gac tat tac aag tca aac gag gat 337Ser Lys Arg Leu Thr Arg Ala Val Asp Tyr Tyr Lys Ser Asn Glu Asp
100 105 110gat aaa gag tca acg aaa ggt cgt ttg tgt att gag ata ttg gaa caa 385Asp Lys Glu Ser Thr Lys Gly Arg Leu Cys Ile Glu Ile Leu Glu Gln
115 120 125gcc gaa aga aga aat cca ttg tta ggt gaa gct tcg gct tct ctt gca 433Ala Glu Arg Arg Asn Pro Leu Leu Gly Glu Ala Ser Ala Ser Leu Ala
130 135 140atg gcc gga gat gat ttg cgt gga aag ttg ttg tac ctt gaa aat cga 481Met Ala Gly Asp Asp Leu Arg Gly Lys Leu Leu Tyr Leu Glu Asn Arg145 150 155 160gtt ggc ctg gct caa ct 498Val Gly Leu Ala Gln
165<210>42<211>165<212>PRT<213>向日葵<400>42Ala Leu Asp Ser Asp Asp Val Glu Xaa Val Thr Met Leu Leu Arg Glu1 5 10 15Gly His Thr Ser Leu Asp Gly Ser Cys Ala Leu His Tyr Ala Val Ala
20 25 30Tyr Ala Asp Ala Lys Thr Thr Thr Glu Leu Leu Asp Leu Ala Leu Ala
35 40 45Asp Val Asn His Lys Asn Ser Arg Gly Phe Thr Val Leu His Val Ala
50 55 60Ala Met Arg Lys Glu Pro Ser Ile Ile Val Ser Leu Leu Thr Lys Gly65 70 75 80Ala Arg Pro Ser Asp Leu Thr Pro Asp Gly Arg Lys Ala Leu Gln Ile
85 90 95Ser Lys Arg Leu Thr Arg Ala Val Asp Tyr Tyr Lys Ser Asn Glu Asp
100 105 110Asp Lys Glu Ser Thr Lys Gly Arg Leu Cys Ile Glu Ile Leu Glu Gln
115 120 125Ala Glu Arg Arg Asn Pro Leu Leu Gly Glu Ala Ser Ala Ser Leu Ala
130 135 140Met Ala Gly Asp Asp Leu Arg Gly Lys Leu Leu Tyr Leu Glu Asn Arg145 150 155 160Val Gly Leu Ala Gln
165<210>43<211>498<212>DNA<213>向日葵<220><221>CDS<222>(2)..(496)<223>向日葵B<400>43g gca ttg gac tct gat gat gtt gag ctt gtg aaa atg att tta gac gaa 49Ala Leu Asp Ser Asp Asp Val Glu Leu Val Lys Met Ile Leu Asp Glu
1 5 10 15tcc aaa atc acg tta gat gaa gcc tgc gct ctt cat tat gcg gtc atg 97Ser Lys Ile Thr Leu Asp Glu Ala Cys Ala Leu His Tyr Ala Val Met
20 25 30tat tgt aat caa gaa gtt gct aag gag att ctt aac tta aac cgt gcg 145Tyr Cys Asn Gln Glu Val Ala Lys Glu Ile Leu Asn Leu Asn Arg Ala
35 40 45gat gtt aat ctt aga aac tca cga gat tac acc gtg ctt cat gtt gct 193Asp Val Asn Leu Arg Asn Ser Arg Asp Tyr Thr Val Leu His Val Ala
50 55 60gcc atg cgt aaa gaa cca tca ctt att gtt tcg att cta agc aaa ggc 241Ala Met Arg Lys Glu Pro Ser Leu Ile Val Ser Ile Leu Ser Lys Gly65 70 75 80gcg tgt gca tcg gat act act ttt gat gga caa agt gcg gtt agt att 289Ala Cys Ala Ser Asp Thr Thr Phe Asp Gly Gln Ser Ala Val Ser Ile
85 90 95tgc agg aga cga aca agg ccc aag gat tat tat gtg aaa acc gaa cac 337Cys Arg Arg Arg Thr Arg Pro Lys Asp Tyr Tyr Val Lys Thr Glu His
100 105 110ggg caa gaa aca aat aaa gat cgt ata tgc atc gat gtt ttg gag cgg 385Gly Gln Glu Thr Asn Lys Asp Arg Ile Cys Ile Asp Val Leu Glu Arg
115 120 125gaa ata aag agg aat ccg atg ata ggc gat gtt tcc gtg tgt tct tca 433Glu Ile Lys Arg Asn Pro Met Ile Gly Asp Val Ser Val Cys Ser Ser
130 135 140gca gtg gct gat gat ttg cat atg aat tta ctc tac ttt gaa aat cga 481Ala Val Ala Asp Asp Leu His Met Asn Leu Leu Tyr Phe Glu Asn Arg145 150 155 160gtt ggc ctt gct caa ct 498Val Gly Leu Ala Gln
165<210>44<211>165<212>PRT<213>向日葵<400>44Ala Leu Asp Ser Asp Asp Val Glu Leu Val Lys Met Ile Leu Asp Glu1 5 10 15Ser Lys Ile Thr Leu Asp Glu Ala Cys Ala Leu His Tyr Ala Val Met
20 25 30Tyr Cys Asn Gln Glu Val Ala Lys Glu Ile Leu Asn Leu Asn Arg Ala
35 40 45Asp Val Asn Leu Arg Ash Ser Arg Asp Tyr Thr Val Leu His Val Ala
50 55 60Ala Met Arg Lys Glu Pro Ser Leu Ile Val Ser Ile Leu Ser Lys Gly65 70 75 80Ala Cys Ala Ser Asp Thr Thr Phe Asp Gly Gln Ser Ala Val Ser Ile
85 90 95Cys Arg Arg Arg Thr Arg Pro Lys Asp Tyr Tyr Val Lys Thr Glu His
100 105 110Gly Gln Glu Thr Asn Lys Asp Arg Ile Cys Ile Asp Val Leu Glu Arg
115 120 125Glu Ile Lys Arg Asn Pro Met Ile Gly Asp Val Ser Val Cys Ser Ser
130 135 140Ala Val Ala Asp Asp Leu His Met Asn Leu Leu Tyr Phe Glu Asn Arg145 150 155 160Val Gly Leu Ala Gln
165<210>45<211>653<212>DNA<213>马铃薯<220><221>CDS<222>(1)..(651)<223>马铃薯A<400>45gak att att gtc aag tct aat gtt gat atc ata acc ctt gat aag tcc 48Xaa Ile Ile Val Lys Ser Asn Val Asp Ile Ile Thr Leu Asp Lys Ser1 5 10 15ttg cct cat gac atc gta aaa caa atc act gat tca cgt gct gaa ctt 96Leu Pro His Asp Ile Val Lys Gln Ile Thr Asp Ser Arg Ala Glu Leu
20 25 30ggt cta caa ggg cct gaa agc aat ggt ttt cct gat aaa cat gtt aag 144Gly Leu Gln Gly Pro Glu Ser Asn Gly Phe Pro Asp Lys His Val Lys
35 40 45agg ata cat agg gca ttg gac tct gat gat gtt gag tta cta agg atg 192Arg Ile His Arg Ala Leu Asp Ser Asp Asp Val Glu Leu Leu Arg Met
50 55 60ttg ctt aaa gaa ggg cat act act ctc gat gat gca tat gct ctc cac 240Leu Leu Lys Glu Gly His Thr Thr Leu Asp Asp Ala Tyr Ala Leu His65 70 75 80tat gct gta gca tat tgc gat gca aag act aca gca gaa ctt tta gat 288Tyr Ala Val Ala Tyr Cys Asp Ala Lys Thr Thr Ala Glu Leu Leu Asp
85 90 95ctt tca ctt gct gat gtt aat cat caa aat cct aga gga tac acg gta 336Leu Ser Leu Ala Asp Val Asn His Gln Asn Pro Arg Gly Tyr Thr Val
100 105 110ctt cat gtt gct gcc atg agg aaa gag cct aaa att ata gtg tcc ctt 384Leu His Val Ala Ala Met Arg Lys Glu Pro Lys Ile Ile Val Ser Leu
115 120 125tta acc aaa gga gct aga cct tct gat ctg aca tct gat ggc aaa aaa 432Leu Thr Lys Gly Ala Arg Pro Ser Asp Leu Thr Ser Asp Gly Lys Lys
130 135 140gca ctt caa att gct aag agg ctc act agg ctt gtg gat ttt act aag 480Ala Leu Gln Ile Ala Lys Arg Leu Thr Arg Leu Val Asp Phe Thr Lys145 150 155 160tct aca gag gaa gga aaa tct gct cca aaa gat cgg tta tgc att gag 528Ser Thr Glu Glu Gly Lys Ser Ala Pro Lys Asp Arg Leu Cys Ile Glu
165 170 175att ctg gag caa gca gaa aga aga gat cca cta cta gga gaa gct tca 576Ile Leu Glu Gln Ala Glu Arg Arg Asp Pro Leu Leu Gly Glu Ala Ser
180 185 190tta tct ctt gct atg gca ggc gat gat ttg cgt atg aag ctg tta tac 624Leu Ser Leu Ala Met Ala Gly Asp Asp Leu Arg Met Lys Leu Leu Tyr
195 200 205ctt gaa aat cga gtt ggc ctk gct caa ct 653Leu Glu Asn Arg Val Gly Xaa Ala Gln
210 215<210>46<211>217<212>PRT<213>马铃薯<400>46Xaa Ile Ile Val Lys Ser Asn Val Asp Ile Ile Thr Leu Asp Lys Ser1 5 10 15Leu Pro His Asp Ile Val Lys Gln Ile Thr Asp Ser Arg Ala Glu Leu
20 25 30Gly Leu Gln Gly Pro Glu Ser Asn Gly Phe Pro Asp Lys His Val Lys
35 40 45Arg Ile His Arg Ala Leu Asp Ser Asp Asp Val Glu Leu Leu Arg Met
50 55 60Leu Leu Lys Glu Gly His Thr Thr Leu Asp Asp Ala Tyr Ala Leu His65 70 75 80Tyr Ala Val Ala Tyr Cys Asp Ala Lys Thr Thr Ala Glu Leu Leu Asp
85 90 95Leu Ser Leu Ala Asp Val Asn His Gln Asn Pro Arg Gly Tyr Thr Val
100 105 110Leu His Val Ala Ala Met Arg Lys Glu Pro Lys Ile Ile Val Ser Leu
115 120 125Leu Thr Lys Gly Ala Arg Pro Ser Asp Leu Thr Ser Asp Gly Lys Lys
130 135 140Ala Leu Gln Ile Ala Lys Arg Leu Thr Arg Leu Val Asp Phe Thr Lys145 150 155 160Ser Thr Glu Glu Gly Lys Ser Ala Pro Lys Asp Arg Leu Cys Ile Glu
165 170 175Ile Leu Glu Gln Ala Glu Arg Arg Asp Pro Leu Leu Gly Glu Ala Ser
180 185 190Leu Ser Leu Ala Met Ala Gly Asp Asp Leu Arg Met Lys Leu Leu Tyr
195 200 205Leu Glu Asn Arg Val Gly Xaa Ala Gln
210 215<210>47<211>498<212>DNA<213>马铃薯<220><221>CDS<222>(2)..(496)<223>马铃薯B<400>47g gca ttg gat tca gat gat gtt gag ttt gtc aag ctt cta ctt aat gag 49Ala Leu Asp Ser Asp Asp Val Glu Phe Val Lys Leu Leu Leu Asn Glu
1 5 10 15tct gac ata agt tta gat gga gcc tac gct ctt cat tac gct gtt gca 97Ser Asp Ile Ser Leu Asp Gly Ala Tyr Ala Leu His Tyr Ala Val Ala
20 25 30tat tgt gac ccc aag gtt gtt act gag gtt ctt gga ctg ggt gtt gct 145Tyr Cys Asp Pro Lys Val Val Thr Glu Val Leu Gly Leu Gly Val Ala
35 40 45aat gtc aac ctt cgg aat aca cgt ggt tac act gtg ctt cac att gct 193Asn Val Asn Leu Arg Asn Thr Arg Gly Tyr Thr Val Leu His Ile Ala
50 55 60gcc atg cgt aag gaa ccc tca atc att gta tca ctt ttg act aag gga 241Ala Met Arg Lys Glu Pro Ser Ile Ile Val Ser Leu Leu Thr Lys Gly65 70 75 80gct cat gca tca gaa att aca ttg gat ggg cag agt gct gtt ggc atc 289Ala His Ala Ser Glu Ile Thr Leu Asp Gly Gln Ser Ala Val Gly Ile
85 90 95tgt agg agg ctg agt agg cct aag gag tac cat gca aaa aca gaa caa 337Cys Arg Arg Leu Ser Arg Pro Lys Glu Tyr His Ala Lys Thr Glu Gln
100 105 110ggc cag gaa gca aac aaa gat cgg gta tgt att gat gtt ttg gag aga 385Gly Gln Glu Ala Asn Lys Asp Arg Val Cys Ile Asp Val Leu Glu Arg
115 120 125gag atg cgt cac aac cca atg acc gga gat gca tta ttt tct tcc ccc 433Glu Met Arg His Asn Pro Met Thr Gly Asp Ala Leu Phe Ser Ser Pro
130 135 140atg ttg gcc gat gat ctg ccc atg aaa ctg ctc tac ctt gaa aat cga 481Met Leu Ala Asp Asp Leu Pro Met Lys Leu Leu Tyr Leu Glu Asn Arg145 150 155 160gtt ggc ctt gct aaa ct 498Val Gly Leu Ala Lys
165<210>48<211>165<212>PRT<213>马铃薯<400>48Ala Leu Asp Ser Asp Asp Val Glu Phe Val Lys Leu Leu Leu Asn Glu1 5 10 15Ser Asp Ile Ser Leu Asp Gly Ala Tyr Ala Leu His Tyr Ala Val Ala
20 25 30Tyr Cys Asp Pro Lys Val Val Thr Glu Val Leu Gly Leu Gly Val Ala
35 40 45Asn Val Asn Leu Arg Asn Thr Arg Gly Tyr Thr Val Leu His Ile Ala
50 55 60Ala Met Arg Lys Glu Pro Ser Ile Ile Val Ser Leu Leu Thr Lys Gly65 70 75 80Ala His Ala Ser Glu Ile Thr Leu Asp Gly Gln Ser Ala Val Gly Ile
85 90 95Cys Arg Arg Leu Ser Arg Pro Lys Glu Tyr His Ala Lys Thr Glu Gln
100 105 110Gly Gln Glu Ala Asn Lys Asp Arg Val Cys Ile Asp Val Leu Glu Arg
115 120 125Glu Met Arg His Asn Pro Met Thr Gly Asp Ala Leu Phe Ser Ser Pro
