CN101111600A - 产率增加的植物及其制备方法 - Google Patents
产率增加的植物及其制备方法 Download PDFInfo
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Abstract
本发明涉及通过调节植物中滑膜肉瘤转位(SYT)多肽或其同源物编码核酸的表达来增加植物产率的方法。一种这样的方法包括向植物中引入SYT核酸或其变体。本发明也涉及已向其中引入了SYT核酸或其变体的转基因植物,所述植物相对于相应的野生型植物具有增加的产率。本发明也涉及在本发明方法中有用的构建体。
Description
发明领域
本发明总体上涉及分子生物学领域,并且涉及相对于相应的野生型植物而言增加植物产率的方法。更具体地,本发明涉及通过调节植物中编码滑膜肉瘤转位(synovial sarcoma translocation,简称SYT)多肽或其同源物编码核酸的表达来增加植物产率的方法。本发明还涉及SYT多肽或其同源物编码核酸的表达受到调节的植物,所述植物相对于相应的野生型植物具有增加的产率。本发明还提供了在本发明方法中有用的构建体
发明背景
不断增加的世界人口和逐渐减少的农业可用耕地迫使研究朝向提高农业的效率。传统的作物和园艺学改良方法利用选育技术来鉴定具有期望性状的植物。然而,此类选育技术有一些缺陷,即这些技术一般为劳动密集型的,而且产生的植物通常包含异质的遗传组分,当这些异质的遗传组分从亲本植物传递时不一定产生期望的性状。分子生物学的进展已经允许人类修饰动物和植物的种质。植物基因工程需要分离和操作遗传物质(一般以DNA或RNA的形式)以及随后将遗传物质引入植物。这类技术具有传递具多种改良的经济、农业或园艺性状的作物或植物的能力。
在经济上特别让人感兴趣的性状是产率,并且在许多植物的情况下是种子产率。产率通常定义为作物可测量经济价值的产出。这可以以数量和/或质量的方式进行定义。诸如谷物、稻类、小麦、芸苔和大豆的作物占人类总卡路里摄取量的一半以上,不论是通过种子本身的直接消耗,还是通过由加工的种子所饲养的肉类产品的消耗。它们也是工业加工所用的糖类、油类和多类代谢物的来源。种子含有胚和胚乳,前者为种子萌发后新的芽和根的来源,后者为在萌发和幼苗早期生长过程中胚生长的营养源。种子的发育涉及许多基因,并且需要代谢物自根、叶和茎转移至正在生长的种子。特别是胚乳,吸收糖类聚合物、油类和蛋白质的代谢前体,将其合成为贮存高分子,以使谷粒长大。增加植物种子产率的能力,无论是增加种子数量、种子生物量、种子发育、种子饱满性状或任何其他种子相关性状,将在农业中具有许多应用,而且甚至具有许多非农业用途,例如诸如药物、抗体或疫苗等物质的生物技术生产。
产率还可以取决于多种因素,如器官的数量和大小、植物构造(例如,分枝的数量)、种子产量等等。根的发育、营养吸收和胁迫耐受性也是决定产率的重要因素。因此优化上述因素也可以促进作物产率的增加。
现已发现,相对于相应的野生型植物,调节植物中SYT多肽或其同源物编码核酸的表达使植物具有增加的产率。
SYT为转录共激活剂,在植物中,其与GRF(生长调控因子)家族蛋白质的转录激活剂形成功能复合物(Kim HJ,Kende H(2004)Proc Nat AcadSc 101:13374-9)。SYT也成为GIF,为GRF相互作用因子(GRF-interactingfactor)的缩写。CRF转录激活剂与酵母染色质重塑复合物的蛋白SWI/SNF质共享结构域(在N末端区域)(van der Knaap E等,(2000)Plant Phys 122:695-704)。认为这些复合物的转录共激活剂参与将SWI/SNF复合物募集至增强子和启动子区域,以实现局部染色质重塑(Nr AM等综述,(2001)Annu Rev Biochem 70:475-501)。局部染色质结构的改变调节转录激活。更准确地说,认为SYT与植物SWI/SNF复合物相互作用,以实现GRF靶基因的转录激活(Kim HJ,Kende H(2004)Proc Nat Acad Sc 101:13374-9)。
SYT属于拟南芥(Arabidopsis)中由三个成员组成的基因家族。所述SYT多肽与人SYT享有同源性。已显示人SYT多肽为转录共激活剂(Thaete等(1999)Hum Molec Genet 8:585-591)。如下三个结构域表征了哺乳动物SYT多肽:
(i)N末端SNH(SYTN末端同源)结构域,在哺乳动物、植物、线虫和鱼类中是保守的;
(ii)C末端QPGY富含结构域,主要由甘氨酸、脯氨酸、谷酰胺和酪氨酸组成,以不定的间隔出现;
(iii)位于前述两个结构域之间的甲硫氨酸富含(Met富含)结构域。
在植物SYT多肽中,SNH结构域充分保守。C末端结构域富含甘氨酸和谷酰胺,但并不富含脯氨酸或酪氨酸。因此其被命名为QG富含结构域,与哺乳动物的QPGY结构域形成对照。同哺乳动物SYT一样,在QG结构域N末端可以鉴定到Met富含结构域。QG富含结构域可视为基本上是蛋白质C末端的剩余部分(除去SHN结构域);Met富含结构域通常包含在QG富含结构域的头一半之内(从N末端到C末端的方向)。可能有第二Met富含结构域位于植物SYT多肽SNH结构域之前(见图1)。
据报道,丧失功能的SYT突变体以及SYT表达降低的转基因植物发育出小且窄的叶子和花瓣,它们具有较少的细胞(Kim HJ,Kende H(2004)Proc Nat Acad Sc 101:13374-9)。
发明内容
根据本发明,提供了增加植物产率的方法,包括调节植物中SYT多肽或其同源物编码核酸的表达。
文中提及的“相应的野生型植物”应理解为是指任何合适的一株或多株对照植物,所述对照植物的选择完全落在本领域技术人员的能力范围之内,并且可以包括,例如,对应的野生型植物或者不含目的基因的对应植物。文中所用的“对照植物”不仅指整株植物,而且还指植物部分,包括种子和种子部分。
有利地,实施根据本发明的方法产生相对于相应的野生型植物产率增加、特别是种子产率增加的植物。
文中所定义的术语“增加的产率”是指任意一种或多种下列参数的增加,每种都相对于相应的野生型植物而言:(i)植物一个或多个部分,特别是地上(可收获)部分增加的生物量(重量)、增加的根生物量或任何其它可收获部分(如果实、坚果和接荚植物可食种子)增加的生物量;(ii)增加的种子总产率,这包括种子生物量(种子重量)的增加,并且其可以是每棵植株或单粒种子基础上的种子重量增加;(iii)增加的(饱满)种子数量;(iv)增加的种子大小,这也可影响种子的组成;(v)增加的种子体积,这也可影响种子的组成(包括油类、蛋白质和糖类的总含量和组成);(vi)增加的单个种子面积;(vii)增加的种子长度或宽度;(viii)增加的收获指数,其表达为可收获部分如种子的产率与总生物量的比率;和(ix)增加的千粒重(TKW),这通过计数饱满种子数量和它们的总重量外推得到。TKW增加可来自于种子大小和/或种子重量的增加。TKW增加可来自胚大小和/或胚乳大小的增加。种子大小、种子体积、种子面积、种子周长、种子宽度和种子长度的增加可以归因于种子特定部分的增加,例如归因于胚和/或胚乳和/或糊粉和/或盾片或种子其他部分大小的增加。
以谷物为例,产率增加可以表现为下列一种或多种情况:每公顷或每英亩植物数量的增加、每棵植株谷穗数量的增加、行数、行粒数、粒重量、千粒重、谷穗长度/直径的增加,种子充实率(其为饱满种子数除以种子总数并乘以100)的增加,等等。以稻为例,产率增加可以表现为下列一个或多个参数的增加:每公顷或每英亩的植物数量、每棵植株的圆锥花序数量、每个圆锥花序的小穗数量、每个圆锥花序的花(小穗floret)的数量(其表达为饱满种子数占初期(primary)圆锥花序的比率)、种子充实率(其为饱满种子数除以种子总数并乘以100)的增加、千粒重的增加,等等。
产率的增加也可以导致改变的构造,或可以作为改变的构造的结果发生。
根据优选的方面,实施本发明的方法产生具有增加的种子产率的植物。因此,本发明提供了增加植物种子产率的方法,该方法包括调节植物中SYT多肽或其同源物编码核酸的表达。
由于本发明的转基因植物具有增加的产率,相对于相应野生型植物在其生命周期相应阶段的生长速率而言,这些植物可能呈现增加的生长速率(至少在其部分生命周期中)。增加的生长速率可以是对植物的一个或多个部分(包括种子)特异性的,或者可以基本上遍及整株植物。具有增加生长速率的植物可以呈现出早期开花。生长速率的增加可以出现在植物生命周期的一个或多个阶段,或者出现在基本上整个植物生命周期的过程中。在植物生命周期的早期阶段,生长速率的增长可以表现为增强的活力。生长速率的增加可以改变植物的收获周期,使植物能够比其他可能的情况更晚播种和/或更快收获。如果生长速率充分增加,可以允许播种同种植物物种更多的种子(例如完全在一个常规的生长期内,播种和收获稻类植物、接着播种和收获更多的稻类植物)。类似的,如果生长速率充分地增加,可能允许播种不同植物物种更多的种子(例如播种和收获稻类植物,随后,例如,播种和任选的收获大豆、马铃薯或任何其他适合的植物)。在一些作物植物的情况下也可能从同一根茎收获增加的次数。改变植物的收获周期可以导致每英亩年生物量产量的增加(这是由于(比方说在一年中)任何特定植物可以生长和收获次数的增加)。与野生型对应物相比,生长速率的增加还能够在更广阔的地域栽培转基因植物,因为种植农作物的地域限制通常由种植时(早季)或收获时(晚季)不利的环境条件所决定。如果缩短收获周期,可以避免这类不利条件。可以通过来自生长曲线的多种参数确定生长速率,这类参数可以是:T-Mid(植物达到其最大大小的50%所需时间)和T-90(植物达到其最大大小90%所需时间)等等。
实施本发明的方法赋予植物相对于相应野生型植物增加的生长速率。因此,本发明提供了增加植物产率的方法,该方法包括调节植物中SYT多肽或其同源物编码核酸的表达。
无论植物处于无胁迫条件下,或植物相对于适宜的对照植物暴露于多种胁迫下,都发生(种子)产率和/或生长速率的增加。通常植物通过更加缓慢的生长来应答接触的胁迫。在重度胁迫条件下,植物甚至可以完全停止生长。另一方面,轻度胁迫在文中定义为当植物接触时不导致植物完全停止生长丧失重新开始生长能力的任何胁迫。由于耕作方法(灌溉、施肥、杀虫剂处理)的发展,栽培的作物植物常常不会遇到重度胁迫。因此,由轻度胁迫诱导的受损的生长通常成为农业中不期望的因素。轻度胁迫是植物可能接触的典型胁迫。这些胁迫可以是植物接触的日常生物的和/或非生物的(环境的)胁迫。典型的非生物的或环境的胁迫包括由反常的热或冷/冰冻温度产生的温度胁迫;盐胁迫;水胁迫(干旱或过量的水)。化学物质也可以引起非生物胁迫。生物的胁迫一般是由病原体如细菌、病毒、真菌和昆虫引起的那些胁迫。
有利地,可以在任何植物中改良产率。
本文所用术语“植物”包含整株植物、植物的祖先和后代以及植物部分,包括种子、枝条、茎、叶、根(包括块茎)、花以及组织和器官,其中上述每一个都包含目的转基因。术语“植物”也包含植物细胞、悬浮培养物、愈伤组织、胚、分生组织区、配子体、孢子体、花粉和小孢子,同样其中上述每一个都包含转基因。
在本发明方法中尤其有用的植物包括所有属于绿色植物(Viridiplantae)总科的植物,尤其是包括选自下列清单的饲料或饲料荚果、观赏植物、食物作物、树木或灌木的单子叶和双子叶植物,其中包括:金合欢属物种(Acacia spp.)、槭树属物种(Acer spp.)、猕猴桃属物种(Actinidia spp.)、七叶树属物种(Aesculus spp.)、新西兰贝壳杉(Agathisaustralis)、Albixia amara、三色桫椤(Alsophila tricolor)、须芒草属物种(Andropogon spp.)、落花生属物种(Arachis spp)、槟榔(Areca catechu)、Astelia fragrans、黄芪(Astragalus cicer)、Baikiaea plurijuga、桦木属物种(Betula spp.)、芸苔属物种(Brassica spp.)、木榄(Bruguiera gymnorrhiza)、Burkea africana、紫铆(Butea frondosa)、Cadaba farinosa、朱缨花属物种(Calliandra spp)、茶(Camellia sinensis)、美人蕉(Canna indica)、辣椒属物种(Capsicum spp.)、决明属物种(Cassia spp.)、距瓣豆(Centroemapubescens)、木瓜属物种(Chaenomeles spp.)、肉桂(Cinnamomum cassia)、小果咖啡(Coffea arabica)、Colophospermum mopane、变异小冠花(Coronillia varia)、枸子(Cotoneaster serotina)、山楂属物种(Crataegus spp.)、香瓜属物种(Cucumis spp.)、柏木属物种(Cupressus spp.)、Cyathea dealbata、木梨(Cydonia oblonga)、圆球柳杉(Cryptomeria japonica)、香茅属物种(Cymbopogon spp.)、Cynthea dealbata、木梨(Cydonia oblonga)、Dalbergiamonetaria、大叶骨碎补(Davallia divaricata)、山马蝗属物种(Desmodiumspp.)、迪卡兰(Dicksonia squarosa)、Diheteropogon amplectens、Dioclea spp、镰扁豆属物种(Dolichos spp.)、Dorycnium rectum、锥穗稗(Echinochloapyramidalis)、Ehrartia spp.、穇子(Eleusine coracana)、Eragrestis spp.、刺桐属物种(Erythrina spp.)、桉属物种(Eucalyptus spp.)、Euclea schimperi、金茅(Eulalia villosa)、荞麦属物种(Fagopyrum spp.)、费约罗(Feijoasellowiana)、草雷属物种(Fragaria spp.)、千斤拔属物种(Flemingia spp)、Freycinetia banksii、Geranium thunbergii、银杏(Ginkgo biloba)、Glycinejavanica、Gliricidia spp、陆地棉(Gossypium hirsutum)、银桦属物种(Grevilleaspp.)、Guibourtia coleosperma、岩黄芪属物种(Hedysarum spp.)、牛鞭草(Hemarthia altissima)、扭黄茅(Heteropogon contortus)、大麦(Hordeumvulgare)、Hyparrhenia rufa、小连翅(Hypericum erectum)、Hypertheliadissoluta、白花庭蓝(Indigo incarnata)、鸢尾属物种(Iris spp.)、Leptarrhenapyrolifolia、胡枝子属物种(Lespediza spp.)、莴苣属物种(Lettuca spp.)、Leucaena leucocephala、Loudetia simplex、Lotonus bainesii、百脉根属物种(Lotus spp.)、硬皮豆(Macrotyloma axillare)、苹果属物种(Malus spp.)、Manihot esculenta、紫苜蓿(Medicago sativa)、水杉(Metasequoiaglyptostroboides)、大蕉(Musa sapientum)、烟草属物种(Nicotianum spp.)、驴食草属物种(Onobrychis spp.)、Ornithopus spp.、稻属物种(Oryza spp.)、非洲双翼豆(Peltophorum africanum)、狼尾草属物种(Pennisetum spp.)、鳄梨(Persea gratissima)、碧冬茄属物种(Petunia spp.)、菜豆属物种(Phaseolusspp.)、槟榔竹(Phoenix canariensis)、Phormium cookianum、石楠属物种(Photinia spp.)、白云杉(Picea glauca)、松属物种(Pinus spp.)、豌豆(Pisumsativum)、新西兰罗汉松(Podocarpus totara)、Pogonarthria fleckii、Pogonarthria squarrosa、杨属物种(Populus spp.)、牧豆树(Prosopiscineraria)、花旗松(Pseudotsuga menziesii)、Pterolobium stellatum、西洋梨(Pyrus communis)、栎属物种(Quercus spp.)、Rhaphiolepsis umbellata、美味棒花棕(Rhopalostylis sapida)、Rhus natalensis、欧洲醋粟(Ribesgrossularia)、茶藨子属物种(Ribes spp.)、洋槐(Robinia pseudoacacia)、蔷薇属物种(Rosa spp.)、悬钩子属物种(Rubus spp.)、柳属物种(Salix spp.)、Schyzachyrium sanguineum、金松(Sciadopitys verticillata)、北美红杉(Sequoia sempervirens)、巨杉(Sequoiadendron giganteum)、两色蜀黍(Sorghum bicolor)、菠菜属物种(Spinacia spp.)、Sporobolus fimbriatus、Stiburus alopecuroides、Stylosanthos humilis、葫芦茶属物种(Tadehagi spp)、落羽杉(Taxodium distichum)、阿拉伯黄背草(Themeda triandra)、车轴草属物种(Trifolium spp.)、小麦属物种(Triticum spp.)、异叶铁杉(Tsugaheterophylla)、越桔属物种(Vaccinium spp.)、野豌豆属物种(Vicia spp.)、葡萄(Vitis vinifera)、锥穗沃森花(Watsonia pyramidata)、马蹄莲(Zantedeschiaaethiopica)、玉蜀黍(Zea mays)、苋属植物(amaranth)、洋蓟(artichoke)、天门冬属(asparagus)、椰菜(broccoli)、孢子甘蓝(Brussel sprouts)、甘蓝、芸苔(canola)、胡萝卜、花椰菜、芹菜、羽衣甘蓝(collard greens)、亚麻、无头甘蓝(kale)、兵豆属(lentil)、油菜籽油菜(oilseed rape)、秋葵(okra)、洋葱、马铃薯、稻、大豆、草莓、甜菜、甘蔗、向日葵、番茄、南瓜(squash)、茶和藻类,等等。根据本发明优选的实施方案,植物为作物植物。作物植物的实例包括大豆、向日葵、芸苔、苜蓿、油菜籽、棉花、番茄、马铃薯或烟草等等。拟南芥通常不认为食作物植物。进一步优选地,植物为单子叶植物,例如甘蔗。更加优选地,植物是谷类,例如稻、玉米、小麦、大麦、粟、黑麦、高粱或燕麦。
本文定义的术语“SYT多肽或其同源物”指这样的多肽,其从N末端到C末端包含:(i)按照递增的偏好顺序,与SEQ ID NO:2的SNH结构域具有至少40%、45%、50%、55%、60%、65%、70%、75%、80%、85%、90%、91%、92%、93%、94%、95%、96%、97%、98%、99%序列同一性的SNH结构域;和(ii)Met富含结构域;和(iii)QG富含结构域。
优选地,与SEQ ID NO:2的SNH结构域具有至少40%同一性的SNH结构域包含图2中显示为黑色的残基。进一步优选地,SNH结构域为SEQ ID
NO:1所代表。
另外,SYT多肽或其同源物可以包含如下一个或多个序列:(a)SEQ IDNO:90;(b)SEQ ID NO:91;和(c)位于SNH结构域之前的N末端的Met富含结构域。
在酵母双杂交系统中,SYT多肽或其同源物通常与GRF(生长调控因子)多肽相互作用。酵母双杂交相互作用测定在本领域众所周知(见Field等(1989)Nature 340(6230):245-246)。例如,SEQ ID NO:4所代表的SYT多肽能够与AtGRF5并与AtGRF9相互作用。本发明人证明SYT多肽及其同源物能够增加植物产率,特别是种子产率。
SYT多肽或其同源物由SYT核酸/基因编码。因此文中定义的术语“SYT核酸/基因”是编码上文所定义的SYT多肽或其同源物的任何核酸/基因。
可以利用本领域内众所周知的常规技术,如通过序列比对,容易地鉴定SYT多肽或其同源物。序列比对的方法是本领域众所周知的,这类方法包括GAP、BESTFIT、BLAST、FASTA和TFASTA。GAP应用Needleman和Wunsch的算法((1970)J.Mol.Biol.48:443-453)来寻找两个完整序列的比对,使匹配数最大化和空位数最小化。BLAST算法(Altschul等(1990)JMol Biol 215:403-10)计算序列同一性的百分比,并对两序列之间的相似性进行统计分析。执行BLAST分析的软件可从生物技术信息国家中心公开地获得。例如,可利用ClustalW多重序列比对算法(版本1.83)容易地进行鉴定:http://clustalw.genome.ip/sit-bin/nph-clustalw,使用默认的两两比对参数以及百分比的记分方法,来鉴定包含与SEQ ID NO:2的SNH结构域具有至少40%同一性的SNH结构域,和/或包含SEQ ID NO:90和/或SEQ IDNO:91的SYT同源物。与SEQ ID NO:2的SNH结构域具有至少40%同一性的序列足以将其鉴定为SYT。
此外,Met富含结构域或QG富含结构域的存在也可以容易地鉴定。如图3所示,Met富含结构域和QG富含结构域位于SNH结构域之后。QG富含结构域可视为基本上是蛋白质C末端剩余部分(除去SNH结构域);Met富含结构域通常包含在QG富含结构域的头一半之内(从N末端到C末端的方向)。决定多肽结构域是否富含特定氨基酸的一级氨基酸组成(以%表示)可以利用来自ExPASy服务器的软件程序,特别是ProtParam工具进行计算(Gasteiger E等(2003)ExPASy:the proteomics server for in-depth proteinknowledge and analysis.Nucleic Acids Res 31:3784-3788)。然后可以将目的蛋白质的组成与Swiss-Prot蛋白质序列数据库中的平均氨基酸组成(以%表示)进行比较。在该数据库内,平均Met(M)含量为2.37%,平均Gln(Q)含量为3.93%,而平均Gly(G)含量为6.93%。如本文所定义的那样,Met富含结构域或QG富含结构域具有高于Swiss-Prot蛋白质序列数据库平均氨基酸组成(以%表示)的Met含量(以%表示)或Gln和Gly含量(以%表示)。
SYT多肽或其同源物的实例包括(由括号中登录号所示的多核苷酸序列编码,参见表1):拟南芥(Arabidopsis thaliana)Arath_SYT1(AY102639.1)SEQ ID NO:4,拟南芥Arath_SYT2(AY102640.1)SEQ ID NO:6,拟南芥Arath_SYT3(AY102641.1)SEQ ID NO:8,Aspergillus officinalisAspof_SYT(CV287542)SEQ ID NO:10,欧洲油菜(Brassica napus)Brana_SYT(CD823592)SEQ ID NO:12,甜橙(Citrus sinensis)Citsi_SYT(CB290588)SEQ ID NO:14,树棉(Gossypium arboreum)Gosar_SYT(BM359324)SEQ ID NO:16,蒺藜状苜蓿(Medicago trunculata)Medtr_SYT(CA858507.1)SEQ ID NO:18,稻(Oryza sativa)Orysa_SYT1(AK058575)SEQ ID NO:20,稻Orysa_SYT2(AK105366)SEQ ID NO:22,稻Orysa SYT3(BP185008)SEQ ID NO:24,马铃薯(Solanum tuberosum)Soltu_SYT(BG590990)SEQ ID NO:26,玉蜀黍(Zea mays)Zeama_SYT1(BG874129.1,CA409022.1)SEQ ID NO:28,玉蜀黍Zeama_SYT2(AY106697)SEQ ID NO:30,智人(Homo sapiens)Homsa_SYT(CAG46900)SEQ ID NO:32,洋葱(Allium cepa)Allce_SYT2(CF437485)SEQ ID NO:34,Aquilegia formosa x Aquilegia pubescens Aqufo_SYT1(DT758802)SEQID NO:36,二穗短柄草(Brachypodium distachyon)Bradi_SYT3(DV480064)SEQ ID NO:38,欧洲油菜Brana_SYT2(CN732814)SEQ ID NO:40,甜橙Citsi_SYT2(CV717501)SEQ ID NO:42,乳浆大戟(Euphorbia esula)Eupes_SYT2(DV144834)SEQ ID NO:44,大豆(Glycine max)Glyma_SYT2(BQ612648)SEQ ID NO:46,野大豆(Glycine soya)Glyso_SYT2(CA799921)SEQ ID NO:48,陆地棉(Gossypium hirsutum)Goshi_SYT1(DT558852)SEQ ID NO:50,陆地棉Goshi_SYT2(DT563805)SEQ ID NO:52,大麦(Hordeum vulgare)Horvu_SYT2(CA032350)SEQ IDNO:54,野莴苣(Lactuca serriola)Lacse_SYT2(DW110765)SEQ ID NO:56,番茄(Lycopersicon esculentum)Lyces_SYT1(AW934450,BP893155)SEQ ID NO:58,驯化苹果(Malus domestica)Maldo_SYT2(CV084230,DR997566)SEQ ID NO:60,蒺藜状苜蓿Medtr_SYT2(CA858743,BI310799,AL382135)SEQ ID NO:62,柳枝稷(Panicum virgatum)Panvi_SYT3(DN152517)SEQ ID NO:64,北美云杉(Picea sitchensis)Picsi_SYT1(DR484100,DR478464)SEQ ID NO:66,火炬松(Pinus taeda)Pinta_SYT1(DT625916)SEQ ID NO:68,欧洲山杨(Populus tremula)Poptr_SYT1(DT476906)SEQ ID NO:70,甘蔗(Saccharum officinarum)Sacof_SYT1(CA078249,CA078630,CA082679,CA234526,CA239244,CA083312)SEQ ID NO:72,甘蔗Sacof_SYT2(CA110367)SEQ ID NO:74,甘蔗Sacof_SYT3(CA161933,CA265085)SEQ ID NO:76,马铃薯Soltu_SYT1(CK265597)SEQ ID NO:78,两色蜀黍(Sorghum bicolor)Sorbi_SYT3(CX611128)SEQ ID NO:80,普通小麦(Triticum aestivum)Triae_SYT2(CD901951)SEQ ID NO:82,普通小麦Triae_SYT3(BJ246754,BJ252709)SEQ ID NO:84,葡萄(Vitis vinifera)Vitvi_SYT1(DV219834)SEQ ID NO:86,玉蜀黍Zeama_SYT3(CO468901)SEQ IDNO:88。
表1:SYT同源物的实例
名称 | NCBI核苷酸登录号 | 核苷酸SEQ ID NO | 翻译多肽SEQ ID NO | 来源 |
Arath_SYT1 | AY102639.1 | 3 | 4 | 拟南芥 |
Arath_SYT2 | AY102640.1 | 5 | 6 | 拟南芥 |
Arath_SYT3 | AY102641.1 | 7 | 8 | 拟南芥 |
Aspof_SYT1 | CV287542 | 9 | 10 | Aspergillus officinalis |
Brana_SYT1 | CD823592 | 11 | 12 | 欧洲油菜 |
Citsi_SYT1 | CB290588 | 13 | 14 | 甜橙 |
Gosar_SYT1 | BM359324 | 15 | 16 | 树棉 |
Medtr_SYT1 | CA858507.1 | 17 | 18 | 蒺藜状苜蓿 |
Orysa_SYT1 | AK058575 | 19 | 20 | 稻 |
Orysa_SYT2 | AK105366 | 21 | 22 | 稻 |
Orysa_SYT3 | BP185008 | 23 | 24 | 稻 |
Soltu_SYT2 | BG590990 | 25 | 26 | 马铃薯 |
Zeama_SYT1 | BG874129.1CA409022.1* | 27 | 28 | 玉蜀黍 |
Zeama_SYT2 | AY106697 | 29 | 30 | 玉蜀黍 |
Homsa_SYT | CR542103 | 31 | 32 | 智人 |
Allce_SYT2 | CF437485 | 33 | 34 | 洋葱 |
Aqufo_SYT1 | DT758802.1 | 35 | 36 | Aquilegia formosa xAquilegia pubescens |
Bradi_SYT3 | DV480064.1 | 37 | 38 | 二穗短柄草 |
Brana_SYT2 | CN732814 | 39 | 40 | 欧洲油菜 |
Citsi_SYT2 | CV717501 | 41 | 42 | 甜橙 |
Eupes_SYT2 | DV144834 | 43 | 44 | 乳浆大戟 |
Glyma_SYT2 | BQ612648 | 45 | 46 | 大豆 |
Glyso_SYT2 | CA799921 | 47 | 48 | 野大豆 |
Goshi_SYT1 | DT558852 | 49 | 50 | 陆地棉 |
Goshi_SYT2 | DT563805 | 51 | 52 | 陆地棉 |
Horvu_SYT2 | CA032350 | 53 | 54 | 大麦 |
Lacse_SYT2 | DW110765 | 55 | 56 | 野莴苣 |
Lyces_SYT1 | AW934450.1BP893155.1* | 57 | 58 | 番茄 |