130 135 140Met Leu Ala Asp Asp Leu Pro Met Lys Leu Leu Tyr Leu Glu Asn Arg145 150 155 160Val Gly Leu Ala Lys
165<210>49<211>477<212>DNA<213>马铃薯<220><221>CDS<222>(2)..(475)<223>马铃薯C<400>49g gca ctg gac tct gat gat gtt gag ttt gtc aag ctt cta ctt aat gag 49Ala Leu Asp Ser Asp Asp Val Glu Phe Val Lys Leu Leu Leu Asn Glu
1 5 10 15tct gac ata agt tta gat gga gcc tac gct ctt cat tac gct gtt gca 97Ser Asp Ile Ser Leu Asp Gly Ala Tyr Ala Leu His Tyr Ala Val Ala
20 25 30tat tgt gac ccc aag gtt gtt act gag gtt ctt gga ctg ggt gtt gct 145Tyr Cys Asp Pro Lys Val Val Thr Glu Val Leu Gly Leu Gly Val Ala
35 40 45aat gtc aac ctt cgg aat aca cgt ggt tac act gtg ctt cac att gct 193Asn Val Asn Leu Arg Asn Thr Arg Gly Tyr Thr Val Leu His Ile Ala
50 55 60gcc atg cgt aag gaa ccc tca atc att gta tca ctt ttg act aag gga 241Ala Met Arg Lys Glu Pro Ser Ile Ile Val Ser Leu Leu Thr Lys Gly65 70 75 80gct cat gca tca gaa att aca ttg gat ggg cag agt gct gtt agc atc 289Ala His Ala Ser Glu Ile Thr Leu Asp Gly Gln Ser Ala Val Ser Ile
85 90 95tgt agg agg ctg act agg cct aag gag tac cat gca aaa aca gaa caa 337Cys Arg Arg Leu Thr Arg Pro Lys Glu Tyr His Ala Lys Thr Glu Gln
100 105 110ggc cag gaa gca aac aaa gat cgg gta tgt att gat gtt ttg gag aga 385Gly Gln Glu Ala Asn Lys Asp Arg Val Cys Ile Asp Val Leu Glu Arg
115 120 125gag atg cgt cgc aac cca atg acc gga gat gca tta ttt tct tcc ccc 433Glu Met Arg Arg Asn Pro Met Thr Gly Asp Ala Leu Phe Ser Ser Pro
130 135 140atg aaa cag ctc tac ctt gaa aat aga gtt ggc ctt gct aaa ct 477Met Lys Gln Leu Tyr Leu Glu Asn Arg Val Gly Leu Ala Lys145 150 155<210>50<211>158<212>PRT<213>马铃薯<400>50Ala Leu Asp Ser Asp Asp Val Glu Phe Val Lys Leu Leu Leu Asn Glu1 5 10 15Ser Asp Ile Ser Leu Asp Gly Ala Tyr Ala Leu His Tyr Ala Val Ala
20 25 30Tyr Cys Asp Pro Lys Val Val Thr Glu Val Leu Gly Leu Gly Val Ala
35 40 45Asn Val Asn Leu Arg Asn Thr Arg Gly Tyr Thr Val Leu His Ile Ala
50 55 60Ala Met Arg Lys Glu Pro Ser Ile Ile Val Ser Leu Leu Thr Lys Gly65 70 75 80Ala His Ala Ser Glu Ile Thr Leu Asp Gly Gln Ser Ala Val Ser Ile
85 90 95Cys Arg Arg Leu Thr Arg Pro Lys Glu Tyr His Ala Lys Thr Glu Gln
100 105 110Gly Gln Glu Ala Asn Lys Asp Arg Val Cys Ile Asp Val Leu Glu Arg
115 120 125Glu Met Arg Arg Asn Pro Met Thr Gly Asp Ala Leu Phe Ser Ser Pro
130 135 140Met Lys Gln Leu Tyr Leu Glu Asn Arg Val Gly Leu Ala Lys145 150 155<210>51<211>501<212>DNA<213>欧洲油菜<220><221>CDS<222>(2)..(499)<223>Canola A<400>51g gca ttg gat tct gat gat gtt gag ttt gtg aag ttg ctt ttg act gag 49Ala Leu Asp Ser Asp Asp Val Glu Phe Val Lys Leu Leu Leu Thr Glu
1 5 10 15tca gat atc act cta gat gaa gcc aat ggt ctt cat tac tca gtg gtg 97Ser Asp Ile Thr Leu Asp Glu Ala Asn Gly Leu His Tyr Ser Val Val
20 25 30tat agt gat ccc aaa gtt gtt gcc gag att ctt act ctt gat atg ggt 145Tyr Ser Asp Pro Lys Val Val Ala Glu Ile Leu Thr Leu Asp Met Gly
35 40 45gat gtc aac cac aga aac tca cgt ggc tac acg gtt ctt cat ctc gca 193Asp Val Asn His Arg Asn Ser Arg Gly Tyr Thr Val Leu His Leu Ala
50 55 60gcc atg cgc aaa gag ccg tcc atc atc ata tct ctt ctc aag aga ggt 241Ala Met Arg Lys Glu Pro Ser Ile Ile Ile Ser Leu Leu Lys Arg Gly65 70 75 80gcc aat gcg tct ggc ttc acg tgt gat gga cgc agt gcg gtt aat ata 289Ala Asn Ala Ser Gly Phe Thr Cys Asp Gly Arg Ser Ala Val Asn Ile
85 90 95tgt aga aga ttg aca act cca aag gat tat cat acg aaa aca gct gcg 337Cys Arg Arg Leu Thr Thr Pro Lys Asp Tyr His Thr Lys Thr Ala Ala
100 105 110aaa ggg agg gaa gct agt aaa gca cgg tta tgt ata gat ctc ttg gaa 385Lys Gly Arg Glu Ala Ser Lys Ala Arg Leu Cys Ile Asp Leu Leu Glu
115 120 125aga gaa gta agg agg aac cct atg gtt gtt gat tca cca atg tgt tcc 433Arg Glu Val Arg Arg Asn Pro Met Val Val Asp Ser Pro Met Cys Ser
130 135 140ctt tct atg cct gaa gat ctc caa atg aga ctg tta tac ctt gaa aat 481Leu Ser Met Pro Glu Asp Leu Gln Met Arg Leu Leu Tyr Leu Glu Asn145 150 155 160cga gtt ggc ctt gct caa ct 501Arg Val Gly Leu Ala Gln
165<210>52<211>166<212>PRT<213>欧洲油菜<400>52Ala Leu Asp Ser Asp Asp Val Glu Phe Val Lys Leu Leu Leu Thr Glu1 5 10 15Ser Asp Ile Thr Leu Asp Glu Ala Asn Gly Leu His Tyr Ser Val Val
20 25 30Tyr Ser Asp Pro Lys Val Val Ala Glu Ile Leu Thr Leu Asp Met Gly
35 40 45Asp Val Asn His Arg Asn Ser Arg Gly Tyr Thr Val Leu His Leu Ala
50 55 60Ala Met Arg Lys Glu Pro Ser Ile Ile Ile Ser Leu Leu Lys Arg Gly65 70 75 80Ala Asn Ala Ser Gly Phe Thr Cys Asp Gly Arg Ser Ala Val Asn Ile
85 90 95Cys Arg Arg Leu Thr Thr Pro Lys Asp Tyr His Thr Lys Thr Ala Ala
100 105 110Lys Gly Arg Glu Ala Ser Lys Ala Arg Leu Cys Ile Asp Leu Leu Glu
115 120 125Arg Glu Val Arg Arg Asn Pro Met Val Val Asp Ser Pro Met Cys Ser
130 135 140Leu Ser Met Pro Glu Asp Leu Gln Met Arg Leu Leu Tyr Leu Glu Asn145 150 155 160Arg Val Gly Leu Ala Gln
165<210>53<211>501<212>DNA<213>欧洲油菜<220><221>CDS<222>(2)..(499)<223>Canola B<400>53g gca ttg gat tct gat gat gtt gag ttt gtg aag ctt ctt ttg acc gag 49Ala Leu Asp Ser Asp Asp Val Glu Phe Val Lys Leu Leu Leu Thr Glu
1 5 10 15tca gat atc act cta gat gaa gcc aat ggt ctt cat tac tca gtg gtg 97Ser Asp Ile Thr Leu Asp Glu Ala Asn Gly Leu His Tyr Ser Val Val
20 25 30tat agt gat ccc aaa gtt gtt gcc gag att ctt act ctt gat atg ggt 145Tyr Ser Asp Pro Lys Val Val Ala Glu Ile Leu Thr Leu Asp Met Gly
35 40 45gat gtt aac cac aga aac tca cgt ggc tac acg gtt ctg cat ctc gca 193Asp Val Asn His Arg Asn Ser Arg Gly Tyr Thr Val Leu His Leu Ala
50 55 60gcc atg cgc aaa gag ccg tcc atc atc ata tct ctt ctc aag aaa ggt 241Ala Met Arg Lys Glu Pro Ser Ile Ile Ile Ser Leu Leu Lys Lys Gly65 70 75 80gcc aat gcg tct ggc ttc acc tgt gat gga cgc agt gcg gtt aat ata 289Ala Asn Ala Ser Gly Phe Thr Cys Asp Gly Arg Ser Ala Val Asn Ile
85 90 95tgt aga aga ttg aca act cca aag gat tat cat act aaa aca gct gcg 337Cys Arg Arg Leu Thr Thr Pro Lys Asp Tyr His Thr Lys Thr Ala Ala
100 105 110aaa ggg agg gaa gct agt aaa gca cgg tta tgt ata gat ctc ttg gaa 385Lys Gly Arg Glu Ala Ser Lys Ala Arg Leu Cys Ile Asp Leu Leu Glu
115 120 125aga gaa gta agg agg aac cct atg gtt gtt gag tca cca atg tgt tct 433Arg Glu Val Arg Arg Asn Pro Met Val Val Glu Ser Pro Met Cys Ser
130 135 140ctt tct atg cct gaa gat ctc caa atg aga ctg tta tac ctt gaa aat 481Leu Ser Met Pro Glu Asp Leu Gln Met Arg Leu Leu Tyr Leu Glu Asn145 150 155 160cga gtt ggc ctg gct caa ct 501Arg Val Gly Leu Ala Gln
165<210>54<211>166<212>PRT<213>欧洲油菜<400>54Ala Leu Asp Ser Asp Asp Val Glu Phe Val Lys Leu Leu Leu Thr Glu1 5 10 15Ser Asp Ile Thr Leu Asp Glu Ala Asn Gly Leu His Tyr Ser Val Val
20 25 30Tyr Ser Asp Pro Lys Val Val Ala Glu Ile Leu Thr Leu Asp Met Gly
35 40 45Asp Val Asn His Arg Asn Ser Arg Gly Tyr Thr Val Leu His Leu Ala
50 55 60Ala Met Arg Lys Glu Pro Ser Ile Ile Ile Ser Leu Leu Lys Lys Gly65 70 75 80Ala Asn Ala Ser Gly Phe Thr Cys Asp Gly Arg Ser Ala Val Asn Ile
85 90 95Cys Arg Arg Leu Thr Thr Pro Lys Asp Tyr His Thr Lys Thr Ala Ala
100 105 110Lys Gly Arg Glu Ala Ser Lys Ala Arg Leu Cys Ile Asp Leu Leu Glu
115 120 125Arg Glu Val Arg Arg Asn Pro Met Val Val Glu Ser Pro Met Cys Ser
130 135 140Leu Ser Met Pro Glu Asp Leu Gln Met Arg Leu Leu Tyr Leu Glu Asn145 150 155 160Arg Val Gly Leu Ala Gln
165<210>55<211>498<212>DNA<213>欧洲油菜<220><221>CDS<222>(2)..(496)<223>Canola C<400>55g gca ctg gat tct gat gat gtt gag ctt gtg aag ctt ctt ttg acc gag 49Ala Leu Asp Ser Asp Asp Val Glu Leu Val Lys Leu Leu Leu Thr Glu
1 5 10 15tca gat atc act cta gat gaa gcc aat ggt ctg cat tac tca gtg gtg 97Ser Asp Ile Thr Leu Asp Glu Ala Asn Gly Leu His Tyr Ser Val Val
20 25 30tat agt gat ccc aaa gtt gtt gca gag ata ctt gcc ctt ggt tta ggt 145Tyr Ser Asp Pro Lys Val Val Ala Glu Ile Leu Ala Leu Gly Leu Gly
35 40 45gat gtc aat cac aga aac tca cgt ggc tac tcg gtt ctt cat ttc gct 193Asp Val Asn His Arg Asn Ser Arg Gly Tyr Ser Val Leu His Phe Ala
50 55 60gcc atg cgt aga gag cct tcc atc atc ata tct ctt ctc aag gaa ggc 241Ala Met Arg Arg Glu Pro Ser Ile Ile Ile Ser Leu Leu Lys Glu Gly65 70 75 80gcc aat gcg tct agc ttc act ttt gat gga cgc agt gcg gtt aat ata 289Ala Asn Ala Ser Ser Phe Thr Phe Asp Gly Arg Ser Ala Val Asn Ile
85 90 95tgt agg aga ctg aca act cca aag gat tat cat aca aag aca tcc aaa 337Cys Arg Arg Leu Thr Thr Pro Lys Asp Tyr His Thr Lys Thr Ser Lys
100 105 110aag agg gaa gct agt aaa gca agg ctg tgc ata gat ctc ttg gaa aga 385Lys Arg Glu Ala Ser Lys Ala Arg Leu Cys Ile Asp Leu Leu Glu Arg
115 120 125gag gtt agg agg aac cct atg ctt gct gat acg cca atg tgt tca ctt 433Glu Val Arg Arg Asn Pro Met Leu Ala Asp Thr Pro Met Cys Ser Leu
130 135 140act atg cct gaa gat ctc caa atg aga ctg tta tac ctt gaa aat cga 481Thr Met Pro Glu Asp Leu Gln Met Arg Leu Leu Tyr Leu Glu Asn Arg145 150 155 160gtt ggt ctt gct aaa ct 498Val Gly Leu Ala Lys
165<210>56<211>165<212>PRT<213>欧洲油菜<400>56Ala Leu Asp Ser Asp Asp Val Glu Leu Val Lys Leu Leu Leu Thr Glu1 5 10 15Ser Asp Ile Thr Leu Asp Glu Ala Asn Gly Leu His Tyr Ser Val Val
20 25 30Tyr Ser Asp Pro Lys Val Val Ala Glu Ile Leu Ala Leu Gly Leu Gly
35 40 45Asp Val Asn His Arg Asn Ser Arg Gly Tyr Ser Val Leu His Phe Ala
50 55 60Ala Met Arg Arg Glu Pro Ser Ile Ile Ile Ser Leu Leu Lys Glu Gly65 70 75 80Ala Asn Ala Ser Ser Phe Thr Phe Asp Gly Arg Ser Ala Val Asn Ile