Maldo_SYT2 | CV084230DR997566* | 59 | 60 | 驯化苹果 |
Medtr_SYT2 | CA858743BI310799.1AL382135.1* | 61 | 62 | 蒺藜状苜蓿 |
Panvi_SYT3 | DN152517 | 63 | 64 | 柳枝稷 |
Picsi_SYT1 | DR484100DR478464.1 | 65 | 66 | 北美云杉 |
Pinta_SYT1 | DT625916 | 67 | 68 | 火炬松 |
Poptr_SYT1 | DT476906 | 69 | 70 | 欧洲山杨 |
Sacof_SYT1 | CA078249.1CA078630CA082679CA234526CA239244CA083312* | 71 | 72 | 甘蔗 |
Sacof_SYT2 | CA110367 | 73 | 74 | 甘蔗 |
Sacof_SYT3 | CA161933.1CA265085* | 75 | 76 | 甘蔗 |
Soltu_SYT1 | CK265597 | 77 | 78 | 马铃薯 |
Sorbi_SYT3 | CX611128 | 79 | 80 | 两色蜀黍 |
Triae_SYT2 | CD901951 | 81 | 82 | 普通小麦 |
Triae_SYT3 | BJ246754BJ252709* | 83 | 84 | 普通小麦 |
VItvi_SYT1 | DV219834 | 85 | 86 | 葡萄 |
Zeama_SYT3 | CO468901 | 87 | 88 | 玉蜀黍 |
*由所引的登录号拼接
应该理解的是,归入“SYT多肽或其同源物”定义的序列不限于由SEQ ID NO:4、SEQ ID NO:6、SEQ ID NO:8、SEQ ID NO:10、SEQ IDNO:12、SEQ ID NO:14、SEQ ID NO:16、SEQ ID NO:18、SEQ ID NO:20、SEQ ID NO:22、SEQ ID NO:24、SEQ ID NO:26、SEQ ID NO:28、SEQ ID NO:30、SEQ ID NO:32、SEQ ID NO:34、SEQ ID NO:36、SEQID NO:38、SEQ ID NO:40、SEQ ID NO:42、SEQ ID NO:44、SEQ ID NO:46、SEQ ID NO:48、SEQ ID NO:50、SEQ ID NO:52、SEQ ID NO:54、SEQ ID NO:56、SEQ ID NO:58、SEQ ID NO:60、SEQ ID NO:62、SEQID NO:64、SEQ ID NO:66、SEQ ID NO:68、SEQ ID NO:70、SEQ ID NO:72、SEQ ID NO:74、SEQ ID NO:76、SEQ ID NO:78、SEQ ID NO:80、SEQ ID NO:82、SEQ ID NO:84、SEQ ID NO:86、SEQ ID NO:88所代表的序列,而是任何这样的的多肽,其从N末端到C末端包含:(i)与SEQ IDNO:2的SNH结构域具有至少40%同一性的SNH结构域;和(ii)Met富含结构域;和(iii)QG富含结构域,均适用于实施本发明的方法。
SYT核酸的实例包括SEQ ID NO:3、SEQ ID NO:5、SEQ ID NO:7、SEQ ID NO:9、SEQ ID NO:11、SEQ ID NO:13、SEQ ID NO:15、SEQID NO:17、SEQ ID NO:19、SEQ ID NO:21、SEQ ID NO:23、SEQ ID NO:25、SEQ ID NO:27、SEQ ID NO:29、SEQ ID NO:31、SEQ ID NO:33、SEQ ID NO:35、SEQ ID NO:37、SEQ ID NO:39、SEQ ID NO:41、SEQID NO:43、SEQ ID NO:45、SEQ ID NO:47、SEQ ID NO:49、SEQ ID NO:51、SEQ ID NO:53、SEQ ID NO:55、SEQ ID NO:57、SEQ ID NO:59、SEQ ID NO:61、SEQ ID NO:63、SEQ ID NO:65、SEQ ID NO:67、SEQID NO:69、SEQ ID NO:71、SEQ ID NO:73、SEQ ID NO:75、SEQ ID NO:77、SEQ ID NO:79、SEQ ID NO:81、SEQ ID NO:83、SEQ ID NO:85、SEQ ID NO:87中任一所代表的那些序列。SYT核酸/基因及其变体可适于实施本发明的方法。SYT核酸/基因的变体通常为与天然存在的SYT核酸/基因具有相同功能的那些核酸/基因,其可以是相同的生物学功能,或者是当植物中所述核酸/基因的表达受到调节时增加产率的功能。这样的变体包括下文所述的SYT核酸/基因的部分和/或能与SYT核酸/基因杂交的核酸。
本文所定义的术语“部分”指编码这样的多肽的DNA片段,所述多肽从N末端到C末端包含:(i)按照递增的偏好顺序,与SEQ ID NO:2的SNH结构域具有至少40%、45%、50%、55%、60%、65%、70%、75%、80%、85%、90%、91%、92%、93%、94%、95%、96%、97%、98%、99%序列同一性的SNH结构域;和(ii)Met富含结构域;和(iii)QG富含结构域。可以,例如,通过在SYT核酸中产生一个或多个缺失来制备部分。部分可以以分离的形式应用,或者它们可以与其它编码(或非编码)序列融合以,例如,产生组合数种活性的蛋白质。当与其它编码序列融合时,翻译产生的多肽可以大于预测的SYT片断。优选地,部分是由以下任一所代表核酸的部分:SEQ ID NO:3、SEQ ID NO:5、SEQ ID NO:7、SEQ ID NO:9、SEQ ID NO:11、SEQ ID NO:13、SEQ ID NO:15、SEQ ID NO:17、SEQID NO:19、SEQ ID NO:21、SEQ ID NO:23、SEQ ID NO:25、SEQ ID NO:27、SEQ ID NO:29、SEQ ID NO:31、SEQ ID NO:33、SEQ ID NO:35、SEQ ID NO:37、SEQ ID NO:39、SEQ ID NO:41、SEQ ID NO:43、SEQID NO:45、SEQ ID NO:47、SEQ ID NO:49、SEQ ID NO:51、SEQ ID NO:53、SEQ ID NO:55、SEQ ID NO:57、SEQ ID NO:59、SEQ ID NO:61、SEQ ID NO:63、SEQ ID NO:65、SEQ ID NO:67、SEQ ID NO:69、SEQID NO:71、SEQ ID NO:73、SEQ ID NO:75、SEQ ID NO:77、SEQ ID NO:79、SEQ ID NO:81、SEQ ID NO:83、SEQ ID NO:85、SEQ ID NO:87。最优选SEQ ID NO:3、SEQ ID NO:5或SEQ ID NO:7所代表核酸的部分。
其他SYT核酸/基因变体是在降低的严格条件下,优选在严格条件下,能够与上文所定义的SYT核酸/基因杂交的核酸,该杂交序列所编码的多肽从N末端到C末端包含:(i)按照递增的偏好顺序,与SEQ ID NO:2的SNH结构域具有至少40%、45%、50%、55%、60%、65%、70%、75%、80%、85%、90%、91%、92%、93%、94%、95%、96%、97%、98%、99%序列同一性的SNH结构域;和(ii)Met富含结构域;和(iii)QG富含结构域。优选杂交序列能与由SEQ ID NO:3、SEQ ID NO:5、SEQ ID NO:7、SEQID NO:9、SEQ ID NO:11、SEQ ID NO:13、SEQ ID NO:15、SEQ ID NO:17、SEQ ID NO:19、SEQ ID NO:21、SEQ ID NO:23、SEQ ID NO:25、SEQ ID NO:27、SEQ ID NO:29、SEQ ID NO:31、SEQ ID NO:33、SEQID NO:35、SEQ ID NO:37、SEQ ID NO:39、SEQ ID NO:41、SEQ ID NO:43、SEQ ID NO:45、SEQ ID NO:47、SEQ ID NO:49、SEQ ID NO:51、SEQ ID NO:53、SEQ ID NO:55、SEQ ID NO:57、SEQ ID NO:59、SEQID NO:61、SEQ ID NO:63、SEQ ID NO:65、SEQ ID NO:67、SEQ ID NO:69、SEQ ID NO:71、SEQ ID NO:73、SEQ ID NO:75、SEQ ID NO:77、SEQ ID NO:79、SEQ ID NO:81、SEQ ID NO:83、SEQ ID NO:85、SEQID NO:87任一所代表的核酸杂交,或与上文所定义的任何上述序列的部分杂交。最优选SEQ ID NO:3、SEQ ID NO:5或SEQ ID NO:7所代表核酸的杂交序列。
本文定义的术语“杂交”指其中基本同源互补的核酸序列彼此退火的过程。杂交过程能够完全在溶液中发生,即互补的核酸都在溶液中。杂交过程也可以在如下情况下进行,即互补核酸之一固定于基质上,如磁珠、琼脂糖珠或任何其它树脂上。此外,杂交过程也可以如此进行,即其中互补核酸之一固定在固相支持物如硝酸纤维素或尼龙膜上,或者如用照相平板印刷固定在例如硅质玻璃支持物上(后者称为核酸阵列或微阵列,或称为核酸芯片)。为了使杂交发生,通常使核酸分子热变性或化学变性,以使双链解链成两条单链,和/或除去单链核酸中的发夹结构或其它二级结构。杂交的严格性受诸如温度、盐浓度、离子强度和杂交缓冲液组成等条件的影响。
在核酸杂交实验如Southern和Northern杂交的情况中,“严格杂交条件”和“严格杂交洗涤条件”依赖于序列,并且在不同的环境参数下不同。熟练的技术人员知晓可以在杂交和洗涤过程中改变的多种参数,从而保持或者改变严格条件。
Tm是在确定的离子强度和pH值下,50%的靶序列与完全匹配的探针杂交的温度。Tm取决于溶液条件和探针的碱基组成及长度。例如,较长的序列在较高温度下特异性杂交。在低于Tm值16℃到32℃获得最大杂交率。在杂交溶液中存在一价阳离子会减少两核酸链之间的静电排斥作用,从而促进杂合体形成;当钠浓度高达0.4M时,这一作用明显。每个百分点的甲酰胺可使DNA-DNA和DNA-RNA双链体的解链温度降低0.6到0.7℃,加入50%甲酰胺能够使杂交在30到45℃完成,尽管这将降低杂交率。碱基对错配降低杂交率和双链体的热稳定性。平均而言,对于大的探针,每个百分点碱基错配使Tm值下降约1℃。Tm值可以用依赖于杂合体类型的下列方程式计算:
1、DNA-DNA杂合体(Meinkoth和Wahl,Anal.Biochem.,138:267-284,1984):
Tm=81.5℃+16.6×log[Na+]a+0.41×%[G/Cb]-500×[Lc]-1-0.61×%甲酰胺
2、DNA-RNA或RNA-RNA杂合体:
Tm=79.8+18.5(log10[Na+]a)+0.58(%G/Cb)+11.8(%G/Cb)2-820/Lc
3、寡DNA或寡RNAd杂合体:
<20个核苷酸:Tm=2(ln)
20-35个核苷酸:Tm=22+1.46(ln)
a或对于其它一价阳离子,但是仅在0.01-0.4M范围内精确。
b仅对于在30%到75%范围内的%GC精确。
cL=双链体的碱基对长度。
d寡,寡核苷酸;ln,引物的有效长度=2×(G/C数)+(A/T数)。
注释:对于每1%甲酰胺,Tm值降低约0.6到0.7℃,而6M尿素的存在可使Tm值降低约30℃。
杂交特异性通常是杂交后洗涤的函数。为了除去非特异杂交产生的背景,用稀释的盐溶液洗涤样品。这类洗涤的关键因素包括最终洗涤溶液的离子强度和温度:盐浓度越低、洗涤温度越高,洗涤的严格性就越高。洗涤条件通常在等于或低于杂交严格性条件下进行。一般地,如上设置适用于核酸杂交测定或基因扩增检测操作的严格条件。也可以选择更高或更低的严格性条件。通常,对于在确定的离子强度和pH值下的特定序列,选择比热解链温度(Tm)低50℃的低严格条件。中等严格条件温度比Tm低20℃,而高严格条件温度比Tm低10℃。例如,严格条件是至少像如条件A-L一样严格;降低的严格条件是至少像如条件M-R一样严格。可以通过许多已知技术中的任一来控制非特异性结合,所述技术诸如,例如用含蛋白质的溶液封闭膜,在杂交缓冲液中添加异源RNA、DNA和SDS,以及用RNA酶处理。在下表2中列出杂交和洗涤条件的实例。
表2:杂交和洗涤条件的实例
严格条件 | 多核苷酸杂合体± | 杂合体长度(bp) | 杂交温度和缓冲液 | 洗涤温度和缓冲液 |
A | DNA∶DNA | >或等于50 | 65℃1×SSC;或42℃,1×SSC和50%甲酰胺 | 65℃;0.3×SSC |
B | DNA∶DNA | <50 | Tb*;1×SSC | Tb*;1×SSC |
C | DNA∶RNA | >或等于50 | 67℃1×SSC;或45℃,1×SSC和50%甲酰胺 | 67℃;0.3×SSC |
D | DNA∶RNA | <50 | Td*;1×SSC | Td*;1×SSC |
E | RNA∶RNA | >或等于50 | 70℃1×SSC;或50℃,1×SSC和50%甲酰胺 | 70℃;0.3×SSC |
F | RNA∶RNA | <50 | Tf*;1×SSC | Tf*;1×SSC |
G | DNA∶DNA | >或等于50 | 65℃4×SSC;或45℃,4×SSC和50%甲酰胺 | 65℃;1×SSC |
H | DNA∶DNA | <50 | Th*;4×SSC | Th*;4×SSC |
I | DNA∶RNA | >或等于50 | 67℃4×SSC;或45℃,4×SSC和50%甲酰胺 | 67℃;1×SSC |
J | DNA∶RNA | <50 | Tj*;4×SSC | Tj*;4×SSC |
K | RNA∶RNA | >或等于50 | 70℃4×SSC;或40℃,6×SSC和50%甲酰胺 | 67℃;1×SSC |
L | RNA∶RNA | <50 | Tl*;2×SSC | Tl*;2×SSC |
M | DNA∶DNA | >或等于50 | 50℃4×SSC;或40℃,6×SSC和50%甲酰胺 | 50℃;2×SSC |
N | DNA∶DNA | <50 | Tn*;6×SSC | Tn*;6×SSC |
O | DNA∶RNA | >或等于50 | 55℃4×SSC;或42℃,6×SSC和50%甲酰胺 | 55℃;2×SSC |
P | DNA∶RNA | <50 | Tp*;6×SSC | Tp*;6×SSC |
Q | RNA∶RNA | >或等于50 | 60℃4×SSC;或45℃,6×SSC和50%甲酰胺 | 60℃;2×SSC |
R | RNA∶RNA | <50 | Tr*;4×SSC | Tr*;4×SSC |
“杂合体长度”是杂交核酸的预期长度。当已知序列的核酸杂交时,可以通过比对序列及鉴别本文所述的保守区域来确定杂合体长度。
在杂交和洗涤缓冲液中,可以用SSPE(1×SSPE是0.15M NaCl,10mM NaH2PO4和1.25mM EDTA,pH7.4)代替SSC(1×SSC是0.15M NaCl和15mM柠檬酸钠);杂交完成后洗涤15分钟。杂交和洗涤可以另外地包括5×Denhardt′s试剂、0.5-1.0%SDS、100μg/ml变性片段化的鲑精DNA、0.5%焦磷酸钠和高达50%的甲酰胺。
*Tb-Tr:对于预期长度小于50个碱基对的杂合体,杂交温度应该比杂合体的解链温度Tm低5-10℃;根据上述方程式确定Tm。
±本发明还包括以PNA或修饰的核酸代替任一或多个DNA或RNA杂交配偶体。
为了定义严格性水平,可以参考Sambrook等(2001)的《分子克隆:实验室手册》,第三版,冷泉港实验室出版,冷泉港,纽约,或者CurrentProtocols in Molecular Biology,John Wiley&Sons,N.Y.(1989)。
SYT核酸或其变体可以来自任何天然或人工的来源,如植物、藻类或动物。可以通过仔细的人为操作在组成和/或基因组环境上修饰所述核酸的天然形式。优选植物来源的核酸,无论来源于同一植物物种(例如对于其被引入的物种而言)还是来源于不同植物物种。优选植物来源的核酸编码SYT1。或者,核酸可以编码SYT2或SYT3,它们在多肽水平上彼此密切相关。可以从双子叶植物物种,优选从十字花科(Brassicaceae),更优选从拟南芥分离所述核酸。更优选地,从拟南芥中分离的三种SYT核酸表示为SEQID NO:3、SEQ ID NO:5和SEQ ID NO:7,并且三种SYT氨基酸序列为SEQ ID NO:4、SEQ ID NO:6和SEQ ID NO:8所代表。
可以通过引入遗传修饰(优选在SYT基因座)调节SYT多肽或其同源物编码核酸的表达。本文所定义的基因座意指基因组区,其包括目的基因和编码区上游或下游的10kb。
例如,可以通过任一(或多个)如下方法引入遗传修饰:T-DNA激活、TILLING、定点诱变、定向进化和同源重组,或通过在植物中引入和表达编码SYT多肽或其同源物的核酸。引入遗传修饰之后的步骤是选择SYT多肽或其同源物编码核酸受调节的表达,所述受调节的表达使植物产率增加,特别是种子产率增加。
T-DNA激活标记(Hayashi等Science(1992)1350-1353)包括将通常含有启动子(也可以是翻译增强子或内含子)的T-DNA插入在目的基因的基因组区或基因编码区上游或下游10kb,从而在构型上使启动子能够指导靶向基因的表达。通常天然启动子对靶向基因表达的调控被破坏,基因由新引入的启动子控制。启动子一般包含于T-DNA中。例如,通过农杆菌(Agrobacterium)感染将此T-DNA随机插入植物基因组并导致在插入T-DNA附近的基因过表达。得到的转基因植物由于引入的启动子附近基因过表达而表现出显性表型。引入的启动子可以是任意能够在期望生物体内(在本案中是植物)指导基因表达的启动子。例如,组成型的、组织偏好的、细胞类型偏好的和诱导型的启动子都适用于T-DNA激活。
也可以通过TILLING(定向诱导的基因组局部突变)技术将遗传修饰引入SYT基因座。这是一种诱变技术,其用于产生和/或鉴定并最终分离诱变的编码具有增强SYT活性之蛋白质的SYT核酸变体。TILLING还允许选择携带此种突变变体的植物。这些突变变体甚至可能比其天然形式基因呈现更高的SYT活性。TILLING结合了高密度诱变和高通量筛选方法。TILLING一般遵循的步骤有:(a)EMS诱变(Redei GP和Koncz C,(1992)InMethods in Arabidopsis Research,Koncz C,Chu a NH,Schell J编辑,新加坡,World Scientific Publishing Co,第16-82页;Feldmann等,(1994)InMeyerowitz EM,Somerville CR编辑卷,Arabidopsis.冷泉港实验室出版社冷泉港,纽约,第137-172页;Lightner J和Caspar T,(1998)In JMartinez-Zapater,J Salinas编辑,Methods on Molecular Biology,82卷Humana Press,Totowa,NJ,第91-104页);(b)DNA制备和个体合并;(c)目的区域的PCR扩增;(d)变性和退火以形成杂双链体;(e)DHPLC,其中库中存在的杂双链体会在色谱图上检测到额外的峰;(f)突变个体的鉴定;和(g)突变PCR产物的测序。TILLING的方法是本领域众所周知的(McCallum等(2002)Nat Biotechnol 18:455-457,由Stemple综述(2004)Nat Rev Genet5(2):145-50)。
定点诱变可用于产生SYT核酸的变体。可以通过几种方法来完成定点诱变,最常见的是基于PCR的方法(current protocols in molecular biology.Wiley编辑http://www.4ulr.com/products/currentprotocols/index.html)。
定向进化也可以用于产生SYT核酸的变体。这包括DNA改组的重复,继之以适当筛选和/或选择,以产生编码具有修饰的生物活性多肽的SYT核酸的变体或其部分(Castle等(2004)Science 304(5674):1151-4;美国专利5,811,238和6,395,547)。
TDNA激活、TILLING、定点诱变和定向进化是能够产生新的SYT等位基因和变体的技术的实例。
同源重组允许向基因组中的指定选择位置引入所选的核酸。同源重组是生物科学中常规使用的标准技术,其用于低等有机体如酵母或小立碗藓(physcomitrella)。在植物中执行同源重组的方法已经不仅在模式植物中描述(Offringa等(1990)EMBO J.9(10):3077-84),而且也在作物植物,如稻中描述(Terada等(2002)Nat Biotech 20(10):1030-4;Iida和Terada(2004)Curr Opin Biotechnol 15(2):132-8)。所靶向的核酸(其可能是上文中定义的SYT核酸或其变体)靶向SYT基因座。所靶向的核酸可以是改良的等位基因,其用于替换内源基因或额外引入到内源基因中。
引入遗传修饰(在本案中不需引入SYT基因座中)的优选方法是在植物中引入和表达编码SYT多肽或其同源物的核酸。SYT多肽或其同源物定义为这样的多肽,其从N末端到C末端包含:(i)按照递增的偏好顺序,与SEQID NO:2的SNH结构域具有至少40%、45%、50%、55%、60%、65%、70%、75%、80%、85%、90%、91%、92%、93%、94%、95%、96%、97%、98%、99%序列同一性的SNH结构域;和(ii)Met富含结构域;和(iii)QG富含结构域。
优选地,与SEQ ID NO:2的SNH结构域具有至少40%同一性的SNH结构域包含图2中显示为黑色的残基。进一步优选地,SNH结构域为SEQ IDNO:1所代表。
欲引入植物的核酸可以是全长的核酸,或者是上述定义的部分或杂交序列。
蛋白质的“同源物”包括肽、寡肽、多肽、蛋白质和酶,其相对于所讨论的未修饰蛋白质具有氨基酸取代、缺失和/或插入,并且具有与其衍生自的未修饰形式蛋白质相似的生物活性和功能活性。为了产生这样的同源物,蛋白质的氨基酸由具有相似性质(如相似的疏水性、亲水性、抗原性、形成或打破α螺旋结构或β片层结构的倾向)的其它氨基酸所替换。保守取代表是本领域内众所周知的(例如见Creighton(1984)Proteins.W.H.Freeman and Company和下表3)。
同源物包括直系同源物(orthologue)和旁系同源物(paralogue),其涵盖用于描述基因祖先关系的进化概念。旁系同源物为相同物种内的基因,其源自于某祖先基因的复制;而直系同源物为来自不同生物体的基因,其起源于物种形成。
例如,可以通过所谓的交互(reciprocal)blast搜索容易地找到例如在单子叶植物种中的直系同源物。这可以通过使用查询序列(例如SEQ ID NO:3、SEQ ID NO:4、SEQ ID NO:5、SEQ ID NO:6、SEQ ID NO:7或SEQID NO:8)针对任何序列数据库,如可在http://www.ncbi.nlm.nih.gov找到的公共可获得的NCBI数据库,进行一次blast而实现。当从核苷酸序列开始时,可使用BLASTN或tBLASTX(利用标准默认值),而当从蛋白质开始时,可使用BLASTP或TBLASTN(利用标准默认值)。Blast结果可以任选地过滤。接着使用过滤的结果或者未过滤的结果中的全长序列针对查询序列源自的相同生物体的序列进行反向blast(二次blast)(在查询序列为SEQ IDNO:3、SEQ ID NO:4、SEQ ID NO:5、SEQ ID NO:6、SEQ ID NO:7或SEQ ID NO:8的情况下,二次blast将会针对稻序列)。然后比较第一次和第二次blast的结果。如果二次blast中得分靠前的命中事件来自查询序列源自的相同物种,则找到了旁系同源物;如果二次blast中得分靠前的命中事件不是来自查询序列源自的相同物种,则找到了直系同源物。得分靠前的命中事件是E值低的命中事件。E值越低,得分越具有显著性(或者换句话说,偶然发现此命中事件的几率越低)。E值的计算是本领域众所周知的。在大家族的情况下可以使用ClustalW,继之以邻近连接树来辅助相关基因的聚类可视化,以鉴定直系同源物和旁系同源物。
同源物可以是蛋白质“取代变体”的形式,即在氨基酸序列中至少有一个残基被除去,并在这一位置插入不同的残基。氨基酸取代通常是单个残基的取代,但是视施加于多肽的功能性限制而定也可能是成簇取代;插入通常在1到10个氨基酸残基的数量级。优选地,氨基酸取代包括保守的氨基酸取代。本领域可以容易地获得保守取代表。下表给出了保守氨基酸取代的实例。
表3:保守氨基酸取代的实例
残基 | 保守取代 | 残基 | 保守取代 |
Ala | Ser | Leu | Ile;Val |
Arg | Lys | Lys | Arg;Gln |
Asn | Gln;His | Met | Leu;Ile |
Asp | Glu | Phe | Met;Leu;Tyr |
Gln | Asn | Ser | Thr;Gly |
Cys | Ser | Thr | Ser;Val |
Glu | Asp | Trp | Tyr |
Gly | Pro | Tyr | Trp;Phe |
His | Asn;Gln | Val | Ile;Leu |
Ile | Leu;Val |
同源物也可以是蛋白质的“插入变体”的形式,即在蛋白质的预定位置引入一个或多个氨基酸残基。插入可以包括氨基端和/或羧基端的融合,以及单个或多个氨基酸的内部序列插入。一般,氨基酸序列内部的插入将小于氨基或羧基端的融合,数量级在约1到10个残基。氨基或羧基端融合蛋白质或肽的实例包括在酵母双杂交系统中应用的转录激活因子的结合结构域或激活结构域、噬菌体外壳蛋白质、(组氨酸)6标签、谷胱甘肽S-转移酶标签、蛋白质A、麦芽糖结合蛋白、二氢叶酸还原酶、Tag·100表位、c-myc表位、FLAG表位、lacZ、CMP(钙调蛋白结合肽)、HA表位、蛋白质C表位和VSV表位。
蛋白质“缺失变体”形式的同源物特征在于从蛋白质中除去一个或多个氨基酸。
可通过本领域众所周知的肽合成技术,如固相肽合成法等,或通过重组DNA操作容易地得到蛋白质的氨基酸变体。用于操纵DNA序列以产生蛋白质的取代、插入或缺失变体的方法是本领域众所周知的。例如,本领域技术人员熟知在DNA预定位置产生取代突变的技术,其包括M13诱变、T7-Gen体外诱变(USB,Cleveland,OH)、QuickChange定点诱变(Stratagene,San Diego,CA)、PCR介导的定点诱变或其它定点诱变方法。
SYT多肽或其同源物可以是衍生物。“衍生物”包括肽、寡肽、多肽、蛋白质和酶,与天然产生形式的蛋白质,例如SEQ ID NO:4、SEQ ID NO:6、SEQ ID NO:8、SEQ ID NO:10、SEQ ID NO:12、SEQ ID NO:14、SEQ ID NO:16、SEQ ID NO:18、SEQ ID NO:20、SEQ ID NO:22、SEQID NO:24、SEQ ID NO:26、SEQ ID NO:28、SEQ ID NO:30、SEQ ID NO:32、SEQ ID NO:34、SEQ ID NO:36、SEQ ID NO:38、SEQ ID NO:40、SEQ ID NO:42、SEQ ID NO:44、SEQ ID NO:46、SEQ ID NO:48、SEQID NO:50、SEQ ID NO:52、SEQ ID NO:54、SEQ ID NO:56、SEQ ID NO:58、SEQ ID NO:60、SEQ ID NO:62、SEQ ID NO:64、SEQ ID NO:66、SEQ ID NO:68、SEQ ID NO:70、SEQ ID NO:72、SEQ ID NO:74、SEQID NO:76、SEQ ID NO:78、SEQ ID NO:80、SEQ ID NO:82、SEQ ID NO:84、SEQ ID NO:86、SEQ ID NO:88所代表的氨基酸序列相比,其可以包括取代、缺失或添加的非天然产生的氨基酸残基。
蛋白质的“衍生物”包括肽、寡肽、多肽、蛋白质和酶,与天然产生形式多肽的氨基酸序列相比,其可以包括天然产生的改变的、糖基化、酰基化、异戊烯化或非天然产生的氨基酸残基。衍生物还可以包括相对于其源自的氨基酸序列的一个或多个非氨基酸取代基,例如共价或非共价地结合于氨基酸序列的报告分子或其它配体,例如与之结合有利于衍生物检测的报告分子,以及相对于天然产生蛋白质的氨基酸序列而言非天然产生的氨基酸残基。
SYT多肽或其同源物可以由SYT核酸/基因的选择性剪接变体编码。本文所用的术语“选择性剪接变体”包括其中选择的内含子和/或外显子已被切除、替换或添加的核酸序列变体,或者其中内含子已被缩短或增长的核酸序列变体。这样的变体保持了蛋白质的生物活性,这可以通过有选择地保留蛋白质的功能性区段来实现。这样的剪接变体可以是天然的或人工的。产生这类剪接变体的方法是本领域众所周知的。优选的剪接变体是编码多肽的核酸的变体,所述多肽从N末端到C末端包含:(i)按照递增的偏好顺序,与SEQ ID NO:2的SNH结构域具有至少40%、45%、50%、55%、60%、65%、70%、75%、80%、85%、90%、91%、92%、93%、94%、95%、96%、97%、98%、99%序列同一性的SNH结构域;和(ii)Met富含结构域;和(iii)QG富含结构域。优选地,与SEQ ID NO:2的SNH结构域具有至少40%同一性的SNH结构域包含图2中显示为黑色的残基。进一步优选地,SNH结构域为SEQ ID NO:1所代表。
另外,SYT多肽或其同源物可以包含如下一个或多个序列:(i)SEQ IDNO:90;和/或(ii)SEQ ID NO:91;和/或(iii)位于SNH结构域之前的N末端的Met富含结构域。