85 90 95Cys Arg Arg Leu Thr Thr Pro Lys Asp Tyr His Thr Lys Thr Ser Lys
100 105 110Lys Arg Glu Ala Ser Lys Ala Arg Leu Cys Ile Asp Leu Leu Glu Arg
115 120 125Glu Val Arg Arg Asn Pro Met Leu Ala Asp Thr Pro Met Cys Ser Leu
130 135 140Thr Met Pro Glu Asp Leu Gln Met Arg Leu Leu Tyr Leu Glu Asn Arg145 150 155 160Val Gly Leu Ala Lys
165<210>57<211>498<212>DNA<213>欧洲油菜<220><221>CDS<222>(2)..(496)<223>Canola D<400>57g gca ctg gac tct gat gat gtt gag ctt gtc aag atg ctt ttg aca gaa 49Ala Leu Asp Ser Asp Asp Val Glu Leu Val Lys Met Leu Leu Thr Glu
1 5 10 15gga cac acg agt cta gac gac gcc tac gct ctt cac tac gct gtt gca 97Gly His Thr Ser Leu Asp Asp Ala Tyr Ala Leu His Tyr Ala Val Ala
20 25 30cat tcc gat gtg aag acg gcc tct gat ctc ata gac ctt gag ctt gcg 145His Ser Asp Val Lys Thr Ala Ser Asp Leu Ile Asp Leu Glu Leu Ala
35 40 45gat gtt gac cat aga aac ctg agg ggg tac acg gcg ctt cac gtt gct 193Asp Val Asp His Arg Asn Leu Arg Gly Tyr Thr Ala Leu His Val Ala
50 55 60gcg atg agg aac gag ccg aag ctg atg gtt tat tta ttg act aaa ggt 241Ala Met Arg Asn Glu Pro Lys Leu Met Val Tyr Leu Leu Thr Lys Gly65 70 75 80gcg aat gcg tcg gag aca acg ttt gac ggt aga acg gct ctt gtg att 289Ala Asn Ala Ser Glu Thr Thr Phe Asp Gly Arg Thr Ala Leu Val Ile
85 90 95gca aaa aga ctc act aaa gct tct gag tat aat gct agt acg gag caa 337Ala Lys Arg Leu Thr Lys Ala Ser Glu Tyr Asn Ala Ser Thr Glu Gln
100 105 110ggg aag cct tct ctg aaa gga ggg cta tgc ata gag gta cta gag cat 385Gly Lys Pro Ser Leu Lys Gly Gly Leu Cys Ile Glu Val Leu Glu His
115 120 125gcg cgg aaa cta ggt agg ttg cct aga gat ggt tta cct tct ctt cca 433Ala Arg Lys Leu Gly Arg Leu Pro Arg Asp Gly Leu Pro Ser Leu Pro
130 135 140gct act cct gat gaa ctg agg atg agg ttg ctc tac ctt gaa aat cga 481Ala Thr Pro Asp Glu Leu Arg Met Arg Leu Leu Tyr Leu Glu Asn Arg145 150 155 160gtt ggc ctg gct caa ct 498Val Gly Leu Ala Gln
165<210>58<211>165<212>PRT<213>欧洲油菜<400>58Ala Leu Asp Ser Asp Asp Val Glu Leu Val Lys Met Leu Leu Thr Glu1 5 10 15Gly His Thr Ser Leu Asp Asp Ala Tyr Ala Leu His Tyr Ala Val Ala
20 25 30His Ser Asp Val Lys Thr Ala Ser Asp Leu Ile Asp Leu Glu Leu Ala
35 40 45Asp Val Asp His Arg Asn Leu Arg Gly Tyr Thr Ala Leu His Val Ala
50 55 60Ala Met Arg Asn Glu Pro Lys Leu Met Val Tyr Leu Leu Thr Lys Gly65 70 75 80Ala Asn Ala Ser Glu Thr Thr Phe Asp Gly Arg Thr Ala Leu Val Ile
85 90 95Ala Lys Arg Leu Thr Lys Ala Ser Glu Tyr Asn Ala Ser Thr Glu Gln
100 105 110Gly Lys Pro Ser Leu Lys Gly Gly Leu Cys Ile Glu Val Leu Glu His
115 120 125Ala Arg Lys Leu Gly Arg Leu Pro Arg Asp Gly Leu Pro Ser Leu Pro
130 135 140Ala Thr Pro Asp Glu Leu Arg Met Arg Leu Leu Tyr Leu Glu Asn Arg145 150 155 160Val Gly Leu Ala Gln
165<210>59<211>31<212>DNA<213>人工序列<220><223>人工序列描述:PCR引物NIM3A<400>59tagatgawgc mtaygctcty caytatgctg t 31<210>60<211>32<212>DNA<213>人工序列<220><223>人工序列描述:PCR引物NIM3B<400>60ggctcyttmc kcatggcagc aayrtgaags ac 32<210>61<211>148<212>DNA<213>番茄<220><221>CDS<222>(4)..(147)<223>番茄B<400>61tag atg atg cat atg ctc ttc att atg ctg ttg cat att gtg acc cca 48
Met Met His Met Leu Phe Ile Met Leu Leu His Ile Val Thr Pro
1 5 10 15agg ttg ttg ctg agg ttc ttg gac tgg gtg ttg cta atg tca acc ttc 96Arg Leu Leu Leu Arg Phe Leu Asp Trp Val Leu Leu Met Ser Thr Phe
20 25 30gga atg cac gtg gtt aca ctg tcc ttc acg ttg ctg cca tgc gga aag 144Gly Met His Val Val Thr Leu Ser Phe Thr Leu Leu Pro Cys Gly Lys
35 40 45agc c 148Ser<210>62<211>48<212>PRT<213>番茄<400>62Met Met His Met Leu Phe Ile Met Leu Leu His Ile Val Thr Pro Arg1 5 10 15Leu Leu Leu Arg Phe Leu Asp Trp Val Leu Leu Met Ser Thr Phe Gly
20 25 30Met His Val Val Thr Leu Ser Phe Thr Leu Leu Pro Cys Gly Lys Ser
35 40 45<210>63<211>2296<212>DNA<213>甜菜<220><221>CDS<222>(113)..(1927)<223>全长甜菜cDNA序列<400>63cacacacaca cccgacgccg tatgcgtatc cattctctct cctcaacctc cctttgactt 60cctcttactc caccatcttc aatgtcgtcg atttccaatc tctaacattc ac atg aca 118
Met Thr
1acc acc tcc aca aca atg gtg atc gat tct cgc acc gct ttc tcc gat 166Thr Thr Ser Thr Thr Met Val Ile Asp Ser Arg Thr Ala Phe Ser Asp
5 10 15tcc aac gac atc agc aat ggc agt agc atc tgc tgc gtc gcc gca aca 214Ser Asn Asp Ile Ser Asn Gly Ser Ser Ile Cys Cys Val Ala Ala Thr
20 25 30aca act aca aca aca acc gcc gca gaa aac tct ctc tcc ttt act ccc 262Thr Thr Thr Thr Thr Thr Ala Ala Glu Asn Ser Leu Ser Phe Thr Pro35 40 45 50gac gcc gcc gct ctt ctc cgc ctc tct gaa aac ctc gac tcg ctt ttc 310Asp Ala Ala Ala Leu Leu Arg Leu Ser Glu Asn Leu Asp Ser Leu Phe
55 60 65caa ccc tcg ctt tct ctc tcc gac tcc gac tct ttc gcc gac gct aaa 358Gln Pro Ser Leu Ser Leu Ser Asp Ser Asp Ser Phe Ala Asp Ala Lys
70 75 80atc gtc gtt tcc ggt gat tcg cgt gaa gtc gcc gtt cat cgg tgt gtt 406Ile Val Val Ser Gly Asp Ser Arg Glu Val Ala Val His Arg Cys Val
85 90 95ctc tcg tct cgg agc tcg ttc ttt cgg tcc gct ttt gct tcg aaa cga 454Leu Ser Ser Arg Ser Ser Phe Phe Arg Ser Ala Phe Ala Ser Lys Arg
100 105 110gag aag gag aag gag agg gat aaa gag aga gtg gtg aag ctt gag ctt 502Glu Lys Glu Lys Glu Arg Asp Lys Glu Arg Val Val Lys Leu Glu Leu115 120 125 130aag gat tta gct ggt gat ttt gag gtt gga ttt gat tcg gtt gtt gcg 550Lys Asp Leu Ala Gly Asp Phe Glu Val Gly Phe Asp Ser Val Val Ala
135 140 145gtt tta ggt tat ttg tat agt ggc aaa gtt agg aat ttg cct aga gga 598Val Leu Gly Tyr Leu Tyr Ser Gly Lys Val Arg Asn Leu Pro Arg Gly
150 155 160att tgt gtt tgt gtt gat gag gat tgc tct cat gaa gct tgt cgt cct 646Ile Cys Val Cys Val Asp Glu Asp Cys Ser His Glu Ala Cys Arg Pro
165 170 175gct gtt gat ttt gtt gtt gag gtt ctc tat ttg tct cac aaa ttc gag 694Ala Val Asp Phe Val Val Glu Val Leu Tyr Leu Ser His Lys Phe Glu
180 185 190att gtc gaa ttg gtt tcg ctt tat cag agg cac cta ctg gat att ctt 742Ile Val Glu Leu Val Ser Leu Tyr Gln Arg His Leu Leu Asp Ile Leu195 200 205 210gac aag att gca cca gat gac gtt cta gta gtg tta tct gtc gct gag 790Asp Lys Ile Ala Pro Asp Asp Val Leu Val Val Leu Ser Val Ala Glu
215 220 225atg tgt gga aat gcg tgt gac gga ttg ctg gca agg tgt att gac aag 838Met Cys Gly Asn Ala Cys Asp Gly Leu Leu Ala Arg Cys Ile Asp Lys
230 235 240att gtg agg tcc gat att gac gta acc acc att gat aaa tcc ttg ccg 886Ile Val Arg Ser Asp Ile Asp Val Thr Thr Ile Asp Lys Ser Leu Pro
245 250 255cag aat gtt gtg aaa cag ata atc gac acg cga aag gaa ctt ggg ttt 934Gln Asn Val Val Lys Gln Ile Ile Asp Thr Arg Lys Glu Leu Gly Phe
260 265 270act gaa cct ggg cgt gtt gag ttt cct gat aag cat gtg aag aga ata 982Thr Glu Pro Gly Arg Val Glu Phe Pro Asp Lys His Val Lys Arg Ile275 280 285 290cac aga gct ttg gaa tcc gat gat gta gag tta gtc aga atg ctt tta 1030His Arg Ala Leu Glu Ser Asp Asp Val Glu Leu Val Arg Met Leu Leu
295 300 305aaa gag cgc cat aca act cta gat gat gca tat gcc ctt cac tat gct 1078Lys Glu Arg His Thr Thr Leu Asp Asp Ala Tyr Ala Leu His Tyr Ala
310 315 320gtg gca cat tgt gat gcc aag acc acc acg gag ctt ctt gag ctt ggg 1126Val Ala His Cys Asp Ala Lys Thr Thr Thr Glu Leu Leu Glu Leu Gly
325 330 335ctt gca gat gtt aat ctt aga aat cta agg ggt cac act gtg cta cat 1174Leu Ala Asp Val Asn Leu Arg Asn Leu Arg Gly His Thr Val Leu His
340 345 350gtg gca gcc atg aga aaa gag cct aag ata att gta tcc ttg tta acc 1222Val Ala Ala Met Arg Lys Glu Pro Lys Ile Ile Val Ser Leu Leu Thr355 360 365 370aag gga gcc cat ccg tct gat ata aca tca gat gat aaa aaa gca ctg 1270Lys Gly Ala His Pro Ser Asp Ile Thr Ser Asp Asp Lys Lys Ala Leu
375 380 385cag ata gca aag aga cta aca aaa gct gtg gac ttc tat aaa act aca 1318Gln Ile Ala Lys Arg Leu Thr Lys Ala Val Asp Phe Tyr Lys Thr Thr
390 395 400gaa caa gga aaa gat gca cca aag gat cgg ttg tgc att gaa ata ctg 1366Glu Gln Gly Lys Asp Ala Pro Lys Asp Arg Leu Cys Ile Glu Ile Leu
405 410 415gag caa gct gaa aga aga gaa cca ttg cta gga gaa ggt tct gtt tct 1414Glu Gln Ala Glu Arg Arg Glu Pro Leu Leu Gly Glu Gly Ser Val Ser
420 425 430ctt gca aag gca gga gat gat ctg cgt atg aag cta tta tat ctt gaa 1462Leu Ala Lys Ala Gly Asp Asp Leu Arg Met Lys Leu Leu Tyr Leu Glu435 440 445 450aat aga gtt gca ctt gct cgg ttg ctc ttt cca atg gaa gcg aaa gtg 1510Asn Arg Val Ala Leu Ala Arg Leu Leu Phe Pro Met Glu Ala Lys Val
455 460 465gct atg gat att gct caa gtg gac gga act tct gaa ttc aca ttg tca 1558Ala Met Asp Ile Ala Gln Val Asp Gly Thr Ser Glu Phe Thr Leu Ser
470 475 480aag aat ata gct gat gca cga aga aat gcg gtg gac ttg aat gag gct 1606Lys Asn Ile Ala Asp Ala Arg Arg Asn Ala Val Asp Leu Asn Glu Ala
485 490 495ccc ttt ata ttg aaa gag gag cat ttg cag agg atg aaa gca ctg tct 1654Pro Phe Ile Leu Lys Glu Glu His Leu Gln Arg Met Lys Ala Leu Ser
500 505 510aaa act gtt gag ctt ggc aag cgt ttc ttt cca cgc tgc tcc gat gtt 1702Lys Thr Val Glu Leu Gly Lys Arg Phe Phe Pro Arg Cys Ser Asp Val515 520 525 530ctt aat aag att atg gac gcc gaa gat cta tca cag ctt gca ttt tta 1750Leu Asn Lys Ile Met Asp Ala Glu Asp Leu Ser Gln Leu Ala Phe Leu
535 540 545gga aaa gat act cca gag gaa cgg caa agg aag aga aaa cga tac ctt 1798Gly Lys Asp Thr Pro Glu Glu Arg Gln Arg Lys Arg Lys Arg Tyr Leu
550 555 560gaa ctg caa gac gct tta act aag gct ttt aca gag gac aaa gaa gag 1846Glu Leu Gln Asp Ala Leu Thr Lys Ala Phe Thr Glu Asp Lys Glu Glu