还优选SEQ ID NO:3、SEQ ID NO:5、SEQ ID NO:7、SEQ ID NO:9、SEQ ID NO:11、SEQ ID NO:13、SEQ ID NO:15、SEQ ID NO:17、SEQ ID NO:19、SEQ ID NO:21、SEQ ID NO:23、SEQ ID NO:25、SEQID NO:27、SEQ ID NO:29、SEQ ID NO:31、SEQ ID NO:33、SEQ ID NO:35、SEQ ID NO:37、SEQ ID NO:39、SEQ ID NO:41、SEQ ID NO:43、SEQ ID NO:45、SEQ ID NO:47、SEQ ID NO:49、SEQ ID NO:51、SEQID NO:53、SEQ ID NO:55、SEQ ID NO:57、SEQ ID NO:59、SEQ ID NO:61、SEQ ID NO:63、SEQ ID NO:65、SEQ ID NO:67、SEQ ID NO:69、SEQ ID NO:71、SEQ ID NO:73、SEQ ID NO:75、SEQ ID NO:77、SEQID NO:79、SEQ ID NO:81、SEQ ID NO:83、SEQ ID NO:85和SEQ IDNO:87所代表的核酸的剪接变体。最优选SEQ ID NO:3、SEQ ID NO:5和SEQ ID NO:7所代表SYT核酸/基因的剪接变体。
同源物还可以由编码SYT多肽或其同源物的核酸的等位基因变体所编码,优选由核酸等位基因变体编码的多肽从N末端到C末端包含:(i)按照递增的偏好顺序,与SEQ ID NO:2的SNH结构域具有至少40%、45%、50%、55%、60%、65%、70%、75%、80%、85%、90%、91%、92%、93%、94%、95%、96%、97%、98%、99%序列同一性的SNH结构域;和(ii)Met富含结构域;和(iii)QG富含结构域。优选地,与SEQ ID NO:2的SNH结构域具有至少40%同一性的SNH结构域包含图2中显示为黑色的残基。进一步优选地,SNH结构域为SEQ ID NO:1所代表。另外,SYT多肽或其同源物可以包含如下一个或多个序列:(i)SEQ ID NO:90;和/或(ii)SEQ IDNO:91;和/或(iii)位于SNH结构域之前的N末端的Met富含结构域。
还优选等位基因变体是SEQ ID NO:3、SEQ ID NO:5、SEQ ID NO:7、SEQ ID NO:9、SEQ ID NO:11、SEQ ID NO:13、SEQ ID NO:15、SEQ ID NO:17、SEQ ID NO:19、SEQ ID NO:21、SEQ ID NO:23、SEQID NO:25、SEQ ID NO:27、SEQ ID NO:29、SEQ ID NO:31、SEQ ID NO:33、SEQ ID NO:35、SEQ ID NO:37、SEQ ID NO:39、SEQ ID NO:41、SEQ ID NO:43、SEQ ID NO:45、SEQ ID NO:47、SEQ ID NO:49、SEQID NO:51、SEQ ID NO:53、SEQ ID NO:55、SEQ ID NO:57、SEQ ID NO:59、SEQ ID NO:61、SEQ ID NO:63、SEQ ID NO:65、SEQ ID NO:67、SEQ ID NO:69、SEQ ID NO:71、SEQ ID NO:73、SEQ ID NO:75、SEQID NO:77、SEQ ID NO:79、SEQ ID NO:81、SEQ ID NO:83、SEQ ID NO:85和SEQ ID NO:87所代表核酸的等位基因变体。最优选等位基因变体是SEQ ID NO:3、SEQ ID NO:5和SEQ ID NO:7中任一所代表核酸的等位基因变体。
等位基因变体天然存在,并且这些天然等位基因的用途包含于本发明的方法中。等位基因变体包括单核苷酸多态性(SNP),以及小型插入/缺失多态性(INDEL)。INDEL的大小通常小于100bp。SNP和INDEL在大多数生物体天然存在的多态性品系中形成最大的一组序列变体。
根据本发明的优选方面,SYT核酸或其变体受调节的表达是增加的表达。增加的表达可以引起SYT mRNA或多肽水平提高,这等同于提高SYT多肽的活性;或者当多肽水平没有变化,或者甚至当多肽水平下降时也可以提高活性。这种情况出现在多肽固有特性发生改变的时候,例如,通过制备比野生型多肽更具活性的突变体形式。增加基因或基因产物表达的方法在本领域有充分的记录,其包括,例如由适当的启动子驱动的过表达、转录增强子或翻译增强子的使用。可以将用作启动子或增强子元件的分离的核酸引入非异源形式多核苷酸的适当位置(一般是上游),从而上调SYT核酸或其变体的表达。例如,可以通过突变、缺失和/或取代,体内地改变内源启动子(见Kmiec,US 5,565,350;Zarling等,PCT/US93/03868),或者将分离的启动子在本发明基因的适当方向和距离引入植物细胞中,从而控制基因的表达。降低基因或基因产物表达的方法在本领域有充分的记录,
如果期望多肽表达,通常要在多肽编码区域的3’末端纳入多聚腺苷酸化区域。多聚腺苷酸化区域可以源自天然基因、多种其它植物基因或T-DNA。例如,待加入的3’末端序列可以源自胭脂碱合酶或章鱼碱合酶基因、或备选地源自其他植物基因、或次优选地源自任何其它真核基因。
也可以在5’非翻译区或部分编码序列的编码序列中加入内含子序列,来增加在胞质中累积的成熟信使的数量。已显示,在植物和动物表达构建体的转录单位中纳入的可剪接内含子均可以在mRNA和蛋白质水平增加高达1000倍的基因表达,Buchman和Berg,Mol.Cell biol.8:4395-4405(1988);Callis等,Genes Dev.1:1183-1200(1987)。通常这类内含子被放置在转录单位5’末端附近时,其增强基因表达的作用达到最大。玉米内含子Adh1-S内含子1、2和6,Bronze-1内含子的使用是本领域公知的。通常见The MaizeHandbook,第116章,Freeling和Walbot编辑,Springer,N.Y.(1994)。
本发明还提供遗传构建体和载体,以促进用于本发明方法中的核苷酸序列的引入和/或表达。
因此,提供的基因构建体含有:
(i)如上文所定义的SYT核酸或其变体;
(ii)一个或多个能够驱动(i)中核酸序列表达的控制序列;和任选的
(iii)转录终止序列。
可以使用本领域技术人员熟知的重组DNA技术构建用于本发明方法的构建体。可以将基因构建体插入可商购的、适合于转化进入植物并适于在转化的细胞中表达目的基因的载体中。
使用含有目的序列(即编码SYT多肽或其同源物的核酸)的载体转化植物。将目的序列有效连接于一个或多个控制序列(至少连接于启动子)。术语“调控元件”、“控制序列”和“启动子”在本文中都可交换的使用,从广义上是指能够影响与之相连的序列表达的调控核酸序列。上述术语包括源自典型真核生物基因组基因的转录调控序列(包括具有或没有CCAAT盒序列的TATA盒,其对于精确的转录起始是必需的),以及另外的调控元件(即上游激活序列、增强子和沉默子),其通过应答发育刺激和/或外部刺激或以组织特异的方式改变基因表达。该术语还包括了经典原核生物基因的转录调控序列,在此情况下可以包括-35盒序列和/或-10盒转录调控序列。术语“调控元件”也包含合成的融合分子或衍生物,其赋予、激活或增强细胞、组织或器官中核酸分子的表达。本文所用的术语“有效连接”指在启动子序列和目的基因之间的功能性连接,以使启动子序列能起始目的基因的转录。
有利地,可以使用任意类型的启动子驱动核酸序列的表达。启动子可以是诱导型启动子,即应答发育、化学、环境或物理刺激,具有诱导的或增加的转录起始。诱导型启动子的实例是胁迫诱导型启动子,即当植物接触多种胁迫条件时激活的启动子。另外或备选的,所述启动子可以是组织偏好的启动子,即能够在某些组织,如在叶、根、种子等组织中优先地起始转录的启动子。
优选地,SYT核酸或其变体有效连接于组成型启动子。组成型启动子在其生长和发育的大多数但不必然是所有阶段都转录激活,并且基本上是普遍表达。优选启动子来源于植物,还优选来源于单子叶植物。最优选使用GOS2启动子(来自稻)(SEQ ID NO:89)。应当清楚本发明的实用性并不局限于SEQ ID NO:3、SEQ ID NO:5或SEQ ID NO:7所代表的SYT核酸,而且本发明的实用性也不局限于由GOS2启动子所驱动的SYT核酸的表达。也可用来驱动SYT核酸表达的其他组成型启动子的实例示于下表4中。
表4:组成型启动子的实例
基因来源 | 表达模式 | 参考文献 |
肌动蛋白 | 组成型 | McElroy等,Plant Cell,2:163-171,1990 |
CAMV35S | 组成型 | Odell等,Nature,313:810-812,1985 |
CaMV 19S | 组成型 | Nilsson等,Physiol.Plant.100:456-462,1997 |
GOS2 | 组成型 | de Pater等,Plant J Nov;2(6):837-44,1992 |
泛素 | 组成型 | Christensen等,Plant Mol.Biol.18:675-689,1992 |
稻亲环蛋白 | 组成型 | Buchholz等,Plant Mol Biol.25(5):837-43,1994 |
玉米H3组蛋白 | 组成型 | Lepetit等,Mol.Gen.Genet.231:276-285,1992 |
肌动蛋白2 | 组成型 | An等,Plant J.10(1);107-121,1996 |
任选的,还可以在引入植物的构建体中使用一个或多个终止子序列。术语“终止子”包括控制序列,其为位于转录单位末端的DNA序列,传递信号引发初级转录本的3’加工和多聚腺苷酸化以及转录的终止。另外的调控元件可以包括转录和翻译的增强子。本领域技术人员将知道适合用于进行本发明的终止子和增强子的序列。这类序列为本领域技术人员所公知或者可以容易地获得。
本发明的遗传构建体还包括在特定细胞类型中维持和/或复制所需的复制起点序列。一个实例是需要将遗传构建体作为附加型遗传元件(如质粒或粘粒分子)在细菌细胞中维持的情况。优选的复制起点包括但不限于f1-ori和colE1。
遗传构建体可以任选地包括可选择的标记基因。如本文所用,术语“可选择的标记基因”包括赋予细胞表型的任意基因,该基因在细胞中表达,有利于鉴定和/或选择经本发明的核酸构建体转染或转化的细胞。适当的标记可以选自赋予抗生素或除草剂抗性的标记,其引入新的代谢性状或允许可视选择。可选择标记基因的实例包括赋予抗生素抗性的基因(例如磷酸化新霉素和卡那霉素的nptII,或磷酸化潮霉素的hpt)、赋予除草剂抗性的基因(例如提供Basta抗性的bar;或提供草甘膦抗性的aroA或gox)、或者提供代谢性状的基因(例如允许植物使用甘露糖作为唯一碳源的manA)。可视标记基因导致形成颜色(例如β-葡糖醛酸糖苷酶,GUS)、发光(例如荧光素酶)或荧光(绿色荧光蛋白GFP及其衍生物)。
本发明还包括可由本发明方法获得的植物。本发明因此提供可由本发明方法获得的植物、植物部分和植物细胞,该植物、植物部分和植物细胞中引入了SYT核酸或其变体,并且该植物、植物部分和植物细胞优选来自作物植物,更优选来自单子叶植物。
本发明还提供产生产率增加的转基因植物的方法,其包括在植物中引入和表达SYT核酸或其变体。
更具体地,本发明提供产生产率增加的转基因植物,优选转基因单子叶植物的方法,该方法包括:
(i)向植物或植物细胞中引入和表达SYT核酸或其变体;和
(ii)在促进植物生长和发育的条件下培养植物细胞。
由培养步骤(ii)获得的植物的后续世代可以通过多种方式繁殖,如用克隆繁殖或经典的育种技术。例如,第一代(或T1)转化的植物可自交得到纯合的第二代(或T2)转化体,而T2植物进一步通过经典育种技术繁殖。
可以将核酸直接引入植物细胞或植物本身(包括引入组织、器官或植物的任何其它部分)。根据本发明的优选方面,通过转化将核酸引入植物。
本文所指术语“转化”包括将外源多核苷酸转移进宿主细胞,不考虑转移所用的方法。通过器官发生或者胚胎发生的能够随即克隆增殖的植物组织都可以使用本发明的遗传构建体转化并从其再生整个植物。具体的组织选择将因可提供和最适于转化的具体物种的克隆增殖系统而改变。示例性的靶组织包括叶盘、花粉、胚、子叶、胚轴、雌配子、愈伤组织、既有的分生组织(例如顶端分生组织、腋芽和根分生组织),以及诱导的分生组织(例如子叶分生组织和胚轴分生组织)。可以将多核苷酸瞬时地或稳定地引入宿主细胞,并且可以,例如作为质粒保持非整合的状态。备选地,其可以整合进入宿主基因组。得到的转化植物细胞可以接着用于以本领域技术人员熟知的方式再生为转化的植物。
植物物种的转化目前是一种相当常规的技术。有利地,可以使用几种转化方法的任一向适当的祖先细胞引入目的基因。转化方法包括用脂质体、电穿孔、增强游离DNA摄取的化学物质、直接向植物注射DNA、粒子枪轰击、用病毒或花粉转化和显微投影(microprojection)。方法可以选自用于原生质体的钙/聚乙二醇方法(Krens,F.A.等,(1882)Nature 296,72-74;Negrutiu I.等,(1987)Plant Mol.Biol.8:363-373);原生质体的电穿孔法(Shillito R.D.等,1985Bio/Technol 3,1099-1102);植物材料的显微注射(Crossway A.等,(1986)Mol.Gen Genet 202:179-185);DNA或RNA包被的粒子轰击(Klein T.M.等,(1987)Nature 327:70);(非整合的)病毒感染,等等。优选使用任何转稻转化的熟知方法,通过农杆菌介导的转化,产生表达SYT基因/核酸的转基因稻类植物,例如在任何以下任一文献中描述的方法:公开的欧洲专利申请EP 1198985A1,Aldemita和Hodges(Planta,199:612-617,1996);Chan等(Plant Mol.Biol.22(3)491-506,1993),Hiei等(PlantJ.6(2):271-282,1994),其公开的内容如同其陈述的全部内容那样并入本文作为参考。至于谷物转化,优选的方法如Ishida等(Nat.Biotechnol.14(6):745-50,1996)或Frame等(Plant Physiol.129(1):13-22,2002)中所述,其公开的内容如同其陈述的全部内容那样并入本文作为参考。
通常在转化以后,选出存在一个或多个标记的植物细胞或细胞群,所述标记由与目的基因共转移的植物可表达基因编码,继之将转化的材料再生成整个植物。
DNA转移和再生之后,可评估推定转化的植物,例如用Southern分析评价目的基因的存在、拷贝数和/或基因组构造。备选的或额外的,可用Northern和/或Western分析、定量PCR监测新引入基因的表达水平,这类技术都是本领域内普通技术人员所熟知的。
产生的转化植物可以通过多种方式繁殖,如用克隆繁殖或经典的育种技术。例如,第一代(或T1)转化的植物可自交得到纯合的第二代(或T2)转化体,T2植物进一步通过经典育种技术繁殖。
产生的转化生物体可以有多种形式。例如,它们可以是转化细胞和非转化细胞的嵌合体;克隆的转化体(例如经过转化包含表达盒的所有细胞);转化和非转化组织的嫁接体(例如在植物中,转化的根茎嫁接到非转化的接穗上)。
本发明显然延及由本文所述方法产生的任何植物细胞或植物,以及所有的植物部分和其无性繁殖体。本发明还涵盖由任意上述方法产生的初级转化或转染的细胞、组织、器官或整个植物的后代,所述后代的唯一要求是与本发明方法产生的亲本呈现同样的基因型和/或表型特性。本发明也包括含有分离的SYT核酸或其变体的宿主细胞。本发明优选的宿主细胞是植物细胞。本发明也延及植物可收获的部分,例如,但不限于种子、叶、果实、花、茎培养物、根茎、块茎和球茎。本发明还涉及由这样的植物的可收获部分衍生的产品,如干丸或干粉、粗粉、油类、脂肪和脂肪酸、淀粉或蛋白质。
本发明还包括SYT核酸或其变体、SYT多肽或其同源物、以及上文定义的构建体在增加植物产率特别是种子产率中的用途。种子产率如上文所定义的那样,并且优选包括增加的种子总产率或增加的TKW。
可以在育种程序中使用SYT核酸或其变体或者SYT多肽或其同源物,其中鉴定可以遗传地连接于SYT基因或其变体的DNA标记。可以使用SYT核酸/基因或其变体或者SYT多肽或其同源物界定分子标记。接着可以将此DNA或蛋白质标记在育种程序中使用,以选择具有增加的产率的植物。例如,SYT基因或其变体可以是SEQ ID NO:3、SEQ ID NO:5、SEQ ID NO:7、SEQ ID NO:9、SEQ ID NO:11、SEQ ID NO:13、SEQ ID NO:15、SEQ ID NO:17、SEQ ID NO:19、SEQ ID NO:21、SEQ ID NO:23、SEQID NO:25、SEQ ID NO:27、SEQ ID NO:29、SEQ ID NO:31、SEQ ID NO:33、SEQ ID NO:35、SEQ ID NO:37、SEQ ID NO:39、SEQ ID NO:41、SEQ ID NO:43、SEQ ID NO:45、SEQ ID NO:47、SEQ ID NO:49、SEQID NO:51、SEQ ID NO:53、SEQ ID NO:55、SEQ ID NO:57、SEQ ID NO:59、SEQ ID NO:61、SEQ ID NO:63、SEQ ID NO:65、SEQ ID NO:67、SEQ ID NO:69、SEQ ID NO:71、SEQ ID NO:73、SEQ ID NO:75、SEQID NO:77、SEQ ID NO:79、SEQ ID NO:81、SEQ ID NO:83、SEQ ID NO:85和SEQ ID NO:87中任一所代表的核酸。
SYT核酸/基因的等位基因变体也可以用于标记辅助的育种程序。这类育种程序有时需要使用,例如EMS诱变,通过植物诱变处理引入等位基因变体;备选的,此程序可以以收集无意产生的所谓“天然”起源的等位基因变体开始。然后通过例如PCR鉴定等位基因变体。随后是选择步骤,用以选择所讨论序列的较好等位基因变体,所述等位基因变体赋予植物增加的产率。一般通过监测含有所研究序列的不同等位基因变体植物的生长行为来进行选择,所述研究序列的不同等位基因变体是例如SEQ ID NO:3、SEQ ID NO:5、SEQ ID NO:7、SEQ ID NO:9、SEQ ID NO:11、SEQ IDNO:13、SEQ ID NO:15、SEQ ID NO:17、SEQ ID NO:19、SEQ ID NO:21、SEQ ID NO:23、SEQ ID NO:25、SEQ ID NO:27、SEQ ID NO:29、SEQ ID NO:31、SEQ ID NO:33、SEQ ID NO:35、SEQ ID NO:37、SEQID NO:39、SEQ ID NO:41、SEQ ID NO:43、SEQ ID NO:45、SEQ ID NO:47、SEQ ID NO:49、SEQ ID NO:51、SEQ ID NO:53、SEQ ID NO:55、SEQ ID NO:57、SEQ ID NO:59、SEQ ID NO:61、SEQ ID NO:63、SEQID NO:65、SEQ ID NO:67、SEQ ID NO:69、SEQ ID NO:71、SEQ ID NO:73、SEQ ID NO:75、SEQ ID NO:77、SEQ ID NO:79、SEQ ID NO:81、SEQ ID NO:83、SEQ ID NO:85和SEQ ID NO:87中任一的不同等位基因变体。可以在温室或田地中监测生长行为。更多任选的步骤包括,将经鉴定含有较好等位基因变体的植物与另一植物杂交。例如,可使用这种方法产生感兴趣表型特征的组合。
SYT核酸或其变体还可以作为探针,用于对为那些基因连锁性状的一部分并作为其标志物的基因进行遗传和物理的作图。这样的信息可以在植物育种中使用,以得到具有所期望表型的品系。SYT核酸或其变体的这类应用仅需要长至少15个核苷酸的核酸序列。SYT核酸或其变体可以用作限制性片段长度多态性(RFLP)标记。可用SYT核酸或其变体探测限制酶切消化的植物基因组DNA的Southern印迹(Sambrook J,Fritsch EF和ManiatisT(1989)《分子克隆:实验室手册》)。随后使用计算机程序如MapMaker(Lander等(1987)Genomics 1:174-181)对产生的带型进行遗传分析,以构建遗传图谱。此外,可以使用核酸在含有一组个体的限制性内切酶处理的基因组DNA中探测Southern印迹,所述一组个体为代表明确的遗传杂交的亲本和子代的一组个体。记录DNA多态性的分离并用于计算在先前用此群体获得的遗传图谱中SYT核酸或其变体的位置(Botstein等(1980)Am.J.Hum.Genet.32:314-331)。
在遗传作图中使用的植物基因衍生探针的产生和用途描述于Bematzky和Tanksley(1986)Plant Mol.Biol.Reporter 4:37-41中。众多出版物中描述过用上述方法或其变通形式对特定cDNA克隆进行遗传作图。例如,可以使用F2杂交群体、回交群体、随机交配群体、近亲同基因系和其它个体组作图。这类方法是本领域技术人员众所周知的。
核酸探针也可以用于物理作图(即在物理图谱上安置序列;见Hoheisel等In:Non-mammalian Genomic Analysis:A Practical Guide,Academicpress 1996,第319-346页,及其中引用的参考文献)。
在另一个实施方案中,核酸探针可用于直接荧光原位杂交(FISH)作图(Trask(1991)Trends Genet.7:149-154)。尽管目前FISH作图的方法倾向用于大的克隆(几个kb到几百个kb;见Laan等(1995)Genome Res.5:13-20),但是灵敏性的提高允许在FISH作图中应用较短的探针。
用于遗传和物理作图的多种基于核酸扩增的方法可以使用所述核酸进行。实例包括等位基因特异性扩增(Kazazian(1989)J.Lab.Clin.Med11:95-96)、PCR扩增片段的多态性(CAPS;Sheffield等(1993)Genomics16:325-332)、等位基因特异性连接(Landegren等(1988)Science 241:1077-1080)、核苷酸延伸反应(Sokolov(1990)Nucleic Acid Res.18:3671)、放射杂交作图(Walter等(1997)Nat.Genet.7:22-28)和Happy作图(Dear和Cook(1989)Nucleic Acid Res.17:6795-6807)。为实施这些方法,使用核酸序列设计和产生用于扩增反应或引物延伸反应的引物对。这类引物的设计是本领域技术人员众所周知的。使用基于PCR的遗传作图的方法,可能需要鉴定跨越相应于本发明核酸序列区域作图的亲本之间DNA序列的差异。然而,这对作图方法通常不是必要的。
根据本发明的方法得到如前所述产率提高的植物。产率提高的性状还可以组合其它经济上有利的性状,如更多提高产率的性状、对多种胁迫的耐受性、改良多种构造特征和/或生化和/或生理学特征的性状。
附图说明
现参考以下附图描述本发明,其中:
图1显示了植物和哺乳动物SYT多肽典型的结构域结构。保守的SNH结构域位于蛋白质的N末端。对于植物SYT多肽而言,蛋白质结构域的C末端剩余部分由QG富含结构域组成,对于哺乳动物SYT多肽而言,其由QPGY富含结构域组成。Met富含结构域通常包含在植物QG富含结构域或哺乳动物QPGY富含结构域的头一半之内(从N末端到C末端的方向)。可能有第二Met富含结构域位于植物SYT多肽的SNH结构域之前。
图2显示了数个SYT多肽N末端的多重比对,使用的是基于修饰的ClustalW算法(InforMax,Bethesda,MD,http://www.informaxinc.com)的VNTI AlignX多重比对程序,采用默认设置,空位开放罚分为10,空位延伸罚分为0.05。植物和人SYT多肽的SNH结构域以加框表示。比对中最后一行包括来源于所比对序列的共有序列。
图3显示了数个植物SYT多肽的多重比对,使用的是基于修饰的ClustalW算法(InforMax,Bethesda,MD,http://www.informaxinc.com)的VNTI AlignX多重比对程序,采用默认设置,空位开放罚分为10,空位延伸罚分为0.05。从N末端到C末端方向的两个主要结构域以加框表示,并被鉴定为SNH结构域和Met富含/QG富含结构域。另外,N末端Met富含结构域也以加框表示,SEQ ID NO:90和SEQ ID NO 91的位置以粗体下划线表示。
图4显示了利用ClustalW 1.83(http://align.genome.jp/sit-bin/clustalw)通过序列比对得到的邻近连接树。SYT1和SYT2/SYT3进化枝以括弧标识。
图5显示了双元载体p0523,用于在稻中表达处于GOS2启动子(内参PRO0129)控制之下的拟南芥AtSYT1。
图6显示了双元载体p0524,用于在稻中表达处于GOS2启动子(内参PRO0129)控制之下的拟南芥AtSYT2。
图7显示了双元载体p0767,用于在稻中表达处于GOS2启动子(内参PRO0129)控制之下的拟南芥AtSYT3。
图8详述了用于执行本发明方法的序列实例。SYT核酸序列从起点到终点表示。这些序列绝大多数来自于EST测序,质量较低。因此可能会遇到核酸取代的情况。
实施例
现参考以下实施例描述本发明,所述实施例仅意在举例说明。
DNA操作除非另外说明,重组DNA技术根据描述于(Sambrook(2001)《分子克隆:实验室手册》,第三版,冷泉港实验室出版,冷泉港,纽约)或者Ausubel等(1994),Current Protocols in Molecular Biology,CurrentProtocols第一卷和第二卷的标准方法执行。植物分子操作的标准材料和方法由R.D.D.Croy描述于Plant Molecular Biology Labfase(1993),由BIOSScientific Publications Ltd(UK)和Blackwell Scientific Publications(UK)出版。
实施例1:AtSYT1、AtSYT2和AtSYT3的基因克隆
使用拟南芥幼苗cDNA文库(Invitrogen,Paisley,UK)作为模板通过PCR扩增拟南芥AtSYT1基因。从幼苗提取的RNA经反转录后,将cDNA克隆进入pCMV Sport 6.0。该库平均插入大小为1.5kb,并且原始克隆数的数量级在1.59×107cfu。在6×1011 cfu/ml的第一次扩增之后,确定原始滴度为9.6×105cfu/ml。提取质粒之后,将200ng模板用于50μl PCR混合物中。PCR扩增所用的引物包括Gateway重组的AttB位点,为prm06681(SEQID NO:92;正义,起始密码子为粗体,AttB1位点为斜体:5’-GGGGACAAGTTTGTACAAAAAAGCAGGCTTAAACAATGCAACAGCACCTGATG-3’)和prm06682(SEQ ID NO:93;反义,互补的,AttB2位点为斜体:5’-GGGGACCACTTTGTACAAGAAAGCTGGGTCATCATTAAGATTCCTTGTGC-3’)。在标准条件下使用Hifi Taq DNA聚合酶进行PCR。同样用标准方法扩增和纯化727bp(包括attB位点)的PCR片段。接着进行Gateway操作的第一步,BP反应,在此期间将PCR片段与pDONR201质粒体内重组以产生Gateway术语所称的“进入(entry)克隆”,p07466。作为Gateway技术一部分的质粒pDPNR201购自Invitrogen。
利用与拟南芥AtSYT1基因相同的方法,通过PCR扩增拟南芥AtSYT2基因。PCR扩增所用的引物包括Gateway重组的AttB位点,为prm06685(SEQ ID NP:94;正义,起始密码子为粗体,AttB1位点为斜体:5’-GGGGACAAGTTTGTACAAAAAAGCAGGCTTAAACAATGCAGCAGCAGCAGTCT 3’)和prm06686(SEQ ID NO:95;反义,终止密码子为粗体,互补的,AttB2位点为斜体:5’GGGGACCACTTTGTACAAGAAAGCTGGGTTCTTTGGATCCTTTTCACTTG 3’)。在标准条件下使用Hifi Taq DNA聚合酶进行PCR。如上扩增和纯化666bp(包括attB位点)的PCR片段。将进入克隆编号为p07467。
利用与拟南芥AtSYT1和AtSYT2基因相同的方法,通过PCR扩增拟南芥AtSYT3基因。PCR扩增所用的引物包括Gateway重组的AttB位点,为prm06683(SEQ ID NO:96;正义,起始密码子为粗体,AttB1位点为斜体:5’GGGGACAAGTTTGTACAAAAAAGCAGGCTTAAACAATGCAGCAATCTCCACAGAT 3’)和prm06684(SEQ ID NO:97;反义,终止密码子为粗体,互补的,AttB2位点为斜体:5’GGGGACCACTTTGTACAAGAAAGCTGGGTTCCTCTATTTCATTTTCCTTCAG 3’)。在标准条件下使用Hifi TaqDNA聚合酶进行PCR。如上扩增和纯化745bp(包括attB位点)的PCR片段。将进入克隆编号为p07604。
实施例2:载体构建
接着,将进入克隆p07466、p07467和p07604与用于稻转化的指定载体p00640一起被用于LR反应此载体在T-DNA边界内包含以下部分作为功能性元件:植物可选择的标记;可筛选的标记表达盒;旨在与已克隆到进入克隆中的目的序列进行体内重组LR的Gateway表达盒。用于组成型表达的稻GOS2启动子(SEQ ID NO:89)(PR00129)在此Gateway盒的上游。
LR重组步骤之后,将所产生的表达载体,分别为用于AtSYT1的p0523、用于AtSYT2的p0524以及用于AtSYT3的p0767(图5至7),转化进入农杆菌菌株LBA4044,随后转化进入稻类植物。使转化的稻类植物生长,随后检测实施例3中描述的参数。
实施例3:处于稻GOS2启动子控制之下的AtSYT1、AtSYT2和AtSYT3的评估及结果
大约产生了15到20个独立的T0稻类转化体。初级转化体由组织培养室转移到温室生长并收获T1种子。6个事件得以保留,其中T1代发生转基因存在/缺乏的3∶1分离。通过监测可视标记的表达,在每一事件中选出大约10个含转基因(杂合子和纯合子)的T1幼苗,和大约10个缺少转基因(无效合子)的T1幼苗。
统计分析:F-检验
使用双因子ANOVA(变异分析)作为植物表型特性整体评估的统计模型。在由本发明基因转化的所有事件的所有植物中,对所有测量参数进行F检验。进行F检验来检查所有转化事件中基因的效应,并验证基因的整体效应,亦称为整体基因效应。真实的整体基因效应显著性的阈值设定为F检验的5%概率水平。显著性F检验值证明基因效应,其意味着不仅仅是基因的存在或位置引起表型的差异。
种子相关参数的测量
成熟的初级圆锥花序被收获、包装、标记条形码,然后在37℃烘箱中干燥三天。然后敲打圆锥花序并对所有种子进行收集和计数。用吹风装置将饱满的壳与空壳分离。丢弃空壳,并再次计数剩余的部分。饱满的壳在分析天平上称重。通过计数分离步骤之后剩余的饱满壳数来确定饱满种子数。通过称量从植物收获的全部饱满的壳来测量总的种子产率。通过计数自植物收获的谷壳数来测量每个植株的种子总数。千粒重(TKW)从已计数的饱满种子数及其总重外推得到。