565 570 575ttt gac cgt tct aca tta tca tca tcg tcg tcg tcg act cca atg ggg 1894Phe Asp Arg Ser Thr Leu Ser Ser Ser Ser Ser Ser Thr Pro Met Gly
580 585 590agg cca tat ggt aag acc aat ttc aag agg taa ctccttagca gctcaaagtt 1947Arg Pro Tyr Gly Lys Thr Asn Phe Lys Arg595 600 605gcatacgacg tcacttgtat aatattcatg tatatgtatg aaaatttctt tttgttctcc 2007ccttctattg atggccacgg tttcgatctt tttggtctgt attataattt ttgaccgatt 2067acttgataga attgtattct atacatcttt ataagctcat agtaacacca gatttaggta 2127ctatccgttg gagacacata ctcttgtgtg cgatgatgaa tcaatcatca gattacatta 2187cacgagccat ttcctgccat attgtaattc atgtatcaag gtacaaataa atagcgtcgt 2247ggggttgcac ctcttgcatt atcgaaaaaa aaaaaaaaaa aaaaaaaaa 2296<210>64<211>604<212>PRT<213>甜菜<400>64Met Thr Thr Thr Ser Thr Thr Met Val Ile Asp Ser Arg Thr Ala Phe1 5 10 15Ser Asp Ser Asn Asp Ile Ser Asn Gly Ser Ser Ile Cys Cys Val Ala
20 25 30Ala Thr Thr Thr Thr Thr Thr Thr Ala Ala Glu Asn Ser Leu Ser Phe
35 40 45Thr Pro Asp Ala Ala Ala Leu Leu Arg Leu Ser Glu Asn Leu Asp Ser
50 55 60Leu Phe Gln Pro Ser Leu Ser Leu Ser Asp Ser Asp Ser Phe Ala Asp65 70 75 80Ala Lys Ile Val Val Ser Gly Asp Ser Arg Glu Val Ala Val His Arg
85 90 95Cys Val Leu Ser Ser Arg Ser Ser Phe Phe Arg Ser Ala Phe Ala Ser
100 105 110Lys Arg Glu Lys Glu Lys Glu Arg Asp Lys Glu Arg Val Val Lys Leu
115 120 125Glu Leu Lys Asp Leu Ala Gly Asp Phe Glu Val Gly Phe Asp Ser Val
130 135 140Val Ala Val Leu Gly Tyr Leu Tyr Ser Gly Lys Val Arg Asn Leu Pro145 150 155 160Arg Gly Ile Cys Val Cys Val Asp Glu Asp Cys Ser His Glu Ala Cys
165 170 175Arg Pro Ala Val Asp Phe Val Val Glu Val Leu Tyr Leu Ser His Lys
180 185 190Phe Glu Ile Val Glu Leu Val Ser Leu Tyr Gln Arg His Leu Leu Asp
195 200 205Ile Leu Asp Lys Ile Ala Pro Asp Asp Val Leu Val Val Leu Ser Val
210 215 220Ala Glu Met Cys Gly Asn Ala Cys Asp Gly Leu Leu Ala Arg Cys Ile225 230 235 240Asp Lys Ile Val Arg Ser Asp Ile Asp Val Thr Thr Ile Asp Lys Ser
245 250 255Leu Pro Gln Asn Val Val Lys Gln Ile Ile Asp Thr Arg Lys Glu Leu
260 265 270Gly Phe Thr Glu Pro Gly Arg Val Glu Phe Pro Asp Lys His Val Lys
275 280 285Arg Ile His Arg Ala Leu Glu Ser Asp Asp Val Glu Leu Val Arg Met
290 295 300Leu Leu Lys Glu Arg His Thr Thr Leu Asp Asp Ala Tyr Ala Leu His305 310 315 320Tyr Ala Val Ala His Cys Asp Ala Lys Thr Thr Thr Glu Leu Leu Glu
325 330 335Leu Gly Leu Ala Asp Val Asn Leu Arg Asn Leu Arg Gly His Thr Val
340 345 350Leu His Val Ala Ala Met Arg Lys Glu Pro Lys Ile Ile Val Ser Leu
355 360 365Leu Thr Lys Gly Ala His Pro Ser Asp Ile Thr Ser Asp Asp Lys Lys
370 375 380Ala Leu Gln Ile Ala Lys Arg Leu Thr Lys Ala Val Asp Phe Tyr Lys385 390 395 400Thr Thr Glu Gln Gly Lys Asp Ala Pro Lys Asp Arg Leu Cys Ile Glu
405 410 415Ile Leu Glu Gln Ala Glu Arg Arg Glu Pro Leu Leu Gly Glu Gly Ser
420 425 430Val Ser Leu Ala Lys Ala Gly Asp Asp Leu Arg Met Lys Leu Leu Tyr
435 440 445Leu Glu Asn Arg Val Ala Leu Ala Arg Leu Leu Phe Pro Met Glu Ala
450 455 460Lys Val Ala Met Asp Ile Ala Gln Val Asp Gly Thr Ser Glu Phe Thr465 470 475 480Leu Ser Lys Asn Ile Ala Asp Ala Arg Arg Asn Ala Val Asp Leu Asn
485 490 495Glu Ala Pro Phe Ile Leu Lys Glu Glu His Leu Gln Arg Met Lys Ala
500 505 510Leu Ser Lys Thr Val Glu Leu Gly Lys Arg Phe Phe Pro Arg Cys Ser
515 520 525Asp Val Leu Asn Lys Ile Met Asp Ala Glu Asp Leu Ser Gln Leu Ala530 535 540Phe Leu Gly Lys Asp Thr Pro Glu Glu Arg Gln Arg Lys Arg Lys Arg545 550 555 560Tyr Leu Glu Leu Gln Asp Ala Leu Thr Lys Ala Phe Thr Glu Asp Lys
565 570 575Glu Glu Phe Asp Arg Ser Thr Leu Ser Ser Ser Ser Ser Ser Thr Pro
580 585 590Met Gly Arg Pro Tyr Gly Lys Thr Asn Phe Lys Arg
595 600<210>65<211>2844<212>DNA<213>向日葵<220><221>CDS<222>(737)..(2512)<223>全长向日葵B cDNA序列<400>65gacgataaaa cccctctctc tttttgctac caagaacctt cctactttct tgcaccaaag 60tttctttgca ggttctttga agcttcttta tcatcatacg ttggtttgat attgtttttg 120atgcatcttt tcacatgggt tttgcttatt gagtgattat ctgttgtggg tatttgatac 180aaattgaaaa aaagatgatt agatttggta tttagggttt tggttattga agattttatt 240aattagggtt tgattagggt ttgattaaga ttcttgtatt ggatgggttg atttagatcc 300agctgtttgt gggtttcaaa tttttgtttt ggtatttgca tatctcattc taatctattc 360agaggttgag gttctttagg tttgactttg actttgactt ttgggtactt tcttgtacat 420gtataatgtt tgatttgatc cattatatgt gttttgtaat tgaatcatag caaattttct 480tgcctgtata tatatgtttt attgaggatt tggttcaagt tttgaccttt ttgggaaaaa 540aagtcaaaca catattcttg ttcatgtagt tttgcaaatc aatcatttca caaatctttc 600tttatgttgg gaatccatct caatcataaa aaagtttctt tctttctttg agttcttgtt 660agctatgaaa gtttatgatt tgtccttttt gtgataaagt caaaccccta atcatcctgg 720gactttgact aaatcg atg gcg aat tca tcc gaa ccg tca tca tcc ata agc 772
Met Ala Asn Ser Ser Glu Pro Ser Ser Ser Ile Ser
1 5 10ttc acc tca tct tca cac ata tct aac ggc gca act agc tac aac ata 820Phe Thr Ser Ser Ser His Ile Ser Asn Gly Ala Thr Ser Tyr Asn Ile
15 20 25ccc cca cca tca atc ccc gag cca cgg tcg aat att gaa atc att ggc 868Pro Pro Pro Ser Ile Pro Glu Pro Arg Ser Asn Ile Glu Ile Ile Gly
30 35 40tta aat aga ctc agc aca aac cta gag aag ctc gta ttc gat tca ggt 916Leu Asn Arg Leu Ser Thr Asn Leu Glu Lys Leu Val Phe Asp Ser Gly45 50 55 60tct gaa tct gat tgc aat tac agc gat gct gaa gtt gtt gtt gag ggt 964Ser Glu Ser Asp Cys Asn Tyr Ser Asp Ala Glu Val Val Val Glu Gly
65 70 75att tct gta ggc att cat cgg tgt att tta gcc act aga agt acg ttt 1012Ile Ser Val Gly Ile His Arg Cys Ile Leu Ala Thr Arg Ser Thr Phe
80 85 90ttt agc gat ttg ttt aag aag aac aaa ggt tgt gta gag aag gac agt 1060Phe Ser Asp Leu Phe Lys Lys Asn Lys Gly Cys Val Glu Lys Asp Ser
95 100 105aag ccg aaa tat aac atg agt gat ttg ttg ccg tat ggg agc gtt ggg 1108Lys Pro Lys Tyr Asn Met Ser Asp Leu Leu Pro Tyr Gly Ser Val Gly
110 115 120tat gat gcg ttt ctc gtg ttt tta agc tat gtt tat act ggg aaa ctg 1156Tyr Asp Ala Phe Leu Val Phe Leu Ser Tyr Val Tyr Thr Gly Lys Leu125 130 135 140aaa gcg tct cct ccg gag gtt tca acc tgc gtt gat gat ggg tgt ctt 1204Lys Ala Ser Pro Pro Glu Val Ser Thr Cys Val Asp Asp Gly Cys Leu
145 150 155cat gat gct tgt tgg cct gct att aac ttt gct gtt gag ttg act tat 1252His Asp Ala Cys Trp Pro Ala Ile Asn Phe Ala Val Glu Leu Thr Tyr
160 165 170gcg tct tcg gtt ttt caa gtt ccg gaa tta gtt tcg ctt ttt cag cgt 1300Ala Ser Ser Val Phe Gln Val Pro Glu Leu Val Ser Leu Phe Gln Arg
175 180 185cgt ctt ctc aac ttt gtg gac aag gct ctt gtt gaa gac gtg atc ccg 1348Arg Leu Leu Asn Phe Val Asp Lys Ala Leu Val Glu Asp Val Ile Pro
190 195 200atc ctt gtt gtg gcc ttt cac tgt cag ttg caa aac gtc tta tct cgt 1396Ile Leu Val Val Ala Phe His Cys Gln Leu Gln Asn Val Leu Ser Arg205 210 215 220tgc att gac cga gta gtt agg tca aag ctc gat act att tcc att gaa 1444Cys Ile Asp Arg Val Val Arg Ser Lys Leu Asp Thr Ile Ser Ile Glu
225 230 235aaa gag ctt cca ttt gaa gtc acc caa atg atc aaa tcc att gat aac 1492Lys Glu Leu Pro Phe Glu Val Thr Gln Met Ile Lys Ser Ile Asp Asn
240 245 250atc atc caa gaa gat gac gaa cat aca gtc gaa tca gaa gtc gtg tta 1540Ile Ile Gln Glu Asp Asp Glu His Thr Val Glu Ser Glu Val Val Leu
255 260 265cgt gaa aag aga att aaa agc ata cac aaa gca tta gac tgt gac gat 1588Arg Glu Lys Arg Ile Lys Ser Ile His Lys Ala Leu Asp Cys Asp Asp
270 275 280gtt gag ctt gtg aaa atg att tta gac gaa tcc aaa atc acg tta gat 1636Val Glu Leu Val Lys Met Ile Leu Asp Glu Ser Lys Ile Thr Leu Asp285 290 295 300gaa gcc tgc gct ctt cat tat gcg gtc atg tat tgt aat caa gaa gtt 1684Glu Ala Cys Ala Leu His Tyr Ala Val Met Tyr Cys Asn Gln Glu Val
305 310 315gct aag gag att ctt aac tta aac cgt gcg gat gtt aat ctt aga aac 1732Ala Lys Glu Ile Leu Asn Leu Asn Arg Ala Asp Val Asn Leu Arg Asn
320 325 330tca cga gat tac acc gtg ctt cat gtt gct gcc atg cgt aaa gaa cca 1780Ser Arg Asp Tyr Thr Val Leu His Val Ala Ala Met Arg Lys Glu Pro
335 340 345tca ctt att gtt tcg att cta agc aaa ggc gcg tgt gca tcg gat act 1828Ser Leu Ile Val Ser Ile Leu Ser Lys Gly Ala Cys Ala Ser Asp Thr
350 355 360act ttt gat gga caa agt gcg gtt agt att tgc agg aga cga aca agg 1876Thr Phe Asp Gly Gln Ser Ala Val Ser Ile Cys Arg Arg Arg Thr Arg365 370 375 380ccc aag gat tat tat gtg aaa acc gaa cac ggg caa gaa aca aat aaa 1924Pro Lys Asp Tyr Tyr Val Lys Thr Glu His Gly Gln Glu Thr Asn Lys
385 390 395gat cgt ata tgc atc gat gtt ttg gag cgg gaa ata aag agg aat ccg 1972Asp Arg Ile Cys Ile Asp Val Leu Glu Arg Glu Ile Lys Arg Asn Pro
400 405 410atg ata ggc gat gtt tcc gtg tgt tct tca gca gtg gct gat gat ttg 2020Met Ile Gly Asp Val Ser Val Cys Ser Ser Ala Val Ala Asp Asp Leu
415 420 425cat atg aat tta ctc tac tta gaa aac cga gtg gca ttt gct cga ctg 2068His Met Asn Leu Leu Tyr Leu Glu Asn Arg Val Ala Phe Ala Arg Leu
430 435 440tta ttt ccg tca gaa gcg aaa cta gca atg gaa att gcg cat gcc caa 2116Leu Phe Pro Ser Glu Ala Lys Leu Ala Met Glu Ile Ala His Ala Gln445 450 455 460acg act gca cag tat ccg ggt cta ttg gca tcg aaa ggg tca aat ggt 2164Thr Thr Ala Gln Tyr Pro Gly Leu Leu Ala Ser Lys Gly Ser Asn Gly
465 470 475aac tta agg gag atg gat ttg aac gag aca ccg ttg gtg cag aac aaa 2212Asn Leu Arg Glu Met Asp Leu Asn Glu Thr Pro Leu Val Gln Asn Lys
480 485 490aga ttg ctt tca aga atg gaa gcc ctt tcc cgg aca gtg gaa atg ggt 2260Arg Leu Leu Ser Arg Met Glu Ala Leu Ser Arg Thr Val Glu Met Gly