利用客户定制装置来测量单个种子参数(包括宽度、长度、面积、重量),所述装置由两个主要组件称重装置和成像装置构成,连接于用于图像分析的软件。
3.1温室中生长的转基因植物的种子总产率和TKW测量结果
T1代AtSYT1、AtSYT2和AtSYT3转基因植物的种子总产率和TKW测量结果示于表5至7中。标出了两参数中任一参数增加了的品系数。也显示了转基因和相应无效合子之间的百分比差异,以及F检验的P值。
对于T1代的AtSYT1、AtSYT2和AtSYT3转基因植物,种子总产率和TKW都显著增加(分别为表5至7)。
表5:T1代AtSYT1转基因植物的种子总产率和TKW测量结果
显示增加的事件数 | %差异 | F检验的P值 | |
种子总产率 | 6个中的5个 | 19 | 0.005 |
TKW | 6个中的6个 | 11 | <0.0001 |
表6:T1代AtSYT2转基因植物的种子总产率和TKW测量结果
显示增加的事件数 | %差异 | F检验的P值 | |
种子总产率 | 6个中的4个 | 37 | 0.05 |
TKW | 6个中的6个 | 5 | <0.0001 |
表7:T1代AtSYT3转基因植物的种子总产率和TKW测量结果
显示增加的事件数 | %差异 | F检验的P值 | |
种子总产率 | 6个中的5个 | 22 | 0.0074 |
TKW | 6个中的5个 | 7 | <0.0001 |
3.2 T2代AtSYT1转基因植物种子的种子大小测量结果
利用客户定制装置对T2代植物的种子测量单个种子参数(宽度、长度和面积),所述装置由两个主要组件称重装置和成像装置构成,连接于用于图像分析的软件。对带壳和脱壳的种子均进行了测量。
AtSYT1转基因稻植物T3种子(自T2代植物收获)的平均单个种子面积、长度和宽度测量结果示于表8中。显示了转基因和相应无效合子之间的百分比差异,以及给定参数增加了的事件数和F检验的P值。
带壳和脱壳T3种子(自T2代AtSYT1转基因稻植物收获)的平均单个种子面积、长度和宽度,与其无效对应物相比,都显著增加。
表8:与其无效对应物相比较的AtSYT1转基因稻植物带壳和脱壳T3种子(自T2代植物收获)的单个种子面积、长度和宽度的测量
显示增加的事件数 | %差异 | F检验的P值 | |
平均种子面积 | 6个中的6个 | 11% | <0.0001 |
平均脱壳种子面积 | 6个中的6个 | 10% | <0.0001 |
平均种子长度 | 6个中的6个 | 6% | <0.0001 |
平均脱壳种子长度 | 6个中的6个 | 5% | <0.0001 |
平均种子宽度 | 6个中的6个 | 5% | <0.0001 |
平均脱壳种子宽度 | 6个中的6个 | 4% | <0.0001 |
3.3T2代AtSYT1转基因植物种子的胚大小和胚乳大小测量结果
还通过纵向对切脱壳种子,并将两半种子于35℃用着色剂2,3,5-氯化三苯基四氮唑染色2至3小时,来测量胚大小和胚乳大小。染色后,将两半种子置于皮氏培养皿的琼脂糖凝胶中以备成像。采用了三个独立的事件,其中每个事件分析了120个转基因纯合种子和120个不含转基因的种子。拍摄种子的数码相片,并用ImagePro软件分析图像。三个事件的结果提供如下。
对于所有三个事件,转基因纯合种子的胚都比不含转基因种子的胚更大。对于三个事件中的每一事件,种子胚的平均面积都显著增加,t检验的p值分别为0.0325、<0.0001和<0.0001。与此类似,对于三个事件中的每一事件,种子胚的平均周长都显著增加,t检验的p值分别为0.0176,<0.0001和<0.0001。此外,对于三个事件中的每一事件,种子胚乳的平均面积和平均周长也都显著增加,得到p值均为<0.0001。
3.4田地中互长的AtSYT1转基因植物的TKW测量结果
在9月份将AtSYT1纯合转基因植物及其相应对照移种至田地中,并在12月份收获。每份记录(4个事件)种植4份重复,每份重复为104株植物。植物之间的植距为20×20cm。向田地中注满水进行灌溉。种子收获以后,如上所述对种子TKW进行测量。测量结果表示在表9中。
表9:田地中生长的T3代AtSYT1转基因植物的TKW测量结果
事件 | TKW的百分比增加(%) |
事件1 | 8 |
事件2 | 6 |
事件3 | 5 |
事件4 | 10 |
对于在田地中评估的所有转基因事件,其TKW都增加。
序列表
<110>克罗普迪塞恩股份有限公司
<120>产率增加的植物及其制备方法
<130>CD-129-PCT
<150>EP 05100537.9
<151>2005-01-27
<150>US 60/649,041
<151>2005-02-01
<150>US 60/730,403
<151>2005-10-26
<160>97
<170>PatentIn version 3.3
<210>1
<211>46
<212>PRT
<213>人工序列
<220>
<223>共有序列
<220>
<221>MISC_FEATURE
<222>(3)..(3)
<223>Xaa可以是Gln或Lys
<220>
<221>MISC_FEATURE
<222>(4)..(4)
<223>Xaa可以是任何氨基酸
<220>
<221>MISC_FEATURE
<222>(6)..(6)
<223>Xaa可以是Asp或Glu
<220>
<221>MISC_FEATURE
<222>(7)..(7)
<223>Xaa可以是Glu或Asp
<220>
<221>MISC_FEATURE
<222>(9)..(9)
<223>Xaa可以是Lys或Asn
<220>
<221>MISC_FEATURE
<222>(10)..(10)
<223>Xaa可以是任何氨基酸
<220>
<221>MISC_FEATURE
<222>(13)..(13)
<223>Xaa可以是任何氨基酸
<220>
<221>MISC_FEATURE
<222>(14)..(14)
<223>Xaa可以是Cys、Ala和Lys中任一
<220>
<221>MISC_FEATURE
<222>(16)..(16)
<223>Xaa可以是Leu、Val或Met中任一
<220>
<221>MISC_FEATURE
<222>(17)..(17)
<223>Xaa可以是Glu、Asp或Ser中任一
<220>
<221>MISC_FEATURE
<222>(18)..(18)
<223>Xaa可以是Ser或Asn
<220>
<221>MISC_FEATURE
<222>(19)..(19)
<223>Xaa可以是Gln或Leu
<220>
<221>MISC_FEATURE
<222>(21)..(21)
<223>Xaa可以是任何氨基酸
<220>
<221>MISC_FEATURE
<222>(23)..(23)
<223>Xaa可以是Lys或Arg
<220>
<221>MISC_FEATURE
<222>(24)..(25)
<223>Xaa可以是任何氨基酸
<220>
<221>MISC_FEATURE
<222>(28)..(28)
<223>Xaa可以是Ala、Glu或Ser中任一
<220>
<221>MISC_FEATURE
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<222>(32)..(32)
<223>Xaa可以是Ala、Ser或Gln中任一
<220>
<221>MISC_FEATURE
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<221>misc_feature
<223>386位上的a和425位上的t可以改变为386位上的g和425位上的c
<400>3
atgcaacagc acctgatgca gatgcagccc atgatggctg gttactaccc cagcaatgtt 60
acctctgatc atatccaaca gtacttggac gaaaacaaat cgttgattct gaagattgtt 120
gagtctcaaa actctggaaa gcttagcgaa tgcgccgaga atcaagcaag gcttcaacgc 180
aacctaatgt acctagctgc aatagcagat tctcagcctc agccaccaag tgtgcatagc 240
cagtatggat ctgctggtgg tgggatgatt cagggagaag gagggtcaca ctatttgcag 300
cagcaacaag cgactcaaca gcaacagatg actcagcagt ctctaatggc ggctcgatct 360
tcaatgttgt atgctcagca acagcagcag cagcagcctt acgcgacgct tcagcatcag 420
caattgcacc atagccagct tggaatgagc tcgagcagcg gaggaggagg aagcagtggt 480
ctccatatcc ttcagggaga ggctggtggg tttcatgatt ttggccgtgg gaagccggaa 540
atgggaagtg gtggtggcgg tgaaggcaga ggaggaagtt caggggatgg tggagaaacc 600
ctttacttga aatcatcaga tgatgggaat tga 633
<210>4
<211>210
<212>PRT
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<220>
<221>MISC_FEATURE
<223>129位上的Gln和141位上的Leu可以改变为129位上的Arg和141位上的Ser
<400>4
Met Gln Gln His Leu Met Gln Met Gln Pro Met Met Ala Gly Tyr Tyr
1 5 10 15
Pro Ser Asn Val Thr Ser Asp His Ile Gln Gln Tyr Leu Asp Glu Asn
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Gln Ser Leu Met Ala Ala Arg Ser Ser Met Leu Tyr Ala Gln Gln Gln
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210
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aacatcacta ccgaacagat ccaaaagtac cttgatgaga acaagaagct gattatggcc 120
atcatggaaa accagaatct cggtaaactt gctgagtgcg cccagtacca agctcttctc 180
cagaagaact tgatgtatct tgctgcaatt gctgatgctc aacccccacc acctacgcca 240
ggaccttcac catctacagc tgtcgctgcc cagatggcaa caccgcattc tgggatgcaa 300
ccacctagct acttcatgca acacccacaa gcatcccctg cagggatttt cgctccaagg 360
ggtcctttac agtttggtag cccactccag tttcaggatc cgcaacagca gcagcagata 420
catcagcaag ctatgcaagg acacatgggg attagaccaa tgggtatgac caacaacggg 480
atgcagcatg cgatgcaaca accagaaacc ggtcttggag gaaacgtggg gcttagagga 540
ggaaagcaag atggagcaga tggacaagga aaagatgatg gcaagtga 588
<210>6
<211>195
<212>PRT
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Met Gln Gln Gln Gln Ser Pro Gln Met Phe Pro Met Val Pro Ser Ile
1 5 10 15
Pro Pro Ala Asn Asn Ile Thr Thr Glu Gln Ile Gln Lys Tyr Leu Asp
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Glu Asn Lys Lys Leu Ile Met Ala Ile Met Glu Asn Gln Asn Leu Gly
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Gly Leu Arg Gly Gly Lys Gln Asp Gly Ala Asp Gly Gln Gly Lys Asp
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Asp Gly Lys
195
<210>7
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aagaatttga tgtatctcgc tgcaattgcg gatgctcaac ctcagccacc agcagctaca 240
ctaacatcag gagccatgac tccccaagca atggctccta atccgtcatc aatgcagcca 300
ccaccaagct acttcatgca gcaacatcaa gctgtgggaa tggctcaaca aatacctcct 360
gggattttcc ctcctagagg tccattgcaa tttggtagcc cgcatcagtt tctggatccg 420
cagcaacagt tacatcaaca agctatgcaa gggcacatgg ggattagacc aatgggtttg 480
aataataaca acggactgca acatcaaatg caccaccatg aaactgctct tgccgcaaac 540
aatgcgggtc ctaacgatgc tagtggagga ggtaaaccgg atgggaccaa tatgagccag 600
agtggagctg atgggcaagg tggctcagcc gctagacatg gcggtggtga tgcaaaaact 660
gaaggaaaat ga 672
<210>8
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<212>PRT
<213>拟南芥
<400>8
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1 5 10 15
Pro Thr Asn Asn Ile Thr Thr Glu Gln Ile Gln Lys Tyr Leu Asp Glu
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Tyr Leu Ala Ala Ile Ala Asp Ala Gln Pro Gln Pro Pro Ala Ala Thr
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Leu Thr Ser Gly Ala Met Thr Pro Gln Ala Met Ala Pro Asn Pro Ser
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Ser Met Gln Pro Pro Pro Ser Tyr Phe Met Gln Gln His Gln Ala Val
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Leu Gln Phe Gly Ser Pro His Gln Phe Leu Asp Pro Gln Gln Gln Leu
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His Gln Gln Ala Met Gln Gly His Met Gly Ile Arg Pro Met Gly Leu
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Asn Asn Asn Asn Gly Leu Gln His Gln Met His His His Glu Thr Ala
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Pro Asp Gly Thr Asn Met Ser Gln Ser Gly Ala Asp Gly Gln Gly Gly
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<210>9
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aggaatctga tgtatcttgc agccattgcg gatagccagc cccaagttcc taccattgct 240
cagtatcctc ccaacgctgt tgctgctatg caatcgagtg ctcgctacat gcaacaacac 300
caagcagctc aacagatgac ccctcaatct ctcatggctg ctcgctcctc aatgctctac 360
tcacagtccc caatgtctgc actccagcag caacagcagc aagcagcaat gcatagccag 420
ctcgccatga gctccggagg caacaacagc agcaccggag gattcaccat tcttcatggt 480
gaagctagca taggaggcaa tggctcaatg aattctggtg gagtctttgg agattttgga 540
cggagcagcg gtgggaagca agagactggg agcgaagggc acgggacaga gactcctatg 600
tacctgaaag gctctgaaga agaaggaaac tga 633
<210>10
<211>210
<212>PRT
<213>Aspergillus officinalis
<400>10
Met Gln Gln His Leu Met Gln Met Gln Pro Met Met Ala Thr Tyr Gly
1 5 10 15
Ser Pro Asn Gln Val Thr Thr Asp Ile Ile Gln Gln Tyr Leu Asp Glu
20 25 30
Asn Lys Gln Leu Ile Leu Ala Ile Leu Glu Asn Gln Asn Ser Gly Lys
35 40 45
Ala Asp Glu Cys Ala Glu Asn Gln Ala Lys Leu Gln Arg Asn Leu Met
50 55 60
Tyr Leu Ala Ala Ile Ala Asp Ser Gln Pro Gln Val Pro Thr Ile Ala
65 70 75 80
Gln Tyr Pro Pro Asn Ala Val Ala Ala Met Gln Ser Ser Ala Arg Tyr
85 90 95
Met Gln Gln His Gln Ala Ala Gln Gln Met Thr Pro Gln Ser Leu Met
100 105 110
Ala Ala Arg Ser Ser Met Leu Tyr Ser Gln Ser Pro Met Ser Ala Leu
115 120 125
Gln Gln Gln Gln Gln Gln Ala Ala Met His Ser Gln Leu Ala Met Ser
130 135 140
Ser Gly Gly Asn Asn Ser Ser Thr Gly Gly Phe Thr Ile Leu His Gly
145 150 155 160
Glu Ala Ser Ile Gly Gly Asn Gly Ser Met Asn Ser Gly Gly Val Phe
165 170 175
Gly Asp Phe Gly Arg Ser Ser Gly Gly Lys Gln Glu Thr Gly Ser Glu
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Gly His Gly Thr Glu Thr Pro Met Tyr Leu Lys Gly Ser Glu Glu Glu
195 200 205
Gly Asn
210
<210>11
<211>591
<212>DNA
<213>欧洲油菜(Brassica napus)
<400>11
atgcagccca tgatggctgg ttactacccc agcaatgtca cctctgatca tatccagcag 60
tacttggatg agaacaagtc tttgattctg aagatagttg agtctcaaaa ctcaggaaag 120
ctcagcgagt gtgccgagaa tcaggcaagg cttcaacgca acctcatgta cttggctgca 180
atagcagatt ctcagcctca acctccaagc gtgcatagcc agtatggatc tgctggtggt 240
gggttgattc agggagaagg agcgtcacac tatttgcagc agcaacaggc gactcaacag 300
cagcagatga ctcagcagtc tcttatggca gctcgttctt caatgatgta tcagcagcag 360
caacagcctt atgcaacgct tcagcatcag cagttgcacc atagccagct tgggatgagc 420
tctagcagcg gaggaggaag cagtggtctc catatccttc agggagaggc tggtgggttt 480
catgaatttg gccgtgggaa gccggagatg ggaagtggtg aaggcagggg tggaagctca 540
ggggatggtg gagaaacact ctacttgaag tcatcagatg atgggaactg a 591
<210>12
<211>203
<212>PRT
<213>欧洲油菜
<400>12
Met Gln Gln His Leu Met Gln Met Gln Pro Met Met Ala Gly Tyr Tyr
1 5 10 15
Pro Ser Asn Val Thr Ser Asp His Ile Gln Gln Tyr Leu Asp Glu Asn
20 25 30
Lys Ser Leu Ile Leu Lys Ile Val Glu Ser Gln Asn Ser Gly Lys Leu
35 40 45
Ser Glu Cys Ala Glu Asn Gln Ala Arg Leu Gln Arg Ash Leu Met Tyr
50 55 60
Leu Ala Ala Ile Ala Asp Ser Gln Pro Gln Pro Pro Ser Val His Ser
65 70 75 80
Gln Tyr Gly Ser Ala Gly Gly Gly Leu Ile Gln Gly Glu Gly Ala Ser
85 90 95
His Tyr Leu Gln Gln Gln Gln Ala Thr Gln Gln Gln Gln Met Thr Gln
100 105 110
Gln Ser Leu Met Ala Ala Arg Ser Ser Met Met Tyr Gln Gln Gln Gln
115 120 125
Gln Pro Tyr Ala Thr Leu Gln His Gln Gln Leu His His Ser Gln Leu
130 135 140
Gly Met Ser Ser Ser Ser Gly Gly Gly Ser Ser Gly Leu His Ile Leu
145 150 155 160
Gln Gly Glu Ala Gly Gly Phe His Glu Phe Gly Arg Gly Lys Pro Glu
165 170 175
Met Gly Ser Gly Glu Gly Arg Gly Gly Ser Ser Gly Asp Gly Gly Glu
180 185 190
Thr Leu Tyr Leu Lys Ser Ser Asp Asp Gly Asn
195 200
<210>13
<211>663
<212>DNA
<213>甜橙(Citrus sinensis)
<400>13
atgcaacagc acctgatgca gatgcagccc atgatggcag cttattatcc caacaacgtc 60
actactgacc acattcaaca gtatctagat gagaacaaat cattgatttt gaagattgtt 120
gagagccaga attcagggaa actgagcgag tgtgcagaga accaggcaag attgcagcgg 180
aatctcatgt acctggctgc tattgctgat gctcaacccc aaccacctag cgttcatgcc 240
cagttctctt ctggtggcat tatgcagcca ggagctcact atatgcaaca ccagcaatct 300
cagccaatga caccacagtc acttatggct gcacgctcat ccatggtgta ctctcaacag 360
caattttcag tgcttcagca acagcaagcc ttgcatggtc agcttggcat gagctctggt 420
ggtagctcag gacttcacat gctgcaaagt gagggtagta ctgcaggagg tagtggttca 480
cttgggggtg ggggattccc tgattttggc cgtggctcat ctggtgaagg cttgcactca 540
aggggaatgg ggagcaagca tgatataggc agttctggat ctgctgaagg acgaggaggg 600
agctcaggaa gccaagatgg aggcgaaact ctctacttga aaggggctga tgatggaaat 660
taa 663
<210>14
<211>219
<212>PRT
<213>甜橙
<400>14
Met Gln Gln His Leu Met Gln Met Gln Pro Met Met Ala Ala Tyr Tyr
1 5 10 15
Pro Asn Asn Val Thr Thr Asp His Ile Gln Gln Tyr Leu Asp Glu Asn
20 25 30
Lys Ser Leu Ile Leu Lys Ile Val Glu Ser Gln Asn Ser Gly Lys Leu
35 40 45
Ser Glu Cys Ala Glu Asn Gln Ala Arg Leu Gln Arg Asn Leu Met Tyr
50 55 60
Leu Ala Ala Ile Ala Asp Ala Gln Pro Gln Pro Pro Ser Val His Ala
65 70 75 80
Gln Phe Ser Ser Gly Gly Ile Met Gln Pro Gly Ala His Tyr Met Gln
85 90 95
His Gln Gln Ser Gln Pro Met Thr Pro Gln Ser Leu Met Ala Ala Arg
100 105 110
Ser Ser Met Val Tyr Ser Gln Gln Gln Phe Ser Val Leu Gln Gln Gln
115 120 125
Gln Ala Leu His Gly Gln Leu Gly Met Ser Ser Gly Gly Ser Ser Gly
130 135 140
Leu His Met Leu Gln Ser Glu Gly Ser Thr Ala Gly Gly Ser Gly Ser
145 150 155 160
Leu Gly Gly Gly Gly Phe Pro Asp Phe Gly Arg Gly Ser Ser Gly Glu
165 170 175
Gly Leu His Ser Arg Gly Met Gly Ser Lys His Asp Ile Gly Ser Ser
180 185 190
Gly Ser Ala Glu Gly Arg Gly Gly Ser Ser Gly Ser Gln Asp Gly Gly
195 200 205
Glu Thr Leu Tyr Leu Lys Gly Ala Asp Asp Gly
210 215
<210>15
<211>660
<212>DNA
<213>树棉(Gossypium arboreum)
<220>
<221>misc_feature
<222>(309)..(309)
<223>n为a、c、g或t
<400>15
atgcagcagc acctgatgca gatgcagccc atgatggcag cttattatcc caacaacgtc 60
actactgatc atattcaaca gtatctcgat gagaacaagt cattgatctt aaagattgtt 120
gagagccaga attctgggaa attgagtgaa tgtgctgaga accaagcaag gctgcagcga 180
aacctcatgt acctggctgc cattgcggat tctcaacccc aaccacccac cgtgcatgca 240
cagtttccat ctggtggtat catgcagcaa ggagctgggc actacatgca gcaccaacaa 300
gctcaacana tgacacaaca gtcgcttatg gctgctcggt cctcaatgtt gtattctcag 360
caaccatttt ctgcactgca acaacaacaa caacaaggct ttgcacagtc agcttggcat 420
gagctctggc gggagcacag gcctttcata tgctgcaaac tgaatctagt actgcagggg 480
gcagtgagac accttgggcc cgagggttgt cctgatttgg acgggggtct tttggagagg 540
catccctggt ggcaggccaa tggccggggg aacaaccaaa aatccgggga ggccggctca 600
cctaagggcc gggaggagcc cttggggcag gggggggtga tggggggaac ctcttcttaa 660
<210>16
<211>219
<212>PRT
<213>树棉
<220>
<221>misc_feature
<222>(103)..(103)
<223>Xaa可以是任何天然存在的氨基酸
<400>16
Met Gln Gln His Leu Met Gln Met Gln Pro Met Met Ala Ala Tyr Tyr
1 5 10 15
Pro Asn Asn Val Thr Thr Asp His Ile Gln Gln Tyr Leu Asp Glu Asn
20 25 30
Lys Ser Leu Ile Leu Lys Ile Val Glu Ser Gln Asn Ser Gly Lys Leu
35 40 45
Ser Glu Cys Ala Glu Asn Gln Ala Arg Leu Gln Arg Asn Leu Met Tyr
50 55 60
Leu Ala Ala Ile Ala Asp Ser Gln Pro Gln Pro Pro Thr Val His Ala
65 70 75 80
Gln Phe Pro Ser Gly Gly Ile Met Gln Gln Gly Ala Gly His Tyr Met
85 90 95
Gln His Gln Gln Ala Gln Xaa Met Thr Gln Gln Ser Leu Met Ala Ala
100 105 110
Arg Ser Ser Met Leu Tyr Ser Gln Gln Pro Phe Ser Ala Leu Gln Gln
115 120 125
Gln Gln Gln Gln Gly Phe Ala Gln Ser Ala Trp His Glu Leu Trp Arg
130 135 140
Glu His Arg Pro Phe Ile Cys Cys Lys Leu Asn Leu Val Leu Gln Gly
145 150 155 160
Ala Val Arg His Leu Gly Pro Glu Gly Cys Pro Asp Leu Asp Gly Gly
165 170 175
Leu Leu Glu Arg His Pro Trp Trp Gln Ala Asn Gly Arg Gly Asn Asn
180 185 190
Gln Lys Ser Gly Glu Ala Gly Ser Pro Lys Gly Arg Glu Glu Pro Leu
195 200 205
Gly Gln Gly Gly Val Met Gly Gly Thr Ser Ser
210 215
<210>17
<211>636
<212>DNA
<213>蒺藜状苜蓿(Medicago trunculata)
<400>17
atgcagcagc acctgatgca gatgcagccc atgatggcag cttactatcc taacaacgtc 60
actactgatc atattcaaca gtatcttgat gagaacaagt ccttgattct caagattgtt 120
gaaagccaga acactggcaa gctcaccgag tgtgctgaga accaatcaag gcttcagaga 180
aatctcatgt acctagctgc aatagctgat tctcaacccc aaccacctac tatgcctggc 240
cagtaccctt caagtggaat gatgcagcag ggaggacact acatgcaggc tcaacaagct 300
cagcagatga cacaacaaca attaatggct gcacgttcct ctcttatgta tgctcaacag 360
cttcaacagc agcaagcctt gcaaagccaa cttggtatga attccagtgg aagtcaaggc 420
cttcacatgt tgcatagtga aggggctaat gttggaggca attcatctct aggggctggt 480
tttcctgatt ttggccgtag ctcagccggt gatggtttgc acggcagtgg taagcaagac 540
attggaagca ctgatggccg cggtggaagc tctagtggtc actctggtga tggcggcgaa 600
acactttacc tgaaatcttc tggtgatggg aattag 636
<210>18
<211>211
<212>PRT
<213>蒺藜状苜蓿
<400>18
Met Gln Gln His Leu Met Gln Met Gln Pro Met Met Ala Ala Tyr Tyr
1 5 10 15
Pro Asn Asn Val Thr Thr Asp His Ile Gln Gln Tyr Leu Asp Glu Asn
20 25 30
Lys Ser Leu Ile Leu Lys Ile Val Glu Ser Gln Asn Thr Gly Lys Leu
35 40 45
Thr Glu Cys Ala Glu Asn Gln Ser Arg Leu Gln Arg Asn Leu Met Tyr
50 55 60
Leu Ala Ala Ile Ala Asp Ser Gln Pro Gln Pro Pro Thr Met Pro Gly
65 70 75 80
Gln Tyr Pro Ser Ser Gly Met Met Gln Gln Gly Gly His Tyr Met Gln
85 90 95
Ala Gln Gln Ala Gln Gln Met Thr Gln Gln Gln Leu Met Ala Ala Arg
100 105 110
Ser Ser Leu Met Tyr Ala Gln Gln Leu Gln Gln Gln Gln Ala Leu Gln
115 120 125
Ser Gln Leu Gly Met Asn Ser Ser Gly Ser Gln Gly Leu His Met Leu
130 135 140
His Ser Glu Gly Ala Asn Val Gly Gly Asn Ser Ser Leu Gly Ala Gly
145 150 155 160
Phe Pro Asp Phe Gly Arg Ser Ser Ala Gly Asp Gly Leu His Gly Ser
165 170 175
Gly Lys Gln Asp Ile Gly Ser Thr Asp Gly Arg Gly Gly Ser Ser Ser
180 185 190
Gly His Ser Gly Asp Gly Gly Glu Thr Leu Tyr Leu Lys Ser Ser Gly
195 200 205
Asp Gly Asn
210
<210>19
<211>684
<212>DNA
<213>稻(Oryza sativa)
<400>19
atgcagcagc aacacctgat gcagatgaac cagggcatga tggggggata tgcttcccct 60
accaccgtca ccactgatct cattcagcag tatctggatg agaacaagca gctgatcctg 120
gccatccttg acaaccagaa caatgggaag gtggaagagt gcgctcggaa ccaagctaag 180
ctccagcaca atctcatgta cctcgccgcc atcgccgaca gccagccgcc gcagacggcc 240
gccatgtccc agtatccgtc gaacctgatg atgcagtccg gggcgaggta catgccgcag 300
cagtcggcgc agatgatggc gccgcagtcg ctgatggcgg cgaggtcttc gatgatgtac 360
gcgcagccgg cgctgtcgcc gctccagcag cagcagcagc agcaggcggc ggcggcgcac 420
gggcagctgg gcatgggctc ggggggcacc accagcgggt tcagcatcct ccacggcgag 480
gccagcatgg gcggcggcgg cggcggcggt ggcgccggta acagcatgat gaacgccggc 540
gtgttctccg acttcggacg cggcggcggc ggcggcggca aggaggggtc cacctcgctg 600
tccgtcgacg tccggggcgc caactccggc gcccagagcg gcgacgggga gtacctcaag 660
ggcaccgagg aggaaggcag ctag 684
<210>20
<211>227
<212>PRT
<213>稻
<400>20
Met Gln Gln Gln His Leu Met Gln Met Asn Gln Gly Met Met Gly Gly
1 5 10 15
Tyr Ala Ser Pro Thr Thr Val Thr Thr Asp Leu Ile Gln Gln Tyr Leu
20 25 30
Asp Glu Asn Lys Gln Leu Ile Leu Ala Ile Leu Asp Asn Gln Asn Asn
35 40 45
Gly Lys Val Glu Glu Cys Ala Arg Asn Gln Ala Lys Leu Gln His Asn
50 55 60
Leu Met Tyr Leu Ala Ala Ile Ala Asp Ser Gln Pro Pro Gln Thr Ala
65 70 75 80
Ala Met Ser Gln Tyr Pro Ser Asn Leu Met Met Gln Ser Gly Ala Arg
85 90 95
Tyr Met Pro Gln Gln Ser Ala Gln Met Met Ala Pro Gln Ser Leu Met
100 105 110
Ala Ala Arg Ser Ser Met Met Tyr Ala Gln Pro Ala Leu Ser Pro Leu
115 120 125
Gln Gln Gln Gln Gln Gln Gln Ala Ala Ala Ala His Gly Gln Leu Gly
130 135 140
Met Gly Ser Gly Gly Thr Thr Ser Gly Phe Ser Ile Leu His Gly Glu
145 150 155 160
Ala Ser Met Gly Gly Gly Gly Gly Gly Gly Gly Ala Gly Asn Ser Met
165 170 175
Met Asn Ala Gly Val Phe Ser Asp Phe Gly Arg Gly Gly Gly Gly Gly
180 185 190
Gly Lys Glu Gly Ser Thr Ser Leu Ser Val Asp Val Arg Gly Ala Asn
195 200 205
Ser Gly Ala Gln Ser Gly Asp Gly Glu Tyr Leu Lys Gly Thr Glu Glu
210 215 220
Glu Gly Ser
225
<210>21
<211>558
<212>DNA
<213>稻
<400>21
atgcagcagc agccgatgcc gatgcccgcg caggcgccgc cgacggccgg aatcaccacc 60
gagcagatcc aaaagtatct ggatgaaaac aagcagctta ttttggctat tttggaaaat 120
cagaatctgg gaaagttggc agaatgtgct cagtatcaag cgcagcttca gaagaatctc 180
ttgtacttgg ctgcaattgc tgatactcaa ccgcagacca ctataagccg tccccagatg 240
gtgccgcatg gtgcatcgcc ggggttaggg gggcaataca tgtcgcaggt gccaatgttc 300
ccccccagga cccctctaac gccccagcag atgcaggagc agcagctgca gcaacagcaa 360
gcccagctgc tctcgttcgg cggtcagatg gttatgaggc ctggcgttgt gaatggcatt 420
cctcagcttc tgcaaggcga aatgcaccgc ggagcagatc accagaacgc tggcggggcc 480
acctcggagc cttccgagag ccacaggagc accggcaccg aaastgacgg tggaagcgac 540
ttcggcgatc aatcctaa 558
<210>22
<211>185
<212>PRT
<213>稻
<400>22
Met Gln Gln Gln Pro Met Pro Met Pro Ala Gln Ala Pro Pro Thr Ala
1 5 10 15
Gly Ile Thr Thr Glu Gln Ile Gln Lys Tyr Leu Asp Glu Asn Lys Gln
20 25 30
Leu Ile Leu Ala Ile Leu Glu Asn Gln Asn Leu Gly Lys Leu Ala Glu
35 40 45
Cys Ala Gln Tyr Gln Ala Gln Leu Gln Lys Asn Leu Leu Tyr Leu Ala
50 55 60
Ala Ile Ala Asp Thr Gln Pro Gln Thr Thr Ile Ser Arg Pro Gln Met
65 70 75 80
Val Pro His Gly Ala Ser Pro Gly Leu Gly Gly Gln Tyr Met Ser Gln
85 90 95
Val Pro Met Phe Pro Pro Arg Thr Pro Leu Thr Pro Gln Gln Met Gln
100 105 110
Glu Gln Gln Leu Gln Gln Gln Gln Ala Gln Leu Leu Ser Phe Gly Gly
115 120 125
Gln Met Val Met Arg Pro Gly Val Val Asn Gly Ile Pro Gln Leu Leu
130 135 140
Gln Gly Glu Met His Arg Gly Ala Asp His Gln Asn Ala Gly Gly Ala
145 150 155 160
Thr Ser Glu Pro Ser Glu Ser His Arg Ser Thr Gly Thr Glu Asn Asp
165 170 175
Gly Gly Ser Asp Phe Gly Asp Gln Ser
180 185
<210>23
<211>618
<212>DNA
<213>稻
<400>23
atgcagcagc agatggccat gccggcgggg gccgccgccg ccgcggtgcc gccggcggcc 60
ggcatcacca ccgagcagat ccaaaagtat ttggatgaaa ataaacagct aattttggcc 120
atcctggaaa atcaaaacct agggaagttg gctgaatgtg ctcagtacca agctcagctt 180
caaaagaatc tcttgtatct ggctgccatt gcagatgccc aaccacctca gaatccagga 240
agtcgccctc agatgatgca gcctggtgct accccaggtg ctgggcatta catgtcccaa 300
gtaccgatgt tccctccaag aactccctta accccacaac agatgcaaga gcagcagcag 360
cagcaactcc agcaacagca agctcaggct ctagccttcc ccggccagat gctaatgaga 420
ccaggtactg tcaatggcat gcaatctatc ccagttgctg accctgctcg cgcagccgat 480
cttcagacgg cagcaccggg ctcggtagat ggccgaggaa acaagcagga tgcaacctcg 540
gagccttccg ggaccgagag ccacaagagt gcgggagcag ataacgacgc aggcggtgac 600
atagcggaga agtcctga 618
<210>24
<211>205
<212>PRT
<213>稻
<400>24
Met Gln Gln Gln Met Ala Met Pro Ala Gly Ala Ala Ala Ala Ala Val
1 5 10 15
Pro Pro Ala Ala Gly Ile Thr Thr Glu Gln Ile Gln Lys Tyr Leu Asp
20 25 30
Glu Asn Lys Gln Leu Ile Leu Ala Ile Leu Glu Asn Gln Asn Leu Gly
35 40 45
Lys Leu Ala Glu Cys Ala Gln Tyr Gln Ala Gln Leu Gln Lys Asn Leu
50 55 60
Leu Tyr Leu Ala Ala Ile Ala Asp Ala Gln Pro Pro Gln Asn Pro Gly
65 70 75 80
Ser Arg Pro Gln Met Met Gln Pro Gly Ala Thr Pro Gly Ala Gly His
85 90 95
Tyr Met Ser Gln Val Pro Met Phe Pro Pro Arg Thr Pro Leu Thr Pro
100 105 110
Gln Gln Met Gln Glu Gln Gln Gln Gln Gln Leu Gln Gln Gln Gln Ala
115 120 125
Gln Ala Leu Ala Phe Pro Gly Gln Met Leu Met Arg Pro Gly Thr Val
130 135 140
Asn Gly Met Gln Ser Ile Pro Val Ala Asp Pro Ala Arg Ala Ala Asp
145 150 155 160
Leu Gln Thr Ala Ala Pro Gly Ser Val Asp Gly Arg Gly Asn Lys Gln
165 170 175
Asp Ala Thr Ser Glu Pro Ser Gly Thr Glu Ser His Lys Ser Ala Gly
180 185 190
Ala Asp Asn Asp Ala Gly Gly Asp Ile Ala Glu Lys Ser
195 200 205
<210>25
<211>540
<212>DNA
<213>马铃薯(Solanum tuberosum)
<400>25
atgcagcagc agcacctgat gcagatgcag cccatgatgg cagcctatta tcccaacaat 60
gtcactactg atcatattca acagttcctg gatgagaaca aatcacttat tctgaagatt 120
gttgagagcc agaactctgg gaaaataagt gaatgtgcag agtcccaagc taaacttcag 180
agaaatctta tgtaccttgc agctattgct gattcacagc cccagcctcc tagtatgcat 240
tcacagttag cttctggtgg gatgatgcag ggaggggcac attatatgca gcaacaacaa 300
gctcaacaac tcacaacgca atcgcttatg gctgcagcaa gatcctcctc ctcaatgctc 360
tatggacaac aacaacaaca acaacaacaa caactatcat cattgcaaca acagcaagca 420
gcctttcata gccagcaact cggaatgagc agctctggtg gaggaagcag tagtggactt 480
cacatgctac aaagcgaaaa cactcatagt gctagcactg gtggtgggtg gtttccctga 540
<210>26
<211>179
<212>PRT
<213>马铃薯
<400>26
Met Gln Gln Gln His Leu Met Gln Met Gln Pro Met Met Ala Ala Tyr
1 5 10 15
Tyr Pro Asn Asn Val Thr Thr Asp His Ile Gln Gln Phe Leu Asp Glu
20 25 30
Asn Lys Ser Leu Ile Leu Lys Ile Val Glu Ser Gln Asn Ser Gly Lys
35 40 45
Ile Ser Glu Cys Ala Glu Ser Gln Ala Lys Leu Gln Arg Asn Leu Met
50 55 60
Tyr Leu Ala Ala Ile Ala Asp Ser Gln Pro Gln Pro Pro Ser Met His
65 70 75 80
Ser Gln Leu Ala Ser Gly Gly Met Met Gln Gly Gly Ala His Tyr Met
85 90 95
Gln Gln Gln Gln Ala Gln Gln Leu Thr Thr Gln Ser Leu Met Ala Ala
100 105 110
Ala Arg Ser Ser Ser Ser Met Leu Tyr Gly Gln Gln Gln Gln Gln Gln
115 120 125
Gln Gln Gln Leu Ser Ser Leu Gln Gln Gln Gln Ala Ala Phe His Ser
130 135 140
Gln Gln Leu Gly Met Ser Ser Ser Gly Gly Gly Ser Ser Ser Gly Leu
145 150 155 160
His Met Leu Gln Ser Glu Asn Thr His Ser Ala Ser Thr Gly Gly Gly
165 170 175
Trp Phe Pro
<210>27
<211>684
<212>DNA
<213>玉蜀黍(Zea mays)
<400>27
atgcagcagc aacacctgat gcagatgaac cagaacatga tggggggcta cacctctcct 60
gccgccgtga ccaccgatct catccagcag cacctggacg agaacaagca gctgatcctg 120
gccatcctcg acaaccagaa caatggcaag gcggaggagt gcgaacggca ccaagctaag 180
ctccagcaca acctcatgta cctggccgcc atcgctgaca gccagccgcc acagaccgcg 240
ccactatcac agtacccgtc caacctgatg atgcagccgg gccctcggta catgccaccg 300
cagtccgggc agatgatgaa cccgcagtcg ctgatggcgg cgcggtcctc catgatgtac 360
gcgcacccgt ccctgtcgcc actccagcag cagcaggcgg cgcacggaca gctgggtatg 420
gctccagggg gcggcggtgg cggcacgacc agcgggttca gcatcctcca cggcgaggcc 480
agcatgggcg gtggtggtgc tggcgcaggc gccggcaaca acatgatgaa cgccggcatg 540
ttctcgggct ttggccgcag cggcagtggc gccaaggaag ggtcgacctc tctgtcggtt 600
gacgtccggg gtggaaccag ctccggcgcg cagagcgggg acggcgagta cctcaaagtc 660
ggcaccgagg aagaaggcag ttag 684
<210>28
<211>227
<212>PRT
<213>玉蜀黍
<400>28
Met Gln Gln Gln His Leu Met Gln Met Asn Gln Asn Met Met Gly Gly
1 5 10 15
Tyr Thr Ser Pro Ala Ala Val Thr Thr Asp Leu Ile Gln Gln His Leu
20 25 30
Asp Glu Asn Lys Gln Leu Ile Leu Ala Ile Leu Asp Asn Gln Asn Asn
35 40 45
Gly Lys Ala Glu Glu Cys Glu Arg His Gln Ala Lys Leu Gln His Asn
50 55 60
Leu Met Tyr Leu Ala Ala Ile Ala Asp Ser Gln Pro Pro Gln Thr Ala
65 70 75 80
Pro Leu Ser Gln Tyr Pro Ser Asn Leu Met Met Gln Pro Gly Pro Arg
85 90 95
Tyr Met Pro Pro Gln Ser Gly Gln Met Met Asn Pro Gln Ser Leu Met
100 105 110
Ala Ala Arg Ser Ser Met Met Tyr Ala His Pro Ser Leu Ser Pro Leu
115 120 125
Gln Gln Gln Gln Ala Ala His Gly Gln Leu Gly Met Ala Pro Gly Gly
130 135 140
Gly Gly Gly Gly Thr Thr Ser Gly Phe Ser Ile Leu His Gly Glu Ala
145 150 155 160
Ser Met Gly Gly Gly Gly Ala Gly Ala Gly Ala Gly Asn Asn Met Met
165 170 175
Asn Ala Gly Met Phe Ser Gly Phe Gly Arg Ser Gly Ser Gly Ala Lys
180 185 190
Glu Gly Ser Thr Ser Leu Ser Val Asp Val Arg Gly Gly Thr Ser Ser
195 200 205
Gly Ala Gln Ser Gly Asp Gly Glu Tyr Leu Lys Val Gly Thr Glu Glu
210 215 220
Glu Gly Ser
225
<210>29
<211>549
<212>DNA
<213>玉蜀黍
<400>29
atgcagcagc cgatgcacat gcagccacag gcgccggcga taaccccagc tgccggaatc 60
agcacggagc agatccaaaa gtatctggat gagaataagc agcttatttt ggctattttg 120
gaaaatcaga acctaggaaa attggcagaa tgtgctcagt atcaatcaca acttcagaag 180
aacctcttgt atctcgctgc aatcgcagat gctcaaccgc agactgctgt aagccgccct 240
cagatggcgc cgcctggtgg atcgcctgga gtagggcagt acatgtcaca ggtgcctatg 300
ttcccaccga ggacacctct tacaccccag cagatgcagg agcagcagct tcagcagcag 360
caggctcagt tgctaaactt cagtggccaa atggttgcta gaccaggcat ggtcaacggc 420
atggctcagt ccatgcaagc tcagctacca ccgggtgtga acaagcagga tgctggtggg 480
gtcgcctctg agccctcggg caccgagagc cacaggagca ctggtggtga cgatggtgga 540
agcgactag 549
<210>30
<211>182
<212>PRT
<213>玉蜀黍
<400>30
Met Gln Gln Pro Met His Met Gln Pro Gln Ala Pro Ala Ile Thr Pro
1 5 10 15
Ala Ala Gly Ile Ser Thr Glu Gln Ile Gln Lys Tyr Leu Asp Glu Asn
20 25 30
Lys Gln Leu Ile Leu Ala Ile Leu Glu Asn Gln Asn Leu Gly Lys Leu
35 40 45
Ala Glu Cys Ala Gln Tyr Gln Ser Gln Leu Gln Lys Asn Leu Leu Tyr
50 55 60
Leu Ala Ala Ile Ala Asp Ala Gln Pro Gln Thr Ala Val Ser Arg Pro
65 70 75 80
Gln Met Ala Pro Pro Gly Gly Ser Pro Gly Val Gly Gln Tyr Met Ser
85 90 95
Gln Val Pro Met Phe Pro Pro Arg Thr Pro Leu Thr Pro Gln Gln Met
100 105 110