495 500 505agg cga tat ttc cct cat tgt tca gag gtt ctg gat aag ttc atg gag 2308Arg Arg Tyr Phe Pro His Cys Ser Glu Val Leu Asp Lys Phe Met Glu
510 515 520gac gat cta cag gat ctt ttt atc ctc gag aag ggt acc gaa gaa gaa 2356Asp Asp Leu Gln Asp Leu Phe Ile Leu Glu Lys Gly Thr Glu Glu Glu525 530 535 540caa gaa atc aaa agg acg cga ttt atg gag ctt aaa gaa gat gtc caa 2404Gln Glu Ile Lys Arg Thr Arg Phe Met Glu Leu Lys Glu Asp Val Gln
545 550 555aga gcc ttt acc aag gac aag gcc gag ctt cat cgc ggt ttg tcc tca 2452Arg Ala Phe Thr Lys Asp Lys Ala Glu Leu His Arg Gly Leu Ser Ser
560 565 570tca atg tac acc ccc aca gtg aga aac ggg tca aag agt aaa gcc cgc 2500Ser Met Tyr Thr Pro Thr Val Arg Asn Gly Ser Lys Ser Lys Ala Arg
575 580 585aaa tac tca tga aacccccgtg tttctttgat gatcttttaa cacgctttta 2552Lys Tyr Ser
590cgtgcctaat attagaggca aaacatatgt atgaagaaat aatggtggtg catgatgatg 2612tttagggctc aggtttaggg tttatatgta ctaaattttg tgatttgacg ctaaaaatgc 2672tatgttgttt tttttttttt ttggataata tggtgtgaaa gctaacgcct tttactagta 2732gcatgttaat gtttgtgttt gaatcatagt tttttatgca tgtttgtttt acttgcacaa 2792caactaataa atataatttt tcataataaa aaaaaaaaaa aaaaaaaaaa aa 2844<210>66<211>591<212>PRT<213>向日葵<400>66Met Ala Asn Ser Ser Glu Pro Ser Ser Ser Ile Ser Phe Thr Ser Ser1 5 10 15Ser His Ile Ser Asn Gly Ala Thr Ser Tyr Asn Ile Pro Pro Pro Ser
20 25 30Ile Pro Glu Pro Arg Ser Asn Ile Glu Ile Ile Gly Leu Asn Arg Leu
35 40 45Ser Thr Asn Leu Glu Lys Leu Val Phe Asp Ser Gly Ser Glu Ser Asp
50 55 60Cys Asn Tyr Ser Asp Ala Glu Val Val Val Glu Gly lle Ser Val Gly65 70 75 80Ile His Arg Cys Ile Leu Ala Thr Arg Ser Thr Phe Phe Ser Asp Leu
85 90 95Phe Lys Lys Asn Lys Gly Cys Val Glu Lys Asp Ser Lys Pro Lys Tyr
100 105 110Asn Met Ser Asp Leu Leu Pro Tyr Gly Ser Val Gly Tyr Asp Ala Phe
115 120 125Leu Val Phe Leu Ser Tyr Val Tyr Thr Gly Lys Leu Lys Ala Ser Pro
130 135 140Pro Glu Val Ser Thr Cys Val Asp Asp Gly Cys Leu His Asp Ala Cys145 150 155 160Trp Pro Ala Ile Asn Phe Ala Val Glu Leu Thr Tyr Ala Ser Ser Val
165 170 175Phe Gln Val Pro Glu Leu Val Ser Leu Phe Gln Arg Arg Leu Leu Asn
180 185 190Phe Val Asp Lys Ala Leu Val Glu Asp Val Ile Pro Ile Leu Val Val
195 200 205Ala Phe His Cys Gln Leu Gln Asn Val Leu Ser Arg Cys Ile Asp Arg
210 215 220Val Val Arg ser Lys Leu Asp Thr Ile Ser Ile Glu Lys Glu Leu Pro225 230 235 240Phe Glu Val Thr Gln Met Ile Lys Ser Ile Asp Asn Ile Ile Gln Glu
245 250 255Asp Asp Glu His Thr Val Glu Ser Glu Val Val Leu Arg Glu Lys Arg
260 265 270Ile Lys Ser Ile His Lys Ala Leu Asp Cys Asp Asp Val Glu Leu Val
275 280 285Lys Met Ile Leu Asp Glu Ser Lys Ile Thr Leu Asp Glu Ala Cys Ala
290 295 300Leu His Tyr Ala Val Met Tyr Cys Asn Gln Glu Val Ala Lys Glu Ile305 310 315 320Leu Asn Leu Asn Arg Ala Asp Val Asn Leu Arg Asn Ser Arg Asp Tyr
325 330 335Thr Val Leu His Val Ala Ala Met Arg Lys Glu Pro Ser Leu Ile Val
340 345 350Ser Ile Leu Ser Lys Gly Ala Cys Ala Ser Asp Thr Thr Phe Asp Gly
355 360 365Gln Ser Ala Val Ser Ile Cys Arg Arg Arg Thr Arg Pro Lys Asp Tyr
370 375 380Tyr Val Lys Thr Glu His Gly Gln Glu Thr Asn Lys Asp Arg Ile Cys385 390 395 400Ile Asp Val Leu Glu Arg Glu Ile Lys Arg Asn Pro Met Ile Gly Asp
405 410 415Val Ser Val Cys Ser Ser Ala Val Ala Asp Asp Leu His Met Asn Leu
420 425 430Leu Tyr Leu Glu Asn Arg Val Ala Phe Ala Arg Leu Leu Phe Pro Ser
435 440 445Glu Ala Lys Leu Ala Met Glu Ile Ala His Ala Gln Thr Thr Ala Gln
450 455 460Tyr Pro Gly Leu Leu Ala Ser Lys Gly Ser Asn Gly Asn Leu Arg Glu465 470 475 480Met Asp Leu Asn Glu Thr Pro Leu Val Gln Asn Lys Arg Leu Leu Ser
485 490 495Arg Met Glu Ala Leu Ser Arg Thr Val Glu Met Gly Arg Arg Tyr Phe
500 505 510Pro His Cys Ser Glu Val Leu Asp Lys Phe Met Glu Asp Asp Leu Gln
515 520 525Asp Leu Phe Ile Leu Glu Lys Gly Thr Glu Glu Glu Gln Glu Ile Lys
530 535 540Arg Thr Arg Phe Met Glu Leu Lys Glu Asp Val Gln Arg Ala Phe Thr545 550 555 560Lys Asp Lys Ala Glu Leu His Arg Gly Leu Ser Ser Ser Met Tyr Thr
565 570 575Pro Thr Val Arg Asn Gly Ser Lys Ser Lys Ala Arg Lys Tyr Ser
580 585 590<210>67<211>1477<212>DNA<213>鼠耳芥<220><221>CDS<222>(1)..(804)<223>AtNMLc2 cDNA序列<220><221>misc_特征<222>(1)..(1477)<223>n=a、t、c或g<400>67atg agc aat ctt gaa gaa tct ttg aga tct cta tcg ttg gat ttc ctg 48Met Ser Asn Leu Glu Glu Ser Leu Arg Ser Leu Ser Leu Asp Phe Leu1 5 10 15aac cta cta atc aac ggt caa gct ttc tcc gac gtg act ttc agc gtt 96Asn Leu Leu Ile Asn Gly Gln Ala Phe Ser Asp Val Thr Phe Ser Val
20 25 30gaa ggt cgt tta gtc cac gct cac cgt tgt atc ctc gcc gca cgg agg 144Glu Gly Arg Leu Val His Ala His Arg Cys Ile Leu Ala Ala Arg Arg
35 40 45ctt ttc ttc cgc aaa ttc ttt tgt ggg aca gac tca cca caa cct gtc 192Leu Phe Phe Arg Lys Phe Phe Cys Gly Thr Asp Ser Pro Gln Pro Val
50 55 60aca ggt ata gac ccg acc caa cat ggg tcc gta ccc gct agc cca aca 240Thr Gly Ile Asp Pro Thr Gln His Gly Ser Val Pro Ala Ser Pro Thr65 70 75 80aga ggc tcc acg gcc cca gct gga att ata cca gtg aac tca gtc ggt 288Arg Gly Ser Thr Ala Pro Ala Gly Ile Ile Pro Val Asn Ser Val Gly
85 90 95tat gag gtt ttt ctg ttg cta ctt cag ttt ctt tat agc gga caa gtc 336Tyr Glu Val Phe Leu Leu Leu Leu Gln Phe Leu Tyr Ser Gly Gln Val
100 105 110tcc atc gtg ccg cag aaa cac gag cct aga cct aat tgt ggc gag aga 384Ser Ile Val Pro Gln Lys His Glu Pro Arg Pro Asn Cys Gly Glu Arg
115 120 125gga tgt tgg cac act cat tgc tca gcc gcc gtt gat ctt gct ctt gat 432Gly Cys Trp His Thr His Cys Ser Ala Ala Val Asp Leu Ala Leu Asp
130 135 140act ctc gcc gcc tct cgt tac ttc ggc gtc gag cag ctc gca ttg ctc 480Thr Leu Ala Ala Ser Arg Tyr Phe Gly Val Glu Gln Leu Ala Leu Leu145 150 155 160acc cag aaa caa ttg gca agc atg gtg gag aaa gcc tct atc gaa gat 528Thr Gln Lys Gln Leu Ala Ser Met Val Glu Lys Ala Ser Ile Glu Asp
165 170 175gtg atg aaa gtt tta ata gca tca aga aag caa gac atg cat caa tta 576Val Met Lys Val Leu Ile Ala Ser Arg Lys Gln Asp Met His Gln Leu
180 185 190tgg acc acc tgc tct cac tta gtt gct aaa tca ggt ctc cca cca gag 624Trp Thr Thr Cys Ser His Leu Val Ala Lys Ser Gly Leu Pro Pro Glu
195 200 205att ctt gcc aag cat ctc cct att gac gtc gtc acc aaa ata gaa gag 672Ile Leu Ala Lys His Leu Pro Ile Asp Val Val Thr Lys Ile Glu Glu
210 215 220ctt cgt ctt aaa tct tct ata gct cgc cgt tct cta atg cct cac aac 720Leu Arg Leu Lys Ser Ser Ile Ala Arg Arg Ser Leu Met Pro His Asn225 230 235 240cac cac cat gat ctc agc ggn gnt caa nac cta aag ntc aaa gtt aga 768His His His Asp Leu Ser Xaa Xaa Gln Xaa Leu Lys Xaa Lys Val Arg
245 250 255agg ttg agc cga ctt gga ttc ttc aac gnng aac tag taaagctgat 814Arg Leu Ser Arg Leu Gly Phe Phe Asn Xaa Asn
260 265ggtaatggan aaggactcca ttcttgatga agtcgtaagc attgcattac cgcttgttaa 874aagctgtaga agagaagttg tgaagncttt ngcttgaagc ttggaagctg ccgatgtgaa 934ttatccggcg ggtccggcaa ggnaaancac ctttgcactt cgcgggntga gatggtctct 994ccagacatgg tggctgttct gttagcccnc catgcttgat cctaatgtga ggacagttgg 1054tggaatcacg cctcttgata tccttagaac attaacttcg gatttcttgt tcaaggggca 1114gttcctggat tgactcacat tgaaccgaat aaacttaggc tttgcctcga gcttgttcaa 1174tccgctgcaa tggtgatatc tcgagaagaa ggaaacaata gcaacaacca aaacaatgat 1234aacaataccg ggatttaccc tcatatgaat gaggagcaca atagtggaag cagtggaggg 1294agcaataaca atttggattc aagattggtt tatctcaatc ttggagcagg tacgggtcag 1354atgggtccag gtcgagatca aggggatgac cataacagtc agagggaagg tatgagtcgg 1414catcatcatc atcatcaaga cccatctaca atgtatcatc accatcatca acatcacttc 1474tag 1477<210>68<211>267<212>PRT<213>鼠耳芥<400>68Met Ser Asn Leu Glu Glu Ser Leu Arg Ser Leu Ser Leu Asp Phe Leu1 5 10 15Asn Leu Leu Ile Asn Gly Gln Ala Phe Ser Asp Val Thr Phe Ser Val
20 25 30Glu Gly Arg Leu Val His Ala His Arg Cys Ile Leu Ala Ala Arg Arg
35 40 45Leu Phe Phe Arg Lys Phe Phe Cys Gly Thr Asp Ser Pro Gln Pro Val
50 55 60Thr Gly Ile Asp Pro Thr Gln His Gly Ser Val Pro Ala Ser Pro Thr65 70 75 80Arg Gly Ser Thr Ala Pro Ala Gly Ile Ile Pro Val Asn Ser Val Gly
85 90 95Tyr Glu Val Phe Leu Leu Leu Leu Gln Phe Leu Tyr Ser Gly Gln Val
100 105 110Ser Ile Val Pro Gln Lys His Glu Pro Arg Pro Asn Cys Gly Glu Arg
115 120 125Gly Cys Trp His Thr His Cys Ser Ala Ala Val Asp Leu Ala Leu Asp
130 135 140Thr Leu Ala Ala Ser Arg Tyr Phe Gly Val Glu Gln Leu Ala Leu Leu145 150 155 160Thr Gln Lys Gln Leu Ala Ser Met Val Glu Lys Ala Ser Ile Glu Asp
165 170 175Val Met Lys Val Leu Ile Ala Ser Arg Lys Gln Asp Met His Gln Leu
180 185 190Trp Thr Thr Cys Ser His Leu Val Ala Lys Ser Gly Leu Pro Pro Glu
195 200 205Ile Leu Ala Lys His Leu Pro Ile Asp Val Val Thr Lys Ile Glu Glu
210 215 220Leu Arg Leu Lys Ser Ser Ile Ala Arg Arg Ser Leu Met Pro His Asn225 230 235 240His His His Asp Leu Ser Xaa Xaa Gln Xaa Leu Lys Xaa Lys Val Arg
245 250 255Arg Leu Ser Arg Leu Gly Phe Phe Asn Xaa Asn
260 265<210>69<211>1725<212>DNA<213>鼠耳芥<220><221>CDS<222>(1)..(1725)<223>AtNMLC4-1 cDNA序列<400>69atg gct gca act gca ata gag cca tct tca tct ata agt ttc aca tct 48Met Ala Ala Thr Ala Ile Glu Pro Ser Ser Ser Ile Ser Phe Thr Ser1 5 10 15tct cac tta tca aac cct tct cct gtt gtt act act tat cac tca gct 96Ser His Leu Ser Asn Pro Ser Pro Val Val Thr Thr Tyr His Ser Ala
20 25 30gcc aat ctt gaa gag ctc agc tct aac ttg gag cag ctt ctc act aat 144Ala Asn Leu Glu Glu Leu Ser Ser Asn Leu Glu Gln Leu Leu Thr Asn