Gln Glu Gln Gln Leu Gln Gln Gln Gln Ala Gln Leu Leu Asn Phe Ser
115 120 125
Gly Gln Met Val Ala Arg Pro Gly Met Val Asn Gly Met Ala Gln Ser
130 135 140
Met Gln Ala Gln Leu Pro Pro Gly Val Asn Lys Gln Asp Ala Gly Gly
145 150 155 160
Val Ala Ser Glu Pro Ser Gly Thr Glu Ser His Arg Ser Thr Gly Gly
165 170 175
Asp Asp Gly Gly Ser Asp
180
<210>31
<211>1173
<212>DNA
<213>智人(Homo sapiens)
<400>31
atgggcggca acatgtctgt ggctttcgcg gccccgaggc agcgaggcaa gggggagatc 60
actcccgctg cgattcagaa gatgttggat gacaataacc atcttattca gtgtataatg 120
gactctcaga ataaaggaaa gacctcagag tgttctcagt atcagcagat gttgcacaca 180
aacttggtat accttgctac aatagcagat tctaatcaaa atatgcagtc tcttttacca 240
gcaccaccca cacagaatat gcctatgggt cctggaggga tgaatcagag cggccctccc 300
ccacctccac gctctcacaa catgccttca gatggaatgg taggtggggg tcctcctgca 360
ccgcacatgc agaaccagat gaacggccag atgcctgggc ctaaccatat gcctatgcag 420
ggacctggac ccaatcaact caatatgaca aacagttcca tgaatatgcc ttcaagtagc 480
catggatcca tgggaggtta caaccattct gtgccatcat cacagagcat gccagtacag 540
aatcagatga caatgagtca gggacaacca atgggaaact atggtcccag accaaatatg 600
agtatgcagc caaaccaagg tccaatgatg catcagcagc ctccttctca gcaatacaat 660
atgccacagg gaggcggaca gcattaccaa ggacagcagc cacctatggg aatgatgggt 720
caagttaacc aaggcaatca tatgatgggt cagagacaga ttcctcccta tagacctcct 780
caacagggcc caccacagca gtactcaggc caggaagact attacgggga ccaatacagt 840
catggtggac aaggtcctcc agaaggcatg aaccagcaat attaccctga tggaaattca 900
cagtatggcc aacagcaaga tgcataccag ggaccacctc cacaacaggg atatccaccc 960
cagcagcagc agtacccagg gcagcaaggt tacccaggac agcagcaggg ctacggtcct 1020
tcacagggtg gtccaggtcc tcagtatcct aactacccac agggacaagg tcagcagtat 1080
ggaggatata gaccaacaca gcctggacca ccacagccac cccagcagag gccttatgga 1140
tatgaccagg gacagtatgg aaattaccag cag 1173
<210>32
<211>391
<212>PRT
<213>智人
<400>32
Met Gly Gly Asn Met Ser Val Ala Phe Ala Ala Pro Arg Gln Arg Gly
1 5 10 15
Lys Gly Glu Ile Thr Pro Ala Ala Ile Gln Lys Met Leu Asp Asp Asn
20 25 30
Asn His Leu Ile Gln Cys Ile Met Asp Ser Gln Asn Lys Gly Lys Thr
35 40 45
Ser Glu Cys Ser Gln Tyr Gln Gln Met Leu His Thr Asn Leu Val Tyr
50 55 60
Leu Ala Thr Ile Ala Asp Ser Asn Gln Asn Met Gln Ser Leu Leu Pro
65 70 75 80
Ala Pro Pro Thr Gln Asn Met Pro Met Gly Pro Gly Gly Met Asn Gln
85 90 95
Ser Gly Pro Pro Pro Pro Pro Arg Ser His Asn Met Pro Ser Asp Gly
100 105 110
Met Val Gly Gly Gly Pro Pro Ala Pro His Met Gln Asn Gln Met Asn
115 120 125
Gly Gln Met Pro Gly Pro Asn His Met Pro Met Gln Gly Pro Gly Pro
130 135 140
Asn Gln Leu Asn Met Thr Asn Ser Ser Met Asn Met Pro Ser Ser Ser
145 150 155 160
His Gly Ser Met Gly Gly Tyr Asn His Ser Val Pro Ser Ser Gln Ser
165 170 175
Met Pro Val Gln Asn Gln Met Thr Met Ser Gln Gly Gln Pro Met Gly
180 185 190
Asn Tyr Gly Pro Arg Pro Asn Met Ser Met Gln Pro Asn Gln Gly Pro
195 200 205
Met Met His Gln Gln Pro Pro Ser Gln Gln Tyr Asn Met Pro Gln Gly
210 215 220
Gly Gly Gln His Tyr Gln Gly Gln Gln Pro Pro Met Gly Met Met Gly
225 230 235 240
Gln Val Asn Gln Gly Asn His Met Met Gly Gln Arg Gln Ile Pro Pro
245 250 255
Tyr Arg Pro Pro Gln Gln Gly Pro Pro Gln Gln Tyr Ser Gly Gln Glu
260 265 270
Asp Tyr Tyr Gly Asp Gln Tyr Ser His Gly Gly Gln Gly Pro Pro Glu
275 280 285
Gly Met Asn Gln Gln Tyr Tyr Pro Asp Gly Asn Ser Gln Tyr Gly Gln
290 295 300
Gln Gln Asp Ala Tyr Gln Gly Pro Pro Pro Gln Gln Gly Tyr Pro Pro
305 310 315 320
Gln Gln Gln Gln Tyr Pro Gly Gln Gln Gly Tyr Pro Gly Gln Gln Gln
325 330 335
Gly Tyr Gly Pro Ser Gln Gly Gly Pro Gly Pro Gln Tyr Pro Asn Tyr
340 345 350
Pro Gln Gly Gln Gly Gln Gln Tyr Gly Gly Tyr Arg Pro Thr Gln Pro
355 360 365
Gly Pro Pro Gln Pro Pro Gln Gln Arg Pro Tyr Gly Tyr Asp Gln Gly
370 375 380
Gln Tyr Gly Asn Tyr Gln Gln
385 390
<210>33
<211>627
<212>DNA
<213>洋葱(Allium cepa)
<400>33
atgcagcagc cgcagccagc gatgggaacc atgggctcgg tgccacctac tagcatcacc 60
accgaacaga ttcaaaggta cttggatgag aacaaacagt taatattggc aattttggat 120
aatcaaaatt taggaagact gaatgagtgt gctcaatatc aagctcagct tcaaaagaat 180
ctgctttacc tggcagcaat agctgatgct cagcctcagt ctcctgcggt gcgtctgcag 240
atgatgcctc aaggtgcagc tgccacgcct caagctggaa accaatttat gcagcagcag 300
agccctaatt tccctcccaa aacaggaatg caatttactc ctcaacaagt acaagaattg 360
cagcagcaac agctacaaca tcagccacat atgatgcctc catttcaagg tcaaatgggt 420
atgagaccta tgaatggaat gcaggcagca atgcatgcag attcatctct tgcttataac 480
actaacaata agcaagatgc aggaaacgca gcttatgaaa atactgctgc caacacagat 540
ggttccattc aaaagaaaac agcaaatgat gatttagacc cttctgcagc aaaccctaga 600
aggtctgaag atgccaaatc atcatga 627
<210>34
<211>208
<212>PRT
<213>洋葱
<400>34
Met Gln Gln Pro Gln Pro Ala Met Gly Thr Met Gly Ser Val Pro Pro
1 5 10 15
Thr Ser Ile Thr Thr Glu Gln Ile Gln Arg Tyr Leu Asp Glu Asn Lys
20 25 30
Gln Leu Ile Leu Ala Ile Leu Asp Asn Gln Asn Leu Gly Arg Leu Asn
35 40 45
Glu Cys Ala Gln Tyr Gln Ala Gln Leu Gln Lys Asn Leu Leu Tyr Leu
50 55 60
Ala Ala Ile Ala Asp Ala Gln Pro Gln Ser Pro Ala Val Arg Leu Gln
65 70 75 80
Met Met Pro Gln Gly Ala Ala Ala Thr Pro Gln Ala Gly Asn Gln Phe
85 90 95
Met Gln Gln Gln Ser Pro Asn Phe Pro Pro Lys Thr Gly Met Gln Phe
100 105 110
Thr Pro Gln Gln Val Gln Glu Leu Gln Gln Gln Gln Leu Gln His Gln
115 120 125
Pro His Met Met Pro Pro Phe Gln Gly Gln Met Gly Met Arg Pro Met
130 135 140
Asn Gly Met Gln Ala Ala Met His Ala Asp Ser Ser Leu Ala Tyr Asn
145 150 155 160
Thr Asn Asn Lys Gln Asp Ala Gly Asn Ala Ala Tyr Glu Asn Thr Ala
165 170 175
Ala Asn Thr Asp Gly Ser Ile Gln Lys Lys Thr Ala Asn Asp Asp Leu
180 185 190
Asp Pro Ser Ala Ala Asn Pro Arg Arg Ser Glu Asp Ala Lys Ser Ser
195 200 205
<210>35
<211>633
<212>DNA
<213>Aquilegia formosa x Aquilegia pubescens
<400>35
atgcaacaca tgcagatgca gcccatgatg ccaccttata gtgccaacag cgtcactact 60
gatcatatcc aacagtactt ggatgaaaat aaggcgttga ttctgaagat acttgagaac 120
caaaattcgg gaaaagttag tgaatgtgca gagaaccaag caagacttca acgaaatctt 180
atgtatctgg ctgcaattgc tgattctcaa ccacagcctc ccaatatgca tgctcagtac 240
tctaatgcgg gtataccacc tggtgcacat tacctacaac accaacaggc ccaacagatg 300
acacaacagt cgctcatggc tgctcgatca aatatgctgt atgctcagcc aatcacagga 360
atgcagcaac agcaagcaat gcatagccag cttggcatga gctctggtgg taacagtgga 420
ctccacatga tgcacaatga gggcagcatg ggaggtagtg gggcacttgg aagctattct 480
gattatggcc gtggcagtgg tggtggagta actatcgcta gcaaacaaga tggtggaagt 540
ggttctggtg aaggacgagg tggaaactct ggaggccaaa gtgcagatgg aggtgaatct 600
ctttacctga aaaacagtga cgaagggaac taa 633
<210>36
<211>210
<212>PRT
<213>Aquilegia formosa x Aquilegia pubescens
<400>36
Met Gln His Met Gln Met Gln Pro Met Met Pro Pro Tyr Ser Ala Asn
1 5 10 15
Ser Val Thr Thr Asp His Ile Gln Gln Tyr Leu Asp Glu Asn Lys Ala
20 25 30
Leu Ile Leu Lys Ile Leu Glu Asn Gln Asn Ser Gly Lys Val Ser Glu
35 40 45
Cys Ala Glu Asn Gln Ala Arg Leu Gln Arg Asn Leu Met Tyr Leu Ala
50 55 60
Ala Ile Ala Asp Ser Gln Pro Gln Pro Pro Asn Met His Ala Gln Tyr
65 70 75 80
Ser Asn Ala Gly Ile Pro Pro Gly Ala His Tyr Leu Gln His Gln Gln
85 90 95
Ala Gln Gln Met Thr Gln Gln Ser Leu Met Ala Ala Arg Ser Asn Met
100 105 110
Leu Tyr Ala Gln Pro Ile Thr Gly Met Gln Gln Gln Gln Ala Met His
115 120 125
Ser Gln Leu Gly Met Ser Ser Gly Gly Asn Ser Gly Leu His Met Met
130 135 140
His Asn Glu Gly Ser Met Gly Gly Ser Gly Ala Leu Gly Ser Tyr Ser
145 150 155 160
Asp Tyr Gly Arg Gly Ser Gly Gly Gly Val Thr Ile Ala Ser Lys Gln
165 170 175
Asp Gly Gly Ser Gly Ser Gly Glu Gly Arg Gly Gly Asn Ser Gly Gly
180 185 190
Gln Ser Ala Asp Gly Gly Glu Ser Leu Tyr Leu Lys Asn Ser Asp Glu
195 200 205
Gly Asn
210
<210>37
<211>615
<212>DNA
<213>二穗短柄草(Brachypodium distachyon)
<400>37
atgcagcagg cgatgtccat gtccccgggg tcggccggcg cggtgccgcc tccggccggc 60
atcaccacag agcagatcca aaagtatttg gatgaaaata agcaacttat tttggccatc 120
ctggaaaatc agaacctagg aaagttgact gaatgtgctc agtatcaagc tcaacttcag 180
aagaatctct tgtatctggc tgccattgcg gatgcccaac caccacagaa ccctggaagt 240
cgcccccaga tggtgcagcc tggtggtatg ccaggtgcag ggcattacat gtcgcaagta 300
ccaatgttcc ctccaagaac ccctttaacc ccacaacaga tgcaagagca acagcaccag 360
cagcttcagc agcagcaagc acaggctctt gctttcccca gccagatggt catgagacca 420
ggtactgtga acggcatgca gcctatgcaa gctgatctcc aagcagcagc agcagcacct 480
ggcctggcag acagccgagg aagtaagcag gacgcagcgg tagctggggc catctcggaa 540
ccttctggca ccgagagtca caagagtaca ggagcggatc atgaggcagg tggcgatgta 600
gctgagcaat cctaa 615
<210>38
<211>204
<212>PRT
<213>二穗短柄草
<400>38
Met Gln Gln Ala Met Ser Met Ser Pro Gly Ser Ala Gly Ala Val Pro
1 5 10 15
Pro Pro Ala Gly Ile Thr Thr Glu Gln Ile Gln Lys Tyr Leu Asp Glu
20 25 30
Asn Lys Gln Leu Ile Leu Ala Ile Leu Glu Asn Gln Asn Leu Gly Lys
35 40 45
Leu Thr Glu Cys Ala Gln Tyr Gln Ala Gln Leu Gln Lys Asn Leu Leu
50 55 60
Tyr Leu Ala Ala Ile Ala Asp Ala Gln Pro Pro Gln Asn Pro Gly Ser
65 70 75 80
Arg Pro Gln Met Val Gln Pro Gly Gly Met Pro Gly Ala Gly His Tyr
85 90 95
Met Ser Gln Val Pro Met Phe Pro Pro Arg Thr Pro Leu Thr Pro Gln
100 105 110
Gln Met Gln Glu Gln Gln His Gln Gln Leu Gln Gln Gln Gln Ala Gln
115 120 125
Ala Leu Ala Phe Pro Ser Gln Met Val Met Arg Pro Gly Thr Val Asn
130 135 140
Gly Met Gln Pro Met Gln Ala Asp Leu Gln Ala Ala Ala Ala Ala Pro
145 150 155 160
Gly Leu Ala Asp Ser Arg Gly Ser Lys Gln Asp Ala Ala Val Ala Gly
165 170 175
Ala Ile Ser Glu Pro Ser Gly Thr Glu Ser His Lys Ser Thr Gly Ala
180 185 190
Asp His Glu Ala Gly Gly Asp Val Ala Glu Gln Ser
195 200
<210>39
<211>636
<212>DNA
<213>欧洲油菜
<400>39
atgcagcagc agcagcagca gcagcagcag cctccgcaaa tgtttccgat ggctccttcg 60
atgccgccaa ctaacatcac caccgaacag atccaaaagt accttgagga gaacaagaag 120
ctgataatgg caatcatgga aaatcagaat cttggcaagc ttgcagagtg tgcacagtac 180
caagctcttc tccagaagaa cttaatgtac ctcgctgcta ttgctgatgc tcaacctcct 240
ccatctaccg ctggagctac accaccacca gctatggctt cccagatggg ggcaccgcat 300
cctgggatgc aaccgccgag ctactttatg caacacccac aagcttcagg gatggctcaa 360
caagcaccac ccgctggtat cttccctccg agaggtcctt tgcagtttgg tagcccacac 420
cagcttcagg atccgcaaca gcagcatatg catcaacagg ctatgcaagg acacatgggg 480
atgcgaccaa tgggtatcaa caacaacaat gggatgcagc atcagatgca gcaacaacaa 540
ccagaaacct ctcttggagg aagcgctgca aacgtggggc ttagaggtgg aaagcaagat 600
ggagcagatg gacaaggaaa agatgatggc aaatga 636
<210>40
<211>203
<212>PRT
<213>欧洲油菜
<400>40
Met Gln Gln His Leu Met Gln Met Gln Pro Met Met Ala Gly Tyr Tyr
1 5 10 15
Pro Ser Asn Val Thr Ser Asp His Ile Gln Gln Tyr Leu Asp Glu Asn
20 25 30
Lys Ser Leu Ile Leu Lys Ile Val Glu Ser Gln Asn Ser Gly Lys Leu
35 40 45
Ser Glu Cys Ala Glu Asn Gln Ala Arg Leu Gln Arg Asn Leu Met Tyr
50 55 60
Leu Ala Ala Ile Ala Asp Ser Gln Pro Gln Pro Pro Ser Val His Ser
65 70 75 80
Gln Tyr Gly Ser Ala Gly Gly Gly Leu Ile Gln Gly Glu Gly Ala Ser
85 90 95
His Tyr Leu Gln Gln Gln Gln Ala Thr Gln Gln Gln Gln Met Thr Gln
100 105 110
Gln Ser Leu Met Ala Ala Arg Ser Ser Met Met Tyr Gln Gln Gln Gln
115 120 125
Gln Pro Tyr Ala Thr Leu Gln His Gln Gln Leu His His Ser Gln Leu
130 135 140
Gly Met Ser Ser Ser Ser Gly Gly Gly Ser Ser Gly Leu His Ile Leu
145 150 155 160
Gln Gly Glu Ala Gly Gly Phe His Glu Phe Gly Arg Gly Lys Pro Glu
165 170 175
Met Gly Ser Gly Glu Gly Arg Gly Gly Ser Ser Gly Asp Gly Gly Glu
180 185 190
Thr Leu Tyr Leu Lys Ser Ser Asp Asp Gly Asn
195 200
<210>41
<211>636
<212>DNA
<213>甜橙
<400>41
atgcagcagc caccgcaaat gatccctgtt atgccttcat ttccacccac caacatcacc 60
acagagcaga ttcaaaagta ccttgatgag aacaaaaagt tgattttggc aattttggac 120
aatcaaaatc ttggaaagct tacagaatgt gcccactatc aagctcagct tcaaaagaat 180
ttaatgtatt tagctgcaat tgctgatgca caaccacaag caccaacaat gcctcctcag 240
atggctccac atcctgcaat gcaagctagt gggtattaca tgcaacatcc tcaggcggca 300
gcaatggctc agcaacaagg aatctttccc caaaagatgc cattacaatt caataaccct 360
catcaactac aggatcctca acagcagcta caccaacatc aagccatgca agcacaaatg 420
ggaatgagac cgggtgccac taacaatggt atgcatccca tgcatgctga aagctctctt 480
ggaggtggca gcagtggagg acccccttca gcatcaggcc caggtgacat acgtggtgga 540
aataagcaag atgcctcgga ggctgggact actggtgctg atggccaggg cagttcggct 600
ggtgggcatg gtggggatgg agaggaggca aagtga 636
<210>42
<211>211
<212>PRT
<213>甜橙
<400>42
Met Gln Gln Pro Pro Gln Met Ile Pro Val Met Pro Ser Phe Pro Pro
1 5 10 15
Thr Asn Ile Thr Thr Glu Gln Ile Gln Lys Tyr Leu Asp Glu Asn Lys
20 25 30
Lys Leu Ile Leu Ala Ile Leu Asp Asn Gln Asn Leu Gly Lys Leu Thr
35 40 45
Glu Cys Ala His Tyr Gln Ala Gln Leu Gln Lys Asn Leu Met Tyr Leu
50 55 60
Ala Ala Ile Ala Asp Ala Gln Pro Gln Ala Pro Thr Met Pro Pro Gln
65 70 75 80
Met Ala Pro His Pro Ala Met Gln Ala Ser Gly Tyr Tyr Met Gln His
85 90 95
Pro Gln Ala Ala Ala Met Ala Gln Gln Gln Gly Ile Phe Pro Gln Lys
100 105 110
Met Pro Leu Gln Phe Asn Asn Pro His Gln Leu Gln Asp Pro Gln Gln
115 120 125
Gln Leu His Gln His Gln Ala Met Gln Ala Gln Met Gly Met Arg Pro
130 135 140
Gly Ala Thr Asn Asn Gly Met His Pro Met His Ala Glu Ser Ser Leu
145 150 155 160
Gly Gly Gly Ser Ser Gly Gly Pro Pro Ser Ala Ser Gly Pro Gly Asp
165 170 175
Ile Arg Gly Gly Asn Lys Gln Asp Ala Ser Glu Ala Gly Thr Thr Gly
180 185 190
Ala Asp Gly Gln Gly Ser Ser Ala Gly Gly His Gly Gly Asp Gly Glu
195 200 205
Glu Ala Lys
210
<210>43
<211>597
<212>DNA
<213>乳浆大戟(Euphorbia esula)
<400>43
atgcagcagc aaccgcagat gatgcctatg atgccttcat atccaccagc aaacattacc 60
acggagcaaa tccaaaagta tcttgatgaa aataaaaaat tgattttggc gatcttggat 120
aatcaaaatc ttggaaaact cgctgagtgt gcacagtatc aagccctgct gcaaaaaaat 180
ctgatgtatt tagccgcaat tgctgatgca caaccccaga ccccacccat gccacctcag 240
atgtccccac atccggctat gcaacaagga gcatattaca tgcaacatcc tcaggctgca 300
gcagcagcaa tggctcatca gtcgggtatt ttcccaccaa agatgtctcc gttacaattc 360
aataatcctc atcaaataca ggacccccag cagttacatc aagcagccct ccaagggcaa 420
atgggaatga ggcccatggg gcccaataac gggatgcatc cgatgcaccc cgaggcaaat 480
cttggaggat ctaatgatgg tcgtggagga aacaaacagg atgctccgga gacgggagca 540
tcgggaggtg atgggcaagg caattctggt ggtgatgggg ctgaagatgg gaaatga 597
<210>44
<211>198
<212>PRT
<213>乳浆大戟
<400>44
Met Gln Gln Gln Pro Gln Met Met Pro Met Met Pro Ser Tyr Pro Pro
1 5 10 15
Ala Asn Ile Thr Thr Glu Gln Ile Gln Lys Tyr Leu Asp Glu Asn Lys
20 25 30
Lys Leu Ile Leu Ala Ile Leu Asp Asn Gln Asn Leu Gly Lys Leu Ala
35 40 45
Glu Cys Ala Gln Tyr Gln Ala Leu Leu Gln Lys Asn Leu Met Tyr Leu
50 55 60
Ala Ala Ile Ala Asp Ala Gln Pro Gln Thr Pro Pro Met Pro Pro Gln
65 70 75 80
Met Ser Pro His Pro Ala Met Gln Gln Gly Ala Tyr Tyr Met Gln His
85 90 95
Pro Gln Ala Ala Ala Ala Ala Met Ala His Gln Ser Gly Ile Phe Pro
100 105 110
Pro Lys Met Ser Pro Leu Gln Phe Asn Asn Pro His Gln Ile Gln Asp
115 120 125
Pro Gln Gln Leu His Gln Ala Ala Leu Gln Gly Gln Met Gly Met Arg
130 135 140
Pro Met Gly Pro Asn Asn Gly Met His Pro Met His Pro Glu Ala Asn
145 150 155 160
Leu Gly Gly Ser Asn Asp Gly Arg Gly Gly Asn Lys Gln Asp Ala Pro
165 170 175
Glu Thr Gly Ala Ser Gly Gly Asp Gly Gln Gly Asn Ser Gly Gly Asp
180 185 190
Gly Ala Glu Asp Gly Lys
195
<210>45
<211>642
<212>DNA
<213>大豆(Glycine max)
<400>45
atgcagcaga caccgccaat gattcctatg atgccttctt tcccacctac gaacataacc 60
accgagcaga ttcaaaaata ccttgatgag aacaagaagc tgattctggc aatattggac 120
aatcaaaatc ttggaaaact tgcagaatgt gcccagtacc aagctcagct tcaaaagaat 180
ttgatgtatt tagctgcaat tgctgatgcc cagcctcaaa ccccggccat gcctccgcag 240
atggcaccgc accctgccat gcaaccagga ttctatatgc aacatcctca ggctgctgca 300
gcagcaatgg ctcagcagca gcaaggaatg ttcccccaga aaatgccatt gcaatttggc 360
aatccacatc aaatgcagga acaacaacag cagctacacc agcaggccat ccaaggtcaa 420
atgggactta gacctggaga tataaataat ggcatgcatc caatgcacag tgaggctgct 480
cttggaggtg gaaacagcgg tggtccacct tcggctactg gtccaaacga tgcacgtggt 540
ggaagcaagc aagatgcctc tgaggctgga acagctggtg gagacggcca aggcagctcc 600
gcggctgctc ataacagtgg agatggtgaa gaggcaaagt ga 642
<210>46
<211>213
<212>PRT
<213>大豆
<400>46
Met Gln Gln Thr Pro Pro Met Ile Pro Met Met Pro Ser Phe Pro Pro
1 5 10 15
Thr Asn Ile Thr Thr Glu Gln Ile Gln Lys Tyr Leu Asp Glu Asn Lys
20 25 30
Lys Leu Ile Leu Ala Ile Leu Asp Asn Gln Asn Leu Gly Lys Leu Ala
35 40 45
Glu Cys Ala Gln Tyr Gln Ala Gln Leu Gln Lys Asn Leu Met Tyr Leu
50 55 60
Ala Ala Ile Ala Asp Ala Gln Pro Gln Thr Pro Ala Met Pro Pro Gln
65 70 75 80
Met Ala Pro His Pro Ala Met Gln Pro Gly Phe Tyr Met Gln His Pro
85 90 95
Gln Ala Ala Ala Ala Ala Met Ala Gln Gln Gln Gln Gly Met Phe Pro
100 105 110
Gln Lys Met Pro Leu Gln Phe Gly Asn Pro His Gln Met Gln Glu Gln
115 120 125
Gln Gln Gln Leu His Gln Gln Ala Ile Gln Gly Gln Met Gly Leu Arg
130 135 140
Pro Gly Asp Ile Asn Asn Gly Met His Pro Met His Ser Glu Ala Ala
145 150 155 160
Leu Gly Gly Gly Asn Ser Gly Gly Pro Pro Ser Ala Thr Gly Pro Asn
165 170 175
Asp Ala Arg Gly Gly Ser Lys Gln Asp Ala Ser Glu Ala Gly Thr Ala
180 185 190
Gly Gly Asp Gly Gln Gly Ser Ser Ala Ala Ala His Asn Ser Gly Asp
195 200 205
Gly Glu Glu Ala Lys
210
<210>47
<211>633
<212>DNA
<213>野大豆(Glycine soya)
<400>47
atgcagcaga caccgcctat gattcctatg atgccttcgt tcccacctac gaacataacc 60
accgagcaga ttcaaaaata ccttgatgag aacaagaagc tgattctggc aatattggac 120
aatcaaaatc ttggaaaact tgcagaatgt gcccagtacc aagctcagct tcaaaagaat 180
ttgatgtatt tagctgcaat tgctgatgcc cagcctcaaa caccagccat gcctccacag 240
atggcaccac accctgccat gcaaccagga ttctatatgc aacatcctca ggctgcagca 300
gcagcaatgg ctcagcagca gcagcaagga atgttccccc agaaaatgcc attgcaattt 360
ggcaatccac atcaaatgca ggaacaacag cagcagctac accagcaagc catccaaggt 420
caaatgggac tgagacctgg aggaataaat aatggcatgc atccaatgca caatgagggc 480
ggcaacagcg gtggtccacc ctcggctacc ggtccgaacg acgcacgtgg tggaagcaag 540
caagatgctt ctgaggctgg aacagctggt ggagatggcc aaggcagctc tgcagctgct 600
cataacagtg gagatggtga agaggcaaag tga 633
<210>48
<211>210
<212>PRT
<213>野大豆
<400>48
Met Gln Gln Thr Pro Pro Met Ile Pro Met Met Pro Ser Phe Pro Pro
1 5 10 15
Thr Asn Ile Thr Thr Glu Gln Ile Gln Lys Tyr Leu Asp Glu Asn Lys
20 25 30
Lys Leu Ile Leu Ala Ile Leu Asp Asn Gln Asn Leu Gly Lys Leu Ala
35 40 45
Glu Cys Ala Gln Tyr Gln Ala Gln Leu Gln Lys Asn Leu Met Tyr Leu
50 55 60
Ala Ala Ile Ala Asp Ala Gln Pro Gln Thr Pro Ala Met Pro Pro Gln
65 70 75 80
Met Ala Pro His Pro Ala Met Gln Pro Gly Phe Tyr Met Gln His Pro
85 90 95
Gln Ala Ala Ala Ala Ala Met Ala Gln Gln Gln Gln Gln Gly Met Phe
100 105 110
Pro Gln Lys Met Pro Leu Gln Phe Gly Asn Pro His Gln Met Gln Glu
115 120 125
Gln Gln Gln Gln Leu His Gln Gln Ala Ile Gln Gly Gln Met Gly Leu
130 135 140
Arg Pro Gly Gly Ile Asn Asn Gly Met His Pro Met His Asn Glu Gly
145 150 155 160
Gly Asn Ser Gly Gly Pro Pro Ser Ala Thr Gly Pro Asn Asp Ala Arg
165 170 175
Gly Gly Ser Lys Gln Asp Ala Ser Glu Ala Gly Thr Ala Gly Gly Asp
180 185 190
Gly Gln Gly Ser Ser Ala Ala Ala His Asn Ser Gly Asp Gly Glu Glu
195 200 205
Ala Lys
210
<210>49
<211>690
<212>DNA
<213>陆地棉(Gossypium hirsutum)
<400>49
atgcagcagc acctgatgca gatgcagccc atgatggcag cttattatcc caacaacgtc 60
actactgatc atattcaaca gtatctcgat gagaacaagt cattgatctt aaagattgtt 120
gagagccaga attctgggaa attgagtgaa tgtgctgaga accaagcaag gctgcagcga 180
aacctcatgt acctggctgc cattgcggat tctcaacccc aaccacccac cgtgcatgca 240
cagtttccat ctggtggtat catgcagcca ggagctgggc actacatgca gcaccaacaa 300
gctcaacaaa tgacacaaca gtcgcttatg gctgctcggt cctcaatgtt gtattctcag 360
caaccatttt ctgcactgca acaacaacag cagcaagctt tgcacagtca gcttggcatg 420
agctctggcg gaagcacagg ccttcatatg ctgcaaactg aatctagtac tgcaggtggc 480
agtggagcac ttggggccgg agggtttcct gattttggac gtggttcttc tggagaaggc 540
atccatggtg gcaggccaat ggcaggtgga agcaagcaag atatcgggag tgccggctca 600
gctgaaggtc gtggaggaag ctctggtggt cagggtggtg gtgatggggg tgaaaccctt 660
tacttaaaag cagccgatga tgggaactga 690
<210>50
<211>229
<212>PRT
<213>陆地棉
<400>50
Met Gln Gln His Leu Met Gln Met Gln Pro Met Met Ala Ala Tyr Tyr
1 5 10 15
Pro Asn Asn Val Thr Thr Asp His Ile Gln Gln Tyr Leu Asp Glu Asn
20 25 30
Lys Ser Leu Ile Leu Lys Ile Val Glu Ser Gln Asn Ser Gly Lys Leu
35 40 45
Ser Glu Cys Ala Glu Asn Gln Ala Arg Leu Gln Arg Asn Leu Met Tyr
50 55 60
Leu Ala Ala Ile Ala Asp Ser Gln Pro Gln Pro Pro Thr Val His Ala
65 70 75 80
Gln Phe Pro Ser Gly Gly Ile Met Gln Pro Gly Ala Gly His Tyr Met
85 90 95
Gln His Gln Gln Ala Gln Gln Met Thr Gln Gln Ser Leu Met Ala Ala
100 105 110
Arg Ser Ser Met Leu Tyr Ser Gln Gln Pro Phe Ser Ala Leu Gln Gln
115 120 125
Gln Gln Gln Gln Ala Leu His Ser Gln Leu Gly Met Ser Ser Gly Gly
130 135 140
Ser Thr Gly Leu His Met Leu Gln Thr Glu Ser Ser Thr Ala Gly Gly
145 150 155 160
Ser Gly Ala Leu Gly Ala Gly Gly Phe Pro Asp Phe Gly Arg Gly Ser
165 170 175
Ser Gly Glu Gly Ile His Gly Gly Arg Pro Met Ala Gly Gly Ser Lys
180 185 190
Gln Asp Ile Gly Ser Ala Gly Ser Ala Glu Gly Arg Gly Gly Ser Ser
195 200 205
Gly Gly Gln Gly Gly Gly Asp Gly Gly Glu Thr Leu Tyr Leu Lys Ala
210 215 220
Ala Asp Asp Gly Asn
225
<210>51
<211>642
<212>DNA
<213>陆地棉
<400>51
atgccgcagc caccgcaaat gattcctgtg atgccttcat atccacctac taatatcact 60
actgaacaga ttcagaagta ccttgatgag aataagaagt tgattttggc aattttggac 120
aatcagaatc ttggaaaact cgctgaatgc gcccagtatc aagctcagct gcaaaagaat 180
ttgatgtatt tagctgcaat tgcggatgct caacctcaat caacgccagc aatgtcgcct 240
cagatggcac cgcatccagc aatgcaaccc ggaggatatt ttatgcaaca tcctcaagct 300
gctgcaatgt cacagcaacc tggcatgtac cctcaaaagg tgccattgca attcaatagt 360
ccgcatcaaa tgcaggaccc tcagcacctc ctatatcagc agcatcaaca agcaatgcaa 420
ggtcaaatgg gaatcaggcc tgggggaccc aataatagca tgcatcccat gcattcagag 480
gctagccttg gaggcggcag cagtggtggt ccccctcaac cttcaggccc aagtgatgga 540
cgtgctggaa acaagcaaga gggctccgaa gctggtggta atgggcaggg cagcacaact 600
ggtgggcatg gtggcggtga tggagcggat gaggcaaagt ga 642
<210>52
<211>213
<212>PRT
<213>陆地棉
<400>52
Met Pro Gln Pro Pro Gln Met Ile Pro Val Met Pro Ser Tyr Pro Pro
1 5 10 15
Thr Asn Ile Thr Thr Glu Gln Ile Gln Lys Tyr Leu Asp Glu Asn Lys
20 25 30
Lys Leu Ile Leu Ala Ile Leu Asp Asn Gln Asn Leu Gly Lys Leu Ala
35 40 45
Glu Cys Ala Gln Tyr Gln Ala Gln Leu Gln Lys Asn Leu Met Tyr Leu
50 55 60
Ala Ala Ile Ala Asp Ala Gln Pro Gln Ser Thr Pro Ala Met Ser Pro
65 70 75 80
Gln Met Ala Pro His Pro Ala Met Gln Pro Gly Gly Tyr Phe Met Gln
85 90 95
His Pro Gln Ala Ala Ala Met Ser Gln Gln Pro Gly Met Tyr Pro Gln
100 105 110
Lys Val Pro Leu Gln Phe Asn Ser Pro His Gln Met Gln Asp Pro Gln
115 120 125
His Leu Leu Tyr Gln Gln His Gln Gln Ala Met Gln Gly Gln Met Gly
130 135 140
Ile Arg Pro Gly Gly Pro Asn Asn Ser Met His Pro Met His Ser Glu
145 150 155 160
Ala Ser Leu Gly Gly Gly Ser Ser Gly Gly Pro Pro Gln Pro Ser Gly
165 170 175
Pro Ser Asp Gly Arg Ala Gly Asn Lys Gln Glu Gly Ser Glu Ala Gly
180 185 190
Gly Asn Gly Gln Gly Ser Thr Thr Gly Gly His Gly Gly Gly Asp Gly
195 200 205
Ala Asp Glu Ala Lys
210
<210>53
<211>561
<212>DNA
<213>大麦(Hordeum vulgare)
<400>53
atgcagcaag cgatgcccat gccgccggcg gcggcggcgc ctgggatgcc tccttctgcc 60
ggcctcagca ccgagcagat ccaaaagtac ctggatgaaa ataaacaact aattttggct 120
atcttggaaa atcagaacct gggaaagttg gcggaatgtg ctcagtatca agctcagctt 180
cagaagaatc ttttgtattt ggctgcgatt gctgatactc agccacagac ctctgtaagc 240
cgtcctcaga tggcaccacc tgctgcatcc ccaggggcag ggcattacat gtcacaggtg 300
ccaatgttcc ctccgaggac ccctctaacg cctcagcaga tgcaggagca gcaactacag 360
caacaacagg ctcagatgct tccgtttgct ggtcaaatgg ttgcgagacc cggggctgtc 420
aatggcattc cccaggcccc tcaagttgaa caaccagcct atgcagcagg tggggccagt 480
tccgagcctt ctggcaccga gagccacagg agcactggcg ccgataacga tggtgggagc 540
ggcttggctg accagtccta a 561
<210>54
<211>186
<212>PRT
<213>大麦
<400>54
Met Gln Gln Ala Met Pro Met Pro Pro Ala Ala Ala Ala Pro Gly Met
1 5 10 15
Pro Pro Ser Ala Gly Leu Ser Thr Glu Gln Ile Gln Lys Tyr Leu Asp
20 25 30
Glu Asn Lys Gln Leu Ile Leu Ala Ile Leu Glu Asn Gln Asn Leu Gly
35 40 45
Lys Leu Ala Glu Cys Ala Gln Tyr Gln Ala Gln Leu Gln Lys Asn Leu
50 55 60
Leu Tyr Leu Ala Ala Ile Ala Asp Thr Gln Pro Gln Thr Ser Val Ser
65 70 75 80
Arg Pro Gln Met Ala Pro Pro Ala Ala Ser Pro Gly Ala Gly His Tyr
85 90 95
Met Ser Gln Val Pro Met Phe Pro Pro Arg Thr Pro Leu Thr Pro Gln
100 105 110
Gln Met Gln Glu Gln Gln Leu Gln Gln Gln Gln Ala Gln Met Leu Pro
115 120 125
Phe Ala Gly Gln Met Val Ala Arg Pro Gly Ala Val Asn Gly Ile Pro
130 135 140
Gln Ala Pro Gln Val Glu Gln Pro Ala Tyr Ala Ala Gly Gly Ala Ser
145 150 155 160
Ser Glu Pro Ser Gly Thr Glu Ser His Arg Ser Thr Gly Ala Asp Asn
165 170 175
Asp Gly Gly Ser Gly Leu Ala Asp Gln Ser
180 185
<210>55
<211>555
<212>DNA
<213>野莴苣(Lactuca serriola)
<220>
<221>misc_feature
<222>(253)..(253)
<223>n为a、c、g或t
<400>55
atgaagcagc cgatgatgcc gaatccaatg atgtcttctt cgtttcctcc tacaaacatc 60
accaccgatc agatccaaaa gttccttgat gaaaacaagc aactaattat agcaataatg 120
agcaacctaa atcttggaaa gcttgctgaa tgtgcccagt accaagctct actccaaaaa 180
aatttgatgt atctagcagc cattgcagat gctcaaccac ctacacctac accaacacta 240
aatatctctt atnagatggg cccggttcca catccaggga tgccacagca aggtggattt 300
tacatggcgc agcagcaccc tcaggcggct gtaatgacgg ctcagccacc ttctggtttt 360
ccacaaccga tgcctggtat gcaatttaac agcccacagg ctattcaagg gcagatgggc 420
gggaggtccg gtgggccgcc aagctcagcc gctagtgatg tctggagagg aagcatgcaa 480
gatggtggtg gtggtgctgc tgctgatggt ggtaaggatg gtcatgctgg cggtggacct 540
gaggaagcaa agtaa 555
<210>56
<211>184
<212>PRT
<213>野莴苣
<220>
<221>misc_feature
<222>(85)..(85)
<223>Xaa可以是任何天然存在的氨基酸
<400>56
Met Lys Gln Pro Met Met Pro Asn Pro Met Met Ser Ser Ser Phe Pro
1 5 10 15
Pro Thr Asn Ile Thr Thr Asp Gln Ile Gln Lys Phe Leu Asp Glu Asn
20 25 30
Lys Gln Leu Ile Ile Ala Ile Met Ser Asn Leu Asn Leu Gly Lys Leu
35 40 45
Ala Glu Cys Ala Gln Tyr Gln Ala Leu Leu Gln Lys Asn Leu Met Tyr
50 55 60
LeuAla Ala Ile Ala Asp Ala Gln Pro Pro Thr Pro Thr Pro Thr Leu
65 70 75 80
Asn Ile Ser Tyr Xaa Met Gly Pro Val Pro His Pro Gly Met Pro Gln
85 90 95
Gln Gly Gly Phe Tyr Met Ala Gln Gln His Pro Gln Ala Ala Val Met
100 105 110
Thr Ala Gln Pro Pro Ser Gly Phe Pro Gln Pro Met Pro Gly Met Gln
115 120 125
Phe Asn Ser Pro Gln Ala Ile Gln Gly Gln Met Gly Gly Arg Ser Gly
130 135 140
Gly Pro Pro Ser Ser Ala Ala Ser Asp Val Trp Arg Gly Ser Met Gln
145 150 155 160
Asp Gly Gly Gly Gly Ala Ala Ala Asp Gly Gly Lys Asp Gly His Ala
165 170 175
Gly Gly Gly Pro Glu Glu Ala Lys
180
<210>57
<211>627
<212>DNA
<213>番茄(Lycopersicon esculentum)
<400>57
atgcagcagc acctgatgca gatgcagccc atgatggcag cttactatcc aacgaacgtc 60
actactgacc atattcaaca gtatttggat gaaaacaaat cactcattct gaagattgtt 120
gagagccaga actctgggaa actcagtgaa tgtgcggaga accaagctag gcttcagagg 180
aatctgatgt accttgctgc gattgctgat tcacaacctc aaccttctag catgcattct 240
cagttctctt ctggtgggat gatgcagcca gggacacaca gttacttgca gcagcagcag 300
cagcaacaac aagcgcaaca aatggcaaca caacaactca tggctgcaag atcctcgtcg 360
atgctctatg gacaacagca gcagcaatct cagttatcgc aatatcaaca aggcttgcat 420
agtagccaac tcggcatgag ttctggcagt ggcggaagca ctggacttca tcacatgctt 480
caaagtgaat catcacctca tggtggtggt ttctctcatg acttcggccg cgcaaataag 540
caagacattg ggagtagtat gtctgctgaa gggcgcggcg gaagttcagg tggtgagaat 600
ctttatctga aagcttctga ggattga 627
<210>58
<211>208
<212>PRT
<213>番茄
<400>58
Met Gln Gln His Leu Met Gln Met Gln Pro Met Met Ala Ala Tyr Tyr
1 5 10 15
Pro Thr Asn Val Thr Thr Asp His Ile Gln Gln Tyr Leu Asp Glu Asn
20 25 30
Lys Ser Leu Ile Leu Lys Ile Val Glu Ser Gln Asn Ser Gly Lys Leu
35 40 45
Ser Glu Cys Ala Glu Asn Gln Ala Arg Leu Gln Arg Asn Leu Met Tyr
50 55 60
Leu Ala Ala Ile Ala Asp Ser Gln Pro Gln Pro Ser Ser Met His Ser
65 70 75 80
Gln Phe Ser Ser Gly Gly Met Met Gln Pro Gly Thr His Ser Tyr Leu
85 90 95
Gln Gln Gln Gln Gln Gln Gln Gln Ala Gln Gln Met Ala Thr Gln Gln
100 105 110
Leu Met Ala Ala Arg Ser Ser Ser Met Leu Tyr Gly Gln Gln Gln Gln
115 120 125
Gln Ser Gln Leu Ser Gln Tyr Gln Gln Gly Leu His Ser Ser Gln Leu