35 40 45cca gat tgc gat tac act gac gca gag atc atc att gaa gaa gaa gct 192Pro Asp Cys Asp Tyr Thr Asp Ala Glu Ile Ile Ile Glu Glu Glu Ala
50 55 60aac cct gtg agt gtt cat aga tgt gtt tta gct gct agg agc aag ttt 240Asn Pro Val Ser Val His Arg Cys Val Leu Ala Ala Arg Ser Lys Phe65 70 75 80ttt ctt gat ctg ttt aag aaa gat aaa gat agt agt gag aag aaa cct 288Phe Leu Asp Leu Phe Lys Lys Asp Lys Asp Ser Ser Glu Lys Lys Pro
85 90 95aag tat caa atg aaa gat tta tta cca tat gga aat gtg gga cgt gag 336Lys Tyr Gln Met Lys Asp Leu Leu Pro Tyr Gly Asn Val Gly Arg Glu
100 105 110gca ttt ctg cat ttc ttg agc tat atc tac act ggg agg tta aag cct 384Ala Phe Leu His Phe Leu Ser Tyr Ile Tyr Thr Gly Arg Leu Lys Pro
115 120 125ttt cct atc gag gtt tca act tgt gtt gat tca gtt tgt gct cat gat 432Phe Pro Ile Glu Val Ser Thr Cys Val Asp Ser Val Cys Ala His Asp
130 135 140tct tgt aaa ccg gcc att gat ttt gct gtt gag ttg atg tat gct tca 480Ser Cys Lys Pro Ala Ile Asp Phe Ala Val Glu Leu Met Tyr Ala Ser145 150 155 160ttt gtg ttc caa atc ccg gat ctt gtt tcg tca ttt cag cgg aag ctt 528Phe Val Phe Gln Ile Pro Asp Leu Val Ser Ser Phe Gln Arg Lys Leu
165 170 175cgt aac tat gtt gag aag tca cta gta gag aat gtt ctt cct atc ctc 576Arg Asn Tyr Val Glu Lys Ser Leu Val Glu Asn Val Leu Pro Ile Leu
180 185 190tta gtt gcg ttt cat tgt gat ttg aca cag ctt ctt gat caa tgc att 624Leu Val Ala Phe His Cys Asp Leu Thr Gln Leu Leu Asp Gln Cys Ile
195 200 205gag aga gtg gcg aga tca gac tta gac aga ttc tgt atc gaa aag gag 672Glu Arg Val Ala Arg Ser Asp Leu Asp Arg Phe Cys Ile Glu Lys Glu
210 215 220ctt cct tta gaa gta ttg gaa aaa atc aaa cag ctt cga gtt aag tcg 720Leu Pro Leu Glu Val Leu Glu Lys Ile Lys Gln Leu Arg Val Lys Ser225 230 235 240gtg aac ata ccc gag gtg gag gat aaa tcg ata gag aga aca ggg aaa 768Val Asn Ile Pro Glu Val Glu Asp Lys Ser Ile Glu Arg Thr Gly Lys
245 250 255gta ctc aag gca ttg gat tca gat gat gta gaa ctc gtg aag ctt ctt 816Val Leu Lys Ala Leu Asp Ser Asp Asp Val Glu Leu Val Lys Leu Leu
260 265 270ttg act gag tca gat ata act cta gac caa gcc aat ggt cta cat tat 864Leu Thr Glu Ser Asp Ile Thr Leu Asp Gln Ala Asn Gly Leu His Tyr
275 280 285gca gtg gca tac agt gat ccg aaa gtt gtg aca cag gtt ctt gat cta 912Ala Val Ala Tyr Ser Asp Pro Lys Val Val Thr Gln Val Leu Asp Leu
290 295 300gat atg gct gat gtt aat ttc aga aat tcc agg ggg tat acg gtt ctt 960Asp Met Ala Asp Val Asn Phe Arg Asn Ser Arg Gly Tyr Thr Val Leu305 310 315 320cat att gct gct atg cgt aga gag cca aca att atc ata cca ctt att 1008His Ile Ala Ala Met Arg Arg Glu Pro Thr Ile Ile Ile Pro Leu Ile
325 330 335caa aaa gga gct aat gct tca gat ttc acg ttt gat gga cgc agt gcg 1056Gln Lys Gly Ala Asn Ala Ser Asp Phe Thr Phe Asp Gly Arg Ser Ala
340 345 350gta aat ata tgt agg aga ctc act agg ccg aaa gat tat cat acc aaa 1104Val Asn Ile Cys Arg Arg Leu Thr Arg Pro Lys Asp Tyr His Thr Lys
355 360 365acc tca agg aaa gaa cct agt aaa tac cgc tta tgc atc gat atc ttg 1152Thr Ser Arg Lys Glu Pro Ser Lys Tyr Arg Leu Cys Ile Asp Ile Leu
370 375 380gaa agg gaa att aga agg aat cca ttg gtt agt ggg gat aca ccc act 1200Glu Arg Glu Ile Arg Arg Asn Pro Leu Val Ser Gly Asp Thr Pro Thr385 390 395 400tgt tcc cat tcg atg ccc gag gat ctc caa atg agg ttg tta tac tta 1248Cys Ser His Ser Met Pro Glu Asp Leu Gln Met Arg Leu Leu Tyr Leu
405 410 415gaa aag cga gtg gga ctt gct cag ttg ttc ttc cca gca gaa gcc aat 1296Glu Lys Arg Val Gly Leu Ala Gln Leu Phe Phe Pro Ala Glu Ala Asn
420 425 430gtg gct atg gac gtt gct aat gtt gaa ggg aca agc gag tgc aca ggt 1344Val Ala Met Asp Val Ala Asn Val Glu Gly Thr Ser Glu Cys Thr Gly
435 440 445ctt cta act cca cct cca tca aat gat aca act gaa aac ttg ggt aaa 1392Leu Leu Thr Pro Pro Pro Ser Asn Asp Thr Thr Glu Asn Leu Gly Lys
450 455 460gtc gat tta aat gaa acg cct tat gtg caa acg aaa aga atg ctt aca 1440Val Asp Leu Asn Glu Thr Pro Tyr Val Gln Thr Lys Arg Met Leu Thr465 470 475 480cgt atg aaa gcc ctc atg aaa aca gtt gag aca ggt cgg aga tac ttc 1488Arg Met Lys Ala Leu Met Lys Thr Val Glu Thr Gly Arg Arg Tyr Phe
485 490 495cca tct tgt tat gag gtt ctg gat aag tac atg gat cag tat atg gac 1536Pro Ser Cys Tyr Glu Val Leu Asp Lys Tyr Met Asp Gln Tyr Met Asp
500 505 510gaa gaa atc cct gat atg tcg tat ccc gag aaa ggc act gtg aaa gag 1584Glu Glu Ile Pro Asp Met Ser Tyr Pro Glu Lys Gly Thr Val Lys Glu
515 520 525aga aga cag aag agg atg aga tat aac gag ctg aag aac gac gtt aaa 1632Arg Arg Gln Lys Arg Met Arg Tyr Asn Glu Leu Lys Asn Asp Val Lys
530 535 540aaa gca tat agc aaa gac aaa gtc gcg cgg tct tgt ctt tct tct tca 1680Lys Ala Tyr Ser Lys Asp Lys Val Ala Arg Ser Cys Leu Ser Ser Ser545 550 555 560tca cca gct tct tct ctt aga gaa gcc tta gag aat cca aca tga 1725Ser Pro Ala Ser Ser Leu Arg Glu Ala Leu Glu Asn Pro Thr
565 570 575<210>70<211>574<212>PRT<213>鼠耳芥<400>70Met Ala Ala Thr Ala Ile Glu Pro Ser Ser Ser Ile Ser Phe Thr Ser1 5 10 15Ser His Leu Ser Asn Pro Ser Pro Val Val Thr Thr Tyr His Ser Ala
20 25 30Ala Asn Leu Glu Glu Leu Ser Ser Asn Leu Glu Gln Leu Leu Thr Asn
35 40 45Pro Asp Cys Asp Tyr Thr Asp Ala Glu Ile Ile Ile Glu Glu Glu Ala
50 55 60Asn Pro Val Ser Val His Arg Cys Val Leu Ala Ala Arg Ser Lys Phe65 70 75 80Phe Leu Asp Leu Phe Lys Lys Asp Lys Asp Ser Ser Glu Lys Lys Pro
85 90 95Lys Tyr Gln Met Lys Asp Leu Leu Pro Tyr Gly Asn Val Gly Arg Glu
100 105 110Ala Phe Leu His Phe Leu Ser Tyr Ile Tyr Thr Gly Arg Leu Lys Pro
115 120 125Phe Pro Ile Glu Val Ser Thr Cys Val Asp Ser Val Cys Ala His Asp
130 135 140Ser Cys Lys Pro Ala Ile Asp Phe Ala Val Glu Leu Met Tyr Ala Ser145 150 155 160Phe Val Phe Gln Ile Pro Asp Leu Val Ser Ser Phe Gln Arg Lys Leu
165 170 175Arg Asn Tyr Val Glu Lys Ser Leu Val Glu Asn Val Leu Pro Ile Leu
180 185 190Leu Val Ala Phe His Cys Asp Leu Thr Gln Leu Leu Asp Gln Cys Ile
195 200 205Glu Arg Val Ala Arg Ser Asp Leu Asp Arg Phe Cys Ile Glu Lys Glu
210 215 220Leu Pro Leu Glu Val Leu Glu Lys Ile Lys Gln Leu Arg Val Lys Ser225 230 235 240Val Asn Ile Pro Glu Val Glu Asp Lys Ser Ile Glu Arg Thr Gly Lys
245 250 255Val Leu Lys Ala Leu Asp Ser Asp Asp Val Glu Leu Val Lys Leu Leu
260 265 270Leu Thr Glu Ser Asp Ile Thr Leu Asp Gln Ala Asn Gly Leu His Tyr
275 280 285Ala Val Ala Tyr Ser Asp Pro Lys Val Val Thr Gln Val Leu Asp Leu
290 295 300Asp Met Ala Asp Val Asn Phe Arg Asn Ser Arg Gly Tyr Thr Val Leu305 310 315 320His Ile Ala Ala Met Arg Arg Glu Pro Thr Ile Ile Ile Pro Leu Ile
325 330 335Gln Lys Gly Ala Asn Ala Ser Asp Phe Thr Phe Asp Gly Arg Ser Ala
340 345 350Val Asn Ile Cys Arg Arg Leu Thr Arg Pro Lys Asp Tyr His Thr Lys
355 360 365Thr Ser Arg Lys Glu Pro Ser Lys Tyr Arg Leu Cys Ile Asp Ile Leu
370 375 380Glu Arg Glu Ile Arg Arg Asn Pro Leu Val Ser Gly Asp Thr Pro Thr385 390 395 400Cys Ser His Ser Met Pro Glu Asp Leu Gln Met Arg Leu Leu Tyr Leu
405 410 415Glu Lys Arg Val Gly Leu Ala Gln Leu Phe Phe Pro Ala Glu Ala Asn
420 425 430Val Ala Met Asp Val Ala Asn Val Glu Gly Thr Ser Glu Cys Thr Gly
435 440 445Leu Leu Thr Pro Pro Pro Ser Asn Asp Thr Thr Glu Asn Leu Gly Lys
450 455 460Val Asp Leu Asn Glu Thr Pro Tyr Val Gln Thr Lys Arg Met Leu Thr465 470 475 480Arg Met Lys Ala Leu Met Lys Thr Val Glu Thr Gly Arg Arg Tyr Phe
485 490 495Pro Ser Cys Tyr Glu Val Leu Asp Lys Tyr Met Asp Gln Tyr Met Asp
500 505 510Glu Glu Ile Pro Asp Met Ser Tyr Pro Glu Lys Gly Thr Val Lys Glu
515 520 525Arg Arg Gln Lys Arg Met Arg Tyr Asn Glu Leu Lys Asn Asp Val Lys
530 535 540Lys Ala Tyr Ser Lys Asp Lys Val Ala Arg Ser Cys Leu Ser Ser Ser545 550 555 560Ser Pro Ala Ser Ser Leu Arg Glu Ala Leu Glu Asn Pro Thr
565 570<210>71<211>1818<212>DNA<213>鼠耳芥<220><221>CDS<222>(13)..(1818)<223>AtNMLc4-2 cDNA序列<400>71gccgatctcg tg atg atg gcc acc acc acc acc acc acc acc gct aga ttc 51
Met Met Ala Thr Thr Thr Thr Thr Thr Thr Ala Arg Phe
1 5 10tct gat tca tac gag ttc agc aac aca agc ggc aat agc ttc ttc gcc 99Ser Asp Ser Tyr Glu Phe Ser Asn Thr Ser Gly Asn Ser Phe Phe Ala
15 20 25gcc gag tca tct ctt gat tat ccg acg gaa ttt ctc acg cca ccg gag 147Ala Glu Ser Ser Leu Asp Tyr Pro Thr Glu Phe Leu Thr Pro Pro Glu30 35 40 45gta tca gct ctt aaa ctt ctg tct aac tgc ctc gag tct gtt ttc gac 195Val Ser Ala Leu Lys Leu Leu Ser Asn Cys Leu Glu Ser Val Phe Asp
50 55 60tcg ccg gag acg ttc tac agc gat gct aag cta gtt ctc gcc ggc ggc 243Ser Pro Glu Thr Phe Tyr Ser Asp Ala Lys Leu Val Leu Ala Gly Gly
65 70 75cgg gaa gtt tct ttt cac cgt tgt att ctt tcc gcg aga att cct gtc 291Arg Glu Val Ser Phe His Arg Cys Ile Leu Ser Ala Arg Ile Pro Val
80 85 90ttc aaa agc gct tta gcc acc gtg aag gaa caa aaa tcc tcc acc acc 339Phe Lys Ser Ala Leu Ala Thr Val Lys Glu Gln Lys Ser Ser Thr Thr
95 100 105gtg aag ctc cag ctg aaa gag atc gcc aga gat tac gaa gtc ggc ttt 387Val Lys Leu Gln Leu Lys Glu Ile Ala Arg Asp Tyr Glu Val Gly Phe110 115 120 125gac tcg gtt gtg gcg gtt ttg gcg tat gtt tac agc ggc aga gtg agg 435Asp Ser Val Val Ala Val Leu Ala Tyr Val Tyr Ser Gly Arg Val Arg
130 135 140tcc ccg ccg aag gga gct tct gct tgc gta gac gac gat tgt tgc cac 483Ser Pro Pro Lys Gly Ala Ser Ala Cys Val Asp Asp Asp Cys Cys His
145 150 155gtg gct tgc cgg tca aag gtg gat ttc atg gtg gag gtt ctt tat ctg 531Val Ala Cys Arg Ser Lys Val Asp Phe Met Val Glu Val Leu Tyr Leu
160 165 170tct ttc gtt ttc cag att caa gaa tta gtt act ctg tat gag agg cag 579Ser Phe Val Phe Gln Ile Gln Glu Leu Val Thr Leu Tyr Glu Arg Gln
175 180 185ttc ttg gaa att gta gac aaa gtt gta gtc gaa gac atc ttg gtt ata 627Phe Leu Glu Ile Val Asp Lys Val Val Val Glu Asp Ile Leu Val Ile190 195 200 205ttc aag ctt gat act cta tgt ggt aca aca tac aag aag ctt ttg gat 675Phe Lys Leu Asp Thr Leu Cys Gly Thr Thr Tyr Lys Lys Leu Leu Asp
210 215 220aga tgc ata gaa att atc gtg aag tct gat ata gaa cta gtt agt ctt 723Arg Cys Ile Glu Ile Ile Val Lys Ser Asp Ile Glu Leu Val Ser Leu
225 230 235gag aag tct tta cct caa cac att ttc aag caa atc ata gac atc cgc 771Glu Lys Ser Leu Pro Gln His Ile Phe Lys Gln Ile Ile Asp Ile Arg
240 245 250gaa gcg ctc tgt cta gag cca cct aaa cta gaa agg cat gtc aag aac 819Glu Ala Leu Cys Leu Glu Pro Pro Lys Leu Glu Arg His Val Lys Asn
255 260 265ata tac aag gcg cta gac tca gat gat gtt gag ctt gtc aag atg ctt 867Ile Tyr Lys Ala Leu Asp Ser Asp Asp Val Glu Leu Val Lys Met Leu270 275 280 285ttg cta gaa gga cac acc aat ctc gat gag gcg tat gct ctt cat ttt 915Leu Leu Glu Gly His Thr Asn Leu Asp Glu Ala Tyr Ala Leu His Phe
290 295 300gct atc gct cac tgc gct gtg aag acc gcg tat gat ctc ctc gag ctt 963Ala Ile Ala His Cys Ala Val Lys Thr Ala Tyr Asp Leu Leu Glu Leu
305 310 315gag ctt gcg gat gtt aac ctt aga aat ccg agg gga tac act gtg ctt 1011Glu Leu Ala Asp Val Asn Leu Arg Asn Pro Arg Gly Tyr Thr Val Leu
320 325 330cat gtt gct gcg atg cgg aag gag ccg aag ttg ata ata tct ttg tta 1059His Val Ala Ala Met Arg Lys Glu Pro Lys Leu Ile Ile Ser Leu Leu
335 340 345atg aaa ggg gca aat att tta gac aca aca ttg gat ggt aga acc gct 1107Met Lys Gly Ala Asn Ile Leu Asp Thr Thr Leu Asp Gly Arg Thr Ala350 355 360 365tta gtg att gta aaa cga ctc act aaa gcg gat gac tac aaa act agt 1155Leu Val Ile Val Lys Arg Leu Thr Lys Ala Asp Asp Tyr Lys Thr Ser
370 375 380acg gag gac ggt acg cct tct ctg aaa ggc gga tta tgc ata gag gta 1203Thr Glu Asp Gly Thr Pro Ser Leu Lys Gly Gly Leu Cys Ile Glu Val
385 390 395ctt gag cat gaa caa aaa cta gaa tat ttg tcg cct ata gag gct tca 1251Leu Glu His Glu Gln Lys Leu Glu Tyr Leu Ser Pro Ile Glu Ala Ser
400 405 410ctt tct ctt cca gta act cca gag gag ttg agg atg agg ttg ctc tat 1299Leu Ser Leu Pro Val Thr Pro Glu Glu Leu Arg Met Arg Leu Leu Tyr
415 420 425tat gaa aac cga gtt gca ctt gct cga ctt ctc ttt cca gtg gaa act 1347Tyr Glu Asn Arg Val Ala Leu Ala Arg Leu Leu Phe Pro Val Glu Thr430 435 440 445gaa act gta cag ggt att gcc aaa ttg gag gaa aca tgc gag ttt aca 1395Glu Thr Val Gln Gly Ile Ala Lys Leu Glu Glu Thr Cys Glu Phe Thr
450 455 460gct tct agt ctc gag cct gat cat cac att ggt gaa aag cgg aca tca 1443Ala Ser Ser Leu Glu Pro Asp His His Ile Gly Glu Lys Arg Thr Ser
465 470 475Cta gac cta aat atg gcg ccg ttc caa atc cat gag aag cat ttg agt 1491Leu Asp Leu Asn Met Ala Pro Phe Gln Ile His Glu Lys His Leu Ser
480 485 490aga cta aga gca ctt tgt aaa acc gtg gaa ctg ggg aaa cgc tacttc 1539Arg Leu Arg Ala Leu Cys Lys Thr Val Glu Leu Gly Lys Arg Tyr Phe
495 500 505aaa cga tgt tcg ctt gat cac ttt atg gat act gag gac ttg aat cat 1587Lys Arg Cys Ser Leu Asp His Phe Met Asp Thr Glu Asp Leu Asn His510 515 520 525ctt gct agc gta gaa gaa gat act cct gag aaa cgg cta caa aag aag 1635Leu Ala Ser Val Glu Glu Asp Thr Pro Glu Lys Arg Leu Gln Lys Lys
530 535 540caa agg tac atg gaa cta caa gag act ctg atg aag acc ttt agt gag 1683Gln Arg Tyr Met Glu Leu Gln Glu Thr Leu Met Lys Thr Phe Ser Glu
545 550 555gac aag gag gaa tgt gga aag tct tcc aca ccg aaa cca acc tct gcg 1731Asp Lys Glu Glu Cys Gly Lys Ser Ser Thr Pro Lys Pro Thr Ser Ala
560 565 570gtg agg tct aat aga aaa ctc tct cac cgg cgc cta aaa gtg gac aaa 1779Val Arg Ser Asn Arg Lys Leu Ser His Arg Arg Leu Lys Val Asp Lys
575 580 585cgg gat ttt ttg aaa cga cct tac ggg aac ggg gat taa 1818Arg Asp Phe Leu Lys Arg Pro Tyr Gly Asn Gly Asp590 595 600<210>72<211>601<212>PRT<213>鼠耳芥<400>72Met Met Ala Thr Thr Thr Thr Thr Thr Thr Ala Arg Phe Ser Asp Ser1 5 10 15Tyr Glu Phe Ser Asn Thr Ser Gly Asn Ser Phe Phe Ala Ala Glu Ser
20 25 30Ser Leu Asp Tyr Pro Thr Glu Phe Leu Thr Pro Pro Glu Val Ser Ala
35 40 45Leu Lys Leu Leu Ser Asn Cys Leu Glu Ser Val Phe Asp Ser Pro Glu
50 55 60Thr Phe Tyr Ser Asp Ala Lys Leu Val Leu Ala Gly Gly Arg Glu Val65 70 75 80Ser Phe His Arg Cys Ile Leu Ser Ala Arg Ile Pro Val Phe Lys Ser
85 90 95Ala Leu Ala Thr Val Lys Glu Gln Lys Ser Ser Thr Thr Val Lys Leu
100 105 110Gln Leu Lys Glu Ile Ala Arg Asp Tyr Glu Val Gly Phe Asp Ser Val
115 120 125Val Ala Val Leu Ala Tyr Val Tyr Ser Gly Arg Val Arg Ser Pro Pro
130 135 140Lys Gly Ala Ser Ala Cys Val Asp Asp Asp Cys Cys His Val Ala Cys145 150 155 160Arg Ser Lys Val Asp Phe Met Val Glu Val Leu Tyr Leu Ser Phe Val
165 170 175Phe Gln Ile Gln Glu Leu Val Thr Leu Tyr Glu Arg Gln Phe Leu Glu
180 185 190Ile Val Asp Lys Val Val Val Glu Asp Ile Leu Val Ile Phe Lys Leu
195 200 205Asp Thr Leu Cys Gly Thr Thr Tyr Lys Lys Leu Leu Asp Arg Cys Ile
210 215 220Glu Ile Ile Val Lys Ser Asp Ile Glu Leu Val Ser Leu Glu Lys Ser225 230 235 240Leu Pro Gln His Ile Phe Lys Gln Ile Ile Asp Ile Arg Glu Ala Leu
245 250 255Cys Leu Glu Pro Pro Lys Leu Glu Arg His Val Lys Asn Ile Tyr Lys
260 265 270Ala Leu Asp Ser Asp Asp Val Glu Leu Val Lys Met Leu Leu Leu Glu
275 280 285Gly His Thr Asn Leu Asp Glu Ala Tyr Ala Leu His Phe Ala Ile Ala
290 295 300His Cys Ala Val Lys Thr Ala Tyr Asp Leu Leu Glu Leu Glu Leu Ala305 310 315 320Asp Val Asn Leu Arg Asn Pro Arg Gly Tyr Thr Val Leu His Val Ala
325 330 335Ala Met Arg Lys Glu Pro Lys Leu Ile Ile Ser Leu Leu Met Lys Gly
340 345 350Ala Asn Ile Leu Asp Thr Thr Leu Asp Gly Arg Thr Ala Leu Val Ile
355 360 365Val Lys Arg Leu Thr Lys Ala Asp Asp Tyr Lys Thr Ser Thr Glu Asp
370 375 380Gly Thr Pro Ser Leu Lys Gly Gly Leu Cys Ile Glu Val Leu Glu His385 390 395 400Glu Gln Lys Leu Glu Tyr Leu Ser Pro Ile Glu Ala Ser Leu Ser Leu
405 410 415Pro Val Thr Pro Glu Glu Leu Arg Met Arg Leu Leu Tyr Tyr Glu Asn
420 425 430Arg Val Ala Leu Ala Arg Leu Leu Phe Pro Val Glu Thr Glu Thr Val
435 440 445Gln Gly Ile Ala Lys Leu Glu Glu Thr Cys Glu Phe Thr Ala Ser Ser
450 455 460Leu Glu Pro Asp His His Ile Gly Glu Lys Arg Thr Ser Leu Asp Leu465 470 475 480Asn Met Ala Pro Phe Gln Ile His Glu Lys His Leu Ser Arg Leu Arg
485 490 495Ala Leu Cys Lys Thr Val Glu Leu Gly Lys Arg Tyr Phe Lys Arg Cys
500 505 510Ser Leu Asp His Phe Met Asp Thr Glu Asp Leu Asn His Leu Ala Ser
515 520 525Val Glu Glu Asp Thr Pro Glu Lys Arg Leu Gln Lys Lys Gln Arg Tyr
530 535 540Met Glu Leu Gln Glu Thr Leu Met Lys Thr Phe Ser Glu Asp Lys Glu545 550 555 560Glu Cys Gly Lys Ser Ser Thr Pro Lys Pro Thr Ser Ala Val Arg Ser
565 570 575Asn Arg Lys Leu Ser His Arg Arg Leu Lys Val Asp Lys Arg Asp Phe
580 585 590Leu Lys Arg Pro Tyr Gly Asn Gly Asp
595 600<210>73<211>2673<212>DNA<213>烟草<220><221>CDS<222>(661)..(1767)<223>全长烟草B cDNA序列<220><221>misc_特征<222>(1)..(2673)<223>n=a、t、c或g<400>73tcgagcggcc gcccgggcag gtaaactcta acccttttaa tctttttttg gttgcatttc 60ggatctaacc tcaggaaaaa aaacagtatt tttagcctct gcaattgcaa attttctcgt 120ttttttagcc gaagtgaatg ttattccaat tgggtaagct gtgatcaagc agttgaagtt 180ttttgttgca aaatttgcca gttatcttga ctttttgtga agttggtaaa tttttcattt 240gggtaagttg tgatcaagca gttgaagatt tgcactttgt attcttactg tgaaattgca 300gttttgttga ttatagatgg ggtggaattg ttaatttctt ctaaagtttt aaagggttga 360tttggtttta cctgaaatag ggagaatatg acttgtagtt ttggaatttg cttcttttct 420tggtctgcat agttgaatgt tattagaaaa cttatggaaa gttttggtca aacttttgtc 480ctttgagaag aatttcttgt attggtgatt ggttatggtc ttggagaggt tctttttttt 540tttgcataga gcctgtgcgg agaatattat acatggttaa aaacattaga ttttctggac 600tttgactatc ttagatgtag ataaattttg tatatgtttt tagaccatta gaattgggaa 660atg gct tgt tct gct gaa cca tca tca tct ata agc ttt act tca tct 708Met Ala Cys Ser Ala Glu Pro Ser Ser Ser Ile Ser Phe Thr Ser Ser1 5 10 15tcc att aca tcg aat ggg tcg att ggc gtt ggc caa aac act cat gct 756Ser Ile Thr Ser Asn Gly Ser Ile Gly Val Gly Gln Asn Thr His Ala
20 25 30tat ggc ggc tct gag aca ggg agt agt tat gaa atc atc agc ttg agt 804Tyr Gly Gly Ser Glu Thr Gly Ser SerTyr Glu Ile Ile Ser Leu Ser
35 40 45aaa ctc agt aac aat tta gag caa ctc ttg tca gat tcc agc tct gat 852Lys Leu Ser Asn Asn Leu Glu Gln Leu Leu Ser Asp Ser Ser Ser Asp
50 55 60ttt act gat gct gag att gtt gtt gag ggt gtt tca ctt ggt gtt cac 900Phe Thr Asp Ala Glu Ile Val Val Glu Gly Val Ser Leu Gly Val His65 70 75 80cgt tgt ata tta gct gcc agg agt aaa ttt ttt cag gat ctt ttt agg 948Arg Cys Ile Leu Ala Ala Arg Ser Lys Phe Phe Gln Asp Leu Phe Arg
85 90 95aaa gag aag gga agt tgt gga aag gaa ggt aaa cca aga tat tct atg 996Lys Glu Lys Gly Ser Cys Gly Lys Glu Gly Lys Pro Arg Tyr Ser Met
100 105 110acc gat att ttg cct tat ggt aag gtt gga tat gag gct ttc gtt acc 1044Thr Asp Ile Leu Pro Tyr Gly Lys Val Gly Tyr Glu Ala Phe Val Thr
115 120 125ttc cta agc tat ttg tac tca gga aaa ttg aag cat ttc cct ccg gag 1092Phe Leu Ser Tyr Leu Tyr Ser Gly Lys Leu Lys His Phe Pro Pro Glu
130 135 140gta tca aca tgt atg gac act ata tgt gct cat gac tct tgc aga cca 1140Val Ser Thr Cys Met Asp Thr Ile Cys Ala His Asp Ser Cys Arg Pro145 150 155 160gca att aat ttt agt gtg gag ttg atg tat gcc tct tcc atg ttt cag 1188Ala Ile Asn Phe Ser Val Glu Leu Met Tyr Ala Ser Ser Met Phe Gln
165 170 175gtt cca gag cta gta tca ctt ttc ctg aga cgc ctt atc aat ttt gtt 1236Val Pro Glu Leu Val Ser Leu Phe Leu Arg Arg Leu Ile Asn Phe Val
180 185 190ggg aag gct ctt gtg gaa gat gtt atc cca ata ctt aga gtt gct ttt 1284Gly Lys Ala Leu Val Glu Asp Val Ile Pro Ile Leu Arg Val Ala Phe
195 200 205cat tgc caa ttg agc gag ctt ctc act cat tcc gtt gat aga gta gca 1332His Cys Gln Leu Ser Glu Leu Leu Thr His Ser Val Asp Arg Val Ala
210 215 220cga tca gat ctt gaa atc aca tgc att gag aaa gag gtt ccc ttt gaa 1380Arg Ser Asp Leu Glu Ile Thr Cys Ile Glu Lys Glu Val Pro Phe Glu225 230 235 240gtt gca gag aat att aaa tta ttg tgg ccg aaa tgt cag gtt gat gaa 1428Val Ala Glu Asn Ile Lys Leu Leu Trp Pro Lys Cys Gln Val Asp Glu
245 250 255agt aag gtt cta cct gtg gat ccc ttg cat gaa aag aga aaa aat agg 1476Ser Lys Val Leu Pro Val Asp Pro Leu His Glu Lys Arg Lys Asn Arg
260 265 270ata tac aag gca ttg gat tcg gat gat gtt gaa ctt gtc aag ctt cta 1524Ile Tyr Lys Ala Leu Asp Ser Asp Asp Val Glu Leu Val Lys Leu Leu
275 280 285ctg agt gag tct aac ata agc tta gat gaa gcc tac gct ctt cat tat 1572Leu Ser Glu Ser Asn Ile Ser Leu Asp Glu Ala Tyr Ala Leu His Tyr
290 295 300gct gtg gca tat tgt gat ccc aag gtt gtg act gag gtt ctt gga ctg 1620Ala Val Ala Tyr Cys Asp Pro Lys Val Val Thr Glu Val Leu Gly Leu305 310 315 320ggt gtt gcg gat gtc aac cta cgt aat act cgt ggt tac act gtg ctt 1668Gly Val Ala Asp Val Asn Leu Arg Asn Thr Arg Gly Tyr Thr Val Leu
325 330 335cac att gct tcc atg cgt aag gag cca gca gta att gta tcg ctt ttg 1716His Ile Ala Ser Met Arg Lys Glu Pro Ala Val Ile Val Ser Leu Leu
340 345 350act aag gga gct cgt gca tca gag act aca ttg gat ggg cag agt gct 1764Thr Lys Gly Ala Arg Ala Ser Glu Thr Thr Leu Asp Gly Gln Ser Ala
355 360 365gtt agtatctgta ggaggctgac taggcctaag gagtaccatg caaaaacaga 1817Valacaaggccag gaagcaaaca aagatcgggt atgtattgat gttttggaga gagagatgcg 1877tcgcaaccca atggctggag atgcattgtt ttcttcccca atgttggccg atgatctgca 1937catgaaactg cactacctgg aaaatagagt ggcatttgca cggttactgt tccctcttga 1997agccagacta gccatgcaaa ttgcaaatgc tgagactgca gctgaagtag cagtccgttt 2057ggcatctaaa agtacatctg ggaacttgag ggaggttgat ttgaatgaga cacccataaa 2117gcagaaagaa agacttcttt caaggatgca agccctctcg aagacagttg aacttggcaa 2177gcgctatttt ccacattgct ctcaagttct ggacaagttt atggaggatg acttacccga 2237cttaattttc cttgagatgg gccctccaga ggagcaaaag atcaagagga agcgatttaa 2297ggagctcaaa gatgacgttc ancgggcatt taacaaagac aaagctgaac ttcattgctc 2357ccgcttgtcc tcatcatcat gttcctcttc ttttaaagat ggngcaagtg tcaaacttag 2417gaaactatga gtaaataggg ttttgtccta tagtttctct nccatctcag ttttgaatgt 2477aagattaata gtttttataa agacttgtct tgtacancct tcattagagc gcctgctttg 2537tcgctatcca tttccctatt cagcttgtta aacttccatg tttncagtag aaagaaattt 2597gcttaggaac aagcttttgg aatagcttat atggaaaatt gattgtaaaa aaaaaaaaaa 2657aaaaaaaaaa aaaaaa 2673<210>74<211>369<212>PRT<213>烟草<400>74Met Ala Cys Ser Ala Glu Pro Ser Ser Ser Ile Ser Phe Thr Ser Ser1 5 10 15Ser Ile Thr Ser Asn Gly Ser Ile Gly Val Gly Gln Asn Thr His Ala
20 25 30Tyr Gly Gly Ser Glu Thr Gly Ser Ser Tyr Glu Ile Ile Ser Leu Ser
35 40 45Lys Leu Ser Asn Asn Leu Glu Gln Leu Leu Ser Asp Ser Ser Ser Asp
50 55 60Phe Thr Asp Ala Glu Ile Val Val Glu Gly Val Ser Leu Gly Val His65 70 75 80Arg Cys Ile Leu Ala Ala Arg Ser Lys Phe Phe Gln Asp Leu Phe Arg
85 90 95Lys Glu Lys Gly Ser Cys Gly Lys Glu Gly Lys Pro Arg Tyr Ser Met
100 105 110Thr Asp Ile Leu Pro Tyr Gly Lys Val Gly Tyr Glu Ala Phe Val Thr
115 120 125Phe Leu Ser Tyr Leu Tyr Ser Gly Lys Leu Lys His Phe Pro Pro Glu
130 135 140Val Ser Thr Cys Met Asp Thr Ile Cys Ala His Asp Ser Cys Arg Pro145 150 155 160Ala Ile Asn Phe Ser Val Glu Leu Met Tyr Ala Set Ser Met Phe Gln
165 170 175Val Pro Glu Leu Val Ser Leu Phe Leu Arg Arg Leu Ile Asn Phe Val
180 185 190Gly Lys Ala Leu Val Glu Asp Val Ile Pro Ile Leu Arg Val Ala Phe
195 200 205His Cys Gln Leu Ser Glu Leu Leu Thr His Ser Val Asp Arg Val Ala
210 215 220Arg Ser Asp LeuGlu Ile Thr Cys Ile Glu Lys Glu Val Pro Phe Glu225 230 235 240Val Ala Glu Asn Ile Lys Leu Leu Trp Pro Lys Cys Gln Val Asp Glu
245 250 255Ser Lys Val Leu Pro Val Asp Pro Leu His Glu Lys Arg Lys Asn Arg
260 265 270Ile Tyr Lys Ala Leu Asp Ser Asp Asp Val Glu Leu Val Lys Leu Leu
275 280 285Leu Ser Glu Ser Asn Ile Ser Leu Asp Glu Ala Tyr Ala Leu His Tyr
290 295 300Ala Val Ala Tyr Cys Asp Pro Lys Val Val Thr Glu Val Leu Gly Leu305 310 315 320Gly Val Ala Asp Val Asn Leu Arg Asn Thr Arg Gly Tyr Thr Val Leu
325 330 335His Ile Ala Ser Met Arg Lys Glu Pro Ala Val Ile Val Ser Leu Leu
340 345 350Thr Lys Gly Ala Arg Ala Ser Glu Thr Thr Leu Asp Gly Gln Ser Ala
355 360 365Val
Claims (23)
1.一种分离的核酸分子,它包括:
(a)编码SEQ ID NO:2、4、6、8、16、18、20、30、32、34、36、38、40、42、44、46、48、50、52、54、56、58、62、64、66、68、70、72或74的核苷酸序列;
(b)SEQ ID NO:1、3、5、7、15、17、19、29、31、33、35、37、39、41、43、45、47、49、51、53、55、57、61、63、65、67、69、71或73;
(c)包括在序列中与SEQ ID NO:1、3、5、7、15、17、19、29、31、33、35、37、39、41、43、45、47、49、51、53、55、57、61、63、65、67、69、71或73中的至少20个连续碱基对部分相同的至少20个连续碱基对部分的核苷酸序列;
(d)可以使用应用表示为SEQ ID NO:9和10、SEQ ID NO:21和24、SEQ ID NO:22和24、SEQ ID NO:25和28、SEQ ID NO:26和28或SEQ ID NO:59和60的引物对的聚合酶链反应从番茄DNA文库扩增的核苷酸序列;
(e)可以使用应用表示为SEQ ID NO:22和24或SEQ ID NO:26和28的引物对的聚合酶链反应从甜菜DNA文库扩增的核苷酸序列;
(f)可以使用应用表示为SEQ ID NO:26和28的引物对的聚合酶链反应从向日葵DNA文库扩增的核苷酸序列;
(g)可以使用应用表示为SEQ ID NO:21和24、SEQ ID NO:21和23、SEQ ID NO:22和24、SEQ ID NO:25和28或SEQ ID NO:26和28的引物对的聚合酶链反应从马铃薯DNA文库扩增的核苷酸序列;
(h)可以使用应用表示为SEQ ID NO:9和10或SEQ ID NO:26和28的引物对的聚合酶链反应从欧洲油菜DNA文库扩增的核苷酸序列;
(i)可以使用应用表示为SEQ ID NO:13和14、SEQ ID NO:21和24或SEQ ID NO:22和24的引物对的聚合酶链反应从鼠耳芥DNA文库扩增的核苷酸序列;
(j)可以使用应用表示为SEQ ID NO:9和10、SEQ ID NO:11和12、SEQ ID NO:21和24、SEQ ID NO:22和24、SEQ ID NO:25和28或SEQ ID NO:26和28的引物对的聚合酶链反应从烟草DNA文库扩增的核苷酸序列;或
(k)可以使用应用引物对的聚合酶链反应从植物DNA文库扩增的核苷酸序列,所述的引物对包括SEQ ID NO:1、3、5、7、15、17、19、29、31、33、35、37、39、41、43、45、47、49、51、53、55、57、61、63、65、67、69、71或73编码序列(CDS)的前20个核苷酸和最后20个核苷酸的反向补体。
2.一种根据权利要求1的分离的核酸分子,它包括编码SEQ IDNO:2、4、6、8、16、18、20、30、32、34、36、38、40、42、44、46、48、50、52、54、56、58、62、64、66、68、70、72或74的核苷酸序列。
3.一种根据权利要求1的分离的核酸分子,它包括SEQ ID NO:1、3、5、7、15、17、19、29、31、33、35、37、39、41、43、45、47、49、51、53、55、57、61、63、65、67、69、71或73。
4.一种根据权利要求1的分离的核酸分子,它包括一种核苷酸序列,该核苷酸序列包括在序列中与SEQ ID NO:1、3、5、7、15、17、19、29、31、33、35、37、39、41、43、45、47、49、51、53、55、57、61、63、65、67、69、71或73中的至少20个连续碱基对部分相同的至少20个连续碱基对部分。
5.一种根据权利要求1的分离的核酸分子,它包括可以使用应用表示为SEQ ID NO:9和10、SEQ ID NO:21和24、SEQ ID NO:22和24、SEQ ID NO:25和28、SEQ ID NO:26和28或SEQ ID NO:59和60的引物对的聚合酶链反应从番茄DNA文库扩增的核苷酸序列。
6.一种根据权利要求1的分离的核酸分子,它包括可以使用应用表示为SEQ ID NO:22和24或SEQ ID NO:26和28的引物对的聚合酶链反应从甜菜DNA文库扩增的核苷酸序列;
7.一种根据权利要求1的分离的核酸分子,它包括可以使用应用表示为SEQ ID NO:26和28的引物对的聚合酶链反应从向日葵DNA文库扩增的核苷酸序列;
8.一种根据权利要求1的分离的核酸分子,它包括可以使用应用表示为SEQ ID NO:21和24、SEQ ID NO:21和23、SEQ ID NO:22和24、SEQ ID NO:25和28或SEQ ID NO:26和28的引物对的聚合酶链反应从马铃薯DNA文库扩增的核苷酸序列;
9.一种根据权利要求1的分离的核酸分子,它包括可以使用应用表示为SEQ ID NO:9和10或SEQ ID NO:26和28的引物对的聚合酶链反应从欧洲油菜DNA文库扩增的核苷酸序列;
10.一种根据权利要求1的分离的核酸分子,它包括可以使用应用表示为SEQ ID NO:13和14、SEQ ID NO:21和24或SEQ ID NO:22和24的引物对的聚合酶链反应从鼠耳芥DNA文库扩增的核苷酸序列;
11.一种根据权利要求1的分离的核酸分子,它包括可以使用应用表示为SEQ ID NO:9和10、SEQ ID NO:11和12、SEQ ID NO:21和24、SEQ ID NO:22和24、SEQ ID NO:25和28或SEQ ID NO:26和28的引物对的聚合酶链反应从烟草DNA文库扩增的核苷酸序列。
12.一种根据权利要求1的分离的核酸分子,它包括可以使用应用引物对的聚合酶链反应从植物DNA文库扩增的核苷酸序列,所述的引物对相当于SEQ ID NO:1、3、5、7、15、17、19、29、31、33、35、37、39、41、43、45、47、49、51、53、55、57、61、63、65、67、69、71或73编码序列(CDS)的前20个核苷酸和最后20个核苷酸的反向补体。
13.一种包括在植物中具有活性的与上述权利要求中任何一项的核酸分子可操作地连接的启动子的嵌合基因。
14.一种包括权利要求13所述的嵌合基因的重组载体。
15.一种包括权利要求13所述的嵌合基因的宿主细胞。
16.一种包括权利要求13所述的嵌合基因的植物。
17.权利要求16所述的植物,它选自下列植物:水稻、小麦、大麦、黑麦、玉米、马铃薯、canola、向日葵、胡萝卜、甘薯、甜菜、菜豆、豌豆、菊苣、莴苣、甘蓝、花椰菜、嫩茎花椰菜、芜菁、萝卜、菠菜、芦笋、洋葱、大蒜、茄子、胡椒、芹菜、南瓜、南瓜、黄瓜、苹果、梨、温柏、甜瓜、洋李、樱桃、桃、油桃、杏、草莓、葡萄、覆盆子、黑莓、凤梨、鳄梨、番瓜、杧果、香蕉、大豆、烟草、番茄、高粱和糖甘蔗。
18.来自权利要求16的植物的种子。
19.一种增加植物中SAR基因表达的方法,该方法包括在所述植物中表达权利要求13所述的嵌合基因。
20.一种提高植物中病害抗性的方法,该方法包括在所述植物中表达权利要求13所述的嵌合基因。
21.一种PCR引物,它选自SEQ ID NO:9-14、21-28、59和60组成的组。
22.一种用于分离与可导致植物中系统获得性抗性的信号转导级联相关的NIM1同源物的方法,该方法包括使用应用引物对的聚合酶链反应从植物DNA文库扩增DNA分子,所述的引物对相当于SEQ ID NO:1、3、5、7、15、17、19、29、31、33、35、37、39、41、43、45、47、49、51、53、55、57、61、63、65、67、69、71或73编码序列(CDS)的前20个核苷酸和最后20个核苷酸的反向补体;或所述的引物对表示为SEQ ID NO:9和10、SEQ ID NO:11和12、SEQ ID NO:13和14、SEQ ID NO:21和24、SEQ ID NO:22和24、SEQ ID NO:21和23、SEQID NO:25和28、SEQ ID NO:26和28或SEQ ID NO:59和60。
23.权利要求22所述的方法,其中所述的植物DNA文库是烟草、番茄、欧洲油菜、鼠耳芥、甜菜、向日葵或马铃薯的DNA文库。
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KR100423262B1 (ko) * | 2000-12-18 | 2004-03-19 | 세미니스코리아주식회사 | 담배 유래의 Tsi1 유전자를 도입하여 제조된 형질전환균주 |
US8777011B2 (en) | 2001-12-21 | 2014-07-15 | Novartis Ag | Capsule package with moisture barrier |
US20090137390A1 (en) * | 2004-06-30 | 2009-05-28 | Eric Wendell Triplett | Materials and methods for enhancing nitrogen fixation in plants |
KR102583332B1 (ko) * | 2020-11-13 | 2023-09-25 | 서울대학교산학협력단 | 셀룰로오스 합성 유전자 Solyc07g043390이 과발현된 토마토 황화잎말림 바이러스병에 대한 내성이 증진된 형질전환 식물체 |
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WO2000053762A3 (en) | 2001-05-31 |
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