130 135 140
Gly Met Ser Ser Gly Ser Gly Gly Ser Thr Gly Leu His His Met Leu
145 150 155 160
Gln Ser Glu Ser Ser Pro His Gly Gly Gly Phe Ser His Asp Phe Gly
165 170 175
Arg Ala Asn Lys Gln Asp Ile Gly Ser Ser Met Ser Ala Glu Gly Arg
180 185 190
Gly Gly Ser Ser Gly Gly Glu Asn Leu Tyr Leu Lys Ala Ser Glu Asp
195 200 205
<210>59
<211>624
<212>DNA
<213>驯化苹果(Malus domestica)
<400>59
atgcagcagc caccacaaat gatccccgtc atgccttcat ttcctcccac caacatcacc 60
accgaacaaa ttcagaagta ccttgatgac aacaaaaagt tgattctggc aatattggat 120
aatcaaaatc ttggaaaact tgctgagtgt gctcagtacc aggctctgct tcaaaagaat 180
ctgatgtatt tagcagcaat tgccgatgcg caaccacagg caccagctgc ccctccccag 240
atggccccac atcctgctat gcaacaggca ggatattaca tgcaacatcc tcaggcagca 300
gcaatggctc agcaacaggg tattttctcc ccaaagatgc cgatgcaatt caataacatg 360
catcaaatgc acgatccaca gcagcaccaa caagccatgc aagggcaaat gggaatgaga 420
cctggagggc ctaacggcat gccttccatg cttcatactg aggccacaca tggtggtggt 480
agtggcggcc caaattcagc tggagaccca aatgatgggc gtggaggaag caagcaagac 540
gcctctgagt ctggggcagg tggtgatggc caggggacct cagccggcgg gcgtggaact 600
ggtgatggag aggacggcaa gtga 624
<210>60
<211>207
<212>PRT
<213>驯化苹果
<400>60
Met Gln Gln Pro Pro Gln Met Ile Pro Val Met Pro Ser Phe Pro Pro
1 5 10 15
Thr Asn Ile Thr Thr Glu Gln Ile Gln Lys Tyr Leu Asp Asp Asn Lys
20 25 30
Lys Leu Ile Leu Ala Ile Leu Asp Asn Gln Asn Leu Gly Lys Leu Ala
35 40 45
Glu Cys Ala Gln Tyr Gln Ala Leu Leu Gln Lys Asn Leu Met Tyr Leu
50 55 60
Ala Ala Ile Ala Asp Ala Gln Pro Gln Ala Pro Ala Ala Pro Pro Gln
65 70 75 80
Met Ala Pro His Pro Ala Met Gln Gln Ala Gly Tyr Tyr Met Gln His
85 90 95
Pro Gln Ala Ala Ala Met Ala Gln Gln Gln Gly Ile Phe Ser Pro Lys
100 105 110
Met Pro Met Gln Phe Asn Asn Met His Gln Met His Asp Pro Gln Gln
115 120 125
His Gln Gln Ala Met Gln Gly Gln Met Gly Met Arg Pro Gly Gly Pro
130 135 140
Asn Gly Met Pro Ser Met Leu His Thr Glu Ala Thr His Gly Gly Gly
145 150 155 160
Ser Gly Gly Pro Asn Ser Ala Gly Asp Pro Asn Asp Gly Arg Gly Gly
165 170 175
Ser Lys Gln Asp Ala Ser Glu Ser Gly Ala Gly Gly Asp Gly Gln Gly
180 185 190
Thr Ser Ala Gly Gly Arg Gly Thr Gly Asp Gly Glu Asp Gly Lys
195 200 205
<210>61
<211>639
<212>DNA
<213>蒺藜状苜蓿
<400>61
atgcagcaga cacctcaaat gattcctatg atgccttcat tcccacaaca aacaaacata 60
accactgagc agattcaaaa atatcttgat gagaacaaga agctgatcct ggcaatattg 120
gacaatcaaa atcttggaaa acttgcagaa tgtgcccagt accaagctca gcttcagaag 180
aatttgatgt atttagctgc aattgctgac gcgcagccac aaacaccggc cttgcctcca 240
cagatggccc cgcaccctgc gatgcaacaa ggattctata tgcaacatcc tcaggctgca 300
gcaatggctc agcaacaagg aatgttcccc caaaaaatgc caatgcagtt cggtaatccg 360
catcaaatgc aggatcagca gcatcagcag caacaacagc agctacatca gcaagctatg 420
caaggtcaaa tgggacttag acctggaggg ataaataacg gcatgcatcc aatgcacaac 480
gaggctgctc tcggaggtag cggcagtggt ggtcaaatga cgggcgtggt ggtggagcaa 540
gcaagatgct tcggagctgg gacagccggc ggtgatggtc aaggaacctc tgccgcagct 600
gcgcacaaca gtggagatgc ttcagaagaa ggaaagtaa 639
<210>62
<211>213
<212>PRT
<213>蒺藜状苜蓿
<400>62
Met Gln Gln Thr Pro Gln Met Ile Pro Met Met Pro Ser Phe Pro Gln
1 5 10 15
Gln Thr Asn Ile Thr Thr Glu Gln Ile Gln Lys Tyr Leu Asp Glu Asn
20 25 30
Lys Lys Leu Ile Leu Ala Ile Leu Asp Asn Gln Asn Leu Gly Lys Leu
35 40 45
Ala Glu Cys Ala Gln Tyr Gln Ala Gln Leu Gln Lys Asn Leu Met Tyr
50 55 60
Leu Ala Ala Ile Ala Asp Ala Gln Pro Gln Thr Pro Ala Leu Pro Pro
65 70 75 80
Gln Met Ala Pro His Pro Ala Met Gln Gln Gly Phe Tyr Met Gln His
85 90 95
Pro Gln Ala Ala Ala Met Ala Gln Gln Gln Gly Met Phe Pro Gln Lys
100 105 110
Met Pro Met Gln Phe Gly Asn Pro His Gln Met Gln Asp Gln Gln His
115 120 125
Gln Gln Gln Gln Gln Gln Leu His Gln Gln Ala Met Gln Gly Gln Met
130 135 140
Gly Leu Arg Pro Gly Gly Ile Asn Asn Gly Met His Pro Met His Asn
145 150 155 160
Glu Ala Ala Leu Gly Gly Ser Gly Ser Gly Gly Pro Asn Asp Gly Arg
165 170 175
Gly Gly Gly Ser Lys Gln Asp Ala Ser Glu Ala Gly Thr Ala Gly Gly
180 185 190
Asp Gly Gln Gly Thr Ser Ala Ala Ala Ala His Asn Ser Gly Asp Ala
195 200 205
Ser Glu Glu Gly Lys
210
<210>63
<211>624
<212>DNA
<213>柳枝稷(Panicum virgatum)
<400>63
atgcagcagc agatgcccat gcagtcggcg cccccggcga ccggcatcac caccgagcag 60
atccaaaagt atttggatga aaataagcag cttattttgg ccatcctgga aaatcagaac 120
ttaggaaagt tggctgaatg tgctcagtat caagctcagc ttcaaaagaa tctcttgtac 180
ctggctgcga ttgcagatgc ccaaccccaa ccaccacaga accctgcaag tcgcccacag 240
atgatgcaac ctggcatggt accaggtgca gggcattaca tgtcccaagt accaatgttc 300
ccgccaagaa caccattaac cccgcaacag atgcaagaac agcagcagca gcagcagcag 360
cttcaacagc agcaagcaca ggctcttgct ttcccgggac agatggtcat gagacctacc 420
attaatggca tgcagcctat gcaagccgac cctgctgccg ccgccgccag cctacagcag 480
tcagcacctg gccctactga tgggcgagga ggcaagcaag atgcaactgc tggggtgagc 540
acagagcctt ctggcaccga gagccacaag agcacaaccg cagcagatca cgatgtgggc 600
actgatgtcg cggagaaatc ctaa 624
<210>64
<211>207
<212>PRT
<213>柳枝稷
<400>64
Met Gln Gln Gln Met Pro Met Gln Ser Ala Pro Pro Ala Thr Gly Ile
1 5 10 15
Thr Thr Glu Gln Ile Gln Lys Tyr Leu Asp Glu Asn Lys Gln Leu Ile
20 25 30
Leu Ala Ile Leu Glu Asn Gln Asn Leu Gly Lys Leu Ala Glu Cys Ala
35 40 45
Gln Tyr Gln Ala Gln Leu Gln Lys Asn Leu Leu Tyr Leu Ala Ala Ile
50 55 60
Ala Asp Ala Gln Pro Gln Pro Pro Gln Asn Pro Ala Ser Arg Pro Gln
65 70 75 80
Met Met Gln Pro Gly Met Val Pro Gly Ala Gly His Tyr Met Ser Gln
85 90 95
Val Pro Met Phe Pro Pro Arg Thr Pro Leu Thr Pro Gln Gln Met Gln
100 105 110
Glu Gln Gln Gln Gln Gln Gln Gln Leu Gln Gln Gln Gln Ala Gln Ala
115 120 125
Leu Ala Phe Pro Gly Gln Met Val Met Arg Pro Thr Ile Asn Gly Met
130 135 140
Gln Pro Met Gln Ala Asp Pro Ala Ala Ala Ala Ala Ser Leu Gln Gln
145 150 155 160
Ser Ala Pro Gly Pro Thr Asp Gly Arg Gly Gly Lys Gln Asp Ala Thr
165 170 175
Ala Gly Val Ser Thr Glu Pro Ser Gly Thr Glu Ser His Lys Ser Thr
180 185 190
Thr Ala Ala Asp His Asp Val Gly Thr Asp Val Ala Glu Lys Ser
195 200 205
<210>65
<211>747
<212>DNA
<213>北美云杉(Picea sitchensis)
<400>65
atgcagcagc atctcatgca aatgcagccc atgatggcgg catacgcctc caacaacatc 60
accactgatc acatccagaa gtacctggat gagaacaagc agttgattct ggcaattctg 120
gacaaccaaa atcttggaaa gctcaatgag tgtgctcagt accaagcaaa acttcagcag 180
aatttgatgt atctggctgc gattgctgat tctcaaccac aagcacaaac tgcacatgct 240
cagattcctc ctaatgcagt gatgcagtct ggtgggcatt acatgcagca ccagcaggca 300
cagcaacaag tgactcctca gtctctgatg gcagctagat cttccatgct gtattctcag 360
cagccgatgg ctgctttgca tcaagctcag caacaacagc agcagcagca tcagcagcaa 420
caacaatctc ttcacagcca gcttggcata aattctggag gaagcagtgg attgcatatg 480
ttgcatggtg agacaaacat gggatgtaat gggcctctct catctggggg cttccctgaa 540
tttgggcgtg ggtctgctac ctctgctgaa ggtatgcagg ccaacagggg cttcactata 600
gatcgtggtt caaataagca ggatggagta ggatcagaga atgcccatcc aggtgctggt 660
gatggaagag ggagttcaac tggagggcag aatgcagatg agtcagaacc atcatacctg 720
aaagcctccg aagaagaagg aaactag 747
<210>66
<211>248
<212>PRT
<213>北美云杉
<400>66
Met Gln Gln His Leu Met Gln Met Gln Pro Met Met Ala Ala Tyr Ala
1 5 10 15
Ser Asn Asn Ile Thr Thr Asp His Ile Gln Lys Tyr Leu Asp Glu Asn
20 25 30
Lys Gln Leu Ile Leu Ala Ile Leu Asp Asn Gln Asn Leu Gly Lys Leu
35 40 45
Asn Glu Cys Ala Gln Tyr Gln Ala Lys Leu Gln Gln Asn Leu Met Tyr
50 55 60
Leu Ala Ala Ile Ala Asp Ser Gln Pro Gln Ala Gln Thr Ala His Ala
65 70 75 80
Gln Ile Pro Pro Asn Ala Val Met Gln Ser Gly Gly His Tyr Met Gln
85 90 95
His Gln Gln Ala Gln Gln Gln Val Thr Pro Gln Ser Leu Met Ala Ala
100 105 110
Arg Ser Ser Met Leu Tyr Ser Gln Gln Pro Met Ala Ala Leu His Gln
115 120 125
Ala Gln Gln Gln Gln Gln Gln Gln His Gln Gln Gln Gln Gln Ser Leu
130 135 140
His Ser Gln Leu Gly Ile Asn Ser Gly Gly Ser Ser Gly Leu His Met
145 150 155 160
Leu His Gly Glu Thr Asn Met Gly Cys Asn Gly Pro Leu Ser Ser Gly
165 170 175
Gly Phe Pro Glu Phe Gly Arg Gly Ser Ala Thr Ser Ala Glu Gly Met
180 185 190
Gln Ala Asn Arg Gly Phe Thr Ile Asp Arg Gly Ser Asn Lys Gln Asp
195 200 205
Gly Val Gly Ser Glu Asn Ala His Pro Gly Ala Gly Asp Gly Arg Gly
210 215 220
Ser Ser Thr Gly Gly Gln Asn Ala Asp Glu Ser Glu Pro Ser Tyr Leu
225 230 235 240
Lys Ala Ser Glu Glu Glu Gly Asn
245
<210>67
<211>735
<212>DNA
<213>火炬松(Pinus taeda)
<400>67
atgcagcagc acctcatgca aatgcagccc atgatggcgg cctacgcctc caacaatatc 60
accactgatc acatccagaa gtacctggat gagaacaagc agttgattct ggcaattttg 120
gacaaccaaa atctcggaaa gctcaatgag tgtgctcaat accaagcaaa acttcagcag 180
aatttgatgt atctggctgc tattgctgat tctcaacctc aagcacaaac tgcacatgct 240
cagattcctc caaatgcggt gatgcagtct ggtgggcatt acatgcagca tcaacaggca 300
cagcaacaag ttactcctca gtctctgatg gcagctagat cttccatact gtatgctcag 360
caacaacagc agcagcagca tcagcagcat cagcagcaac agcagcaaca acagtctctt 420
cacagccagc ttggcataaa ttctggagga agcagcggtt tgcatatgtt gcatggtgag 480
acaaacatgg gatgtaatgg gcctctgtca tctgggggat tccctgaatt tgggcgtggg 540
tctgctacct ctgctgatgg tatgcaggtg aacaggggct ttgctataga tcgtggttca 600
aacaagcagg atggagttgg atcagagaat gcccatgctg gtgctggtga tggaagaggg 660
agttcaactg gagggcagaa tgcagatgag tcagaaccat catacctgaa ggcctccgag 720
gaagaaggaa actag 735
<210>68
<211>244
<212>PRT
<213>火炬松
<400>68
Met Gln Gln His Leu Met Gln Met Gln Pro Met Met Ala Ala Tyr Ala
1 5 10 15
Ser Asn Asn Ile Thr Thr Asp His Ile Gln Lys Tyr Leu Asp Glu Asn
20 25 30
Lys Gln Leu Ile Leu Ala Ile Leu Asp Asn Gln Asn Leu Gly Lys Leu
35 40 45
Asn Glu Cys Ala Gln Tyr Gln Ala Lys Leu Gln Gln Asn Leu Met Tyr
50 55 60
Leu Ala Ala Ile Ala Asp Ser Gln Pro Gln Ala Gln Thr Ala His Ala
65 70 75 80
Gln Ile Pro Pro Asn Ala Val Met Gln Ser Gly Gly His Tyr Met Gln
85 90 95
His Gln Gln Ala Gln Gln Gln Val Thr Pro Gln Ser Leu Met Ala Ala
100 105 110
Arg Ser Ser Ile Leu Tyr Ala Gln Gln Gln Gln Gln Gln Gln His Gln
115 120 125
Gln His Gln Gln Gln Gln Gln Gln Gln Gln Ser Leu His Ser Gln Leu
130 135 140
Gly Ile Asn Ser Gly Gly Ser Ser Gly Leu His Met Leu His Gly Glu
145 150 155 160
Thr Asn Met Gly Cys Asn Gly Pro Leu Ser Ser Gly Gly Phe Pro Glu
165 170 175
Phe Gly Arg Gly Ser Ala Thr Ser Ala Asp Gly Met Gln Val Asn Arg
180 185 190
Gly Phe Ala Ile Asp Arg Gly Ser Asn Lys Gln Asp Gly Val Gly Ser
195 200 205
Glu Asn Ala His Ala Gly Ala Gly Asp Gly Arg Gly Ser Ser Thr Gly
210 215 220
Gly Gln Asn Ala Asp Glu Ser Glu Pro Ser Tyr Leu Lys Ala Ser Glu
225 230 235 240
Glu Glu Gly Asn
<210>69
<211>663
<212>DNA
<213>欧洲山杨(Populus tremula)
<400>69
atgcaacagc acctgatgca gatgcagccc atgatggcag cctattaccc cagcaacgtc 60
actactgatc atattcaaca gtatctggac gaaaacaagt cattgatttt gaagattgtt 120
gagagccaga attcagggaa actcagtgag tgtgcagaga accaagcaag actgcaacaa 180
aatctcatgt acttggctgc aattgctgat tgtcagcccc aaccacctac catgcatgcc 240
cagttccctt ccagcggcat tatgcagcca ggagcacatt acatgcagca tcaacaagct 300
caacagatga caccacaagc ccttatggct gcacgctctt ctatgctgca gtatgctcaa 360
cagccattct cagcgcttca acaacagcaa gccttacaca gccagctcgg catgagctct 420
ggtggaagcg caggacttca tatgatgcaa agcgaggcta acactgcagg aggcagtgga 480
gctcttggtg ctggacgatt tcctgatttt ggcatggatg cctccagtag aggaatcgca 540
agtgggagca agcaagatat tcggagtgca gggtctagtg aagggcgagg aggaagctct 600
ggaggccagg gtggtgatgg aggtgaaacc ctttacttga aatctgctga tgatgggaac 660
tga 663
<210>70
<211>220
<212>PRT
<213>欧洲山杨
<400>70
Met Gln Gln His Leu Met Gln Met Gln Pro Met Met Ala Ala Tyr Tyr
1 5 10 15
Pro Ser Asn Val Thr Thr Asp His Ile Gln Gln Tyr Leu Asp Glu Asn
20 25 30
Lys Ser Leu Ile Leu Lys Ile Val Glu Ser Gln Asn Ser Gly Lys Leu
35 40 45
Ser Glu Cys Ala Glu Asn Gln Ala Arg Leu Gln Gln Asn Leu Met Tyr
50 55 60
Leu Ala Ala Ile Ala Asp Cys Gln Pro Gln Pro Pro Thr Met His Ala
65 70 75 80
Gln Phe Pro Ser Ser Gly Ile Met Gln Pro Gly Ala His Tyr Met Gln
85 90 95
His Gln Gln Ala Gln Gln Met Thr Pro Gln Ala Leu Met Ala Ala Arg
100 105 110
Ser Ser Met Leu Gln Tyr Ala Gln Gln Pro Phe Ser Ala Leu Gln Gln
115 120 125
Gln Gln Ala Leu His Ser Gln Leu Gly Met Ser Ser Gly Gly Ser Ala
130 135 140
Gly Leu His Met Met Gln Ser Glu Ala Asn Thr Ala Gly Gly Ser Gly
145 150 155 160
Ala Leu Gly Ala Gly Arg Phe Pro Asp Phe Gly Met Asp Ala Ser Ser
165 170 175
Arg Gly Ile Ala Ser Gly Ser Lys Gln Asp Ile Arg Ser Ala Gly Ser
180 185 190
Ser Glu Gly Arg Gly Gly Ser Ser Gly Gly Gln Gly Gly Asp Gly Gly
195 200 205
Glu Thr Leu Tyr Leu Lys Ser Ala Asp Asp Gly Asn
210 215 220
<210>71
<211>678
<212>DNA
<213>甘蔗(Saccharum officinarum)
<400>71
atgcagcagc aacacctgat gcagatgaac cagaacatga ttgggggcta cacctctcct 60
gccgctgtga caaccgatct catccagcag tacctggatg agaacaagca gctgatcctg 120
gccatcctcg acaaccagaa caatggcaag gtggaggagt gcgaacggca ccaagctaag 180
ctccagcaca acctcatgta cctggccgcc atcgccgaca gccagccacc acagactgca 240
ccactatcac aatacccgtc caacctgatg atgcagccgg gccctcggta catgccaccg 300
cagtccgggc agatgatgag cccgcagtcg ctaatggcgg cgcggtcctc catgatgtac 360
gcgcacccgt ccatgtcacc actccagcag cagcaggcag cgcacgggca gctgggcatg 420
gcttcagggg gcggcggtgg cacgaccagt gggttcaaca tcctccatgg cgaggccagt 480
atgggcggtg ctggtggcgc ttgtgccggc aacaacatga tgaacgccgg catgttctca 540
ggctttggcc gcagcggcag tggcgccaag gagggatcga cctcgctgtc ggttgacgtc 600
cgtggtggca ccagctccgg cgcgcaaagc ggggacggcg agtacctgaa agcaggcacc 660
gaggaagaag gcagttaa 678
<210>72
<211>225
<212>PRT
<213>甘蔗
<400>72
Met Gln Gln Gln His Leu Met Gln Met Asn Gln Asn Met Ile Gly Gly
1 5 10 15
Tyr Thr Ser Pro Ala Ala Val Thr Thr Asp Leu Ile Gln Gln Tyr Leu
20 25 30
Asp Glu Asn Lys Gln Leu Ile Leu Ala Ile Leu Asp Asn Gln Asn Asn
35 40 45
Gly Lys Val Glu Glu Cys Glu Arg His Gln Ala Lys Leu Gln His Asn
50 55 60
Leu Met Tyr Leu Ala Ala Ile Ala Asp Ser Gln Pro Pro Gln Thr Ala
65 70 75 80
Pro Leu Ser Gln Tyr Pro Ser Asn Leu Met Met Gln Pro Gly Pro Arg
85 90 95
Tyr Met Pro Pro Gln Ser Gly Gln Met Met Ser Pro Gln Ser Leu Met
100 105 110
Ala Ala Arg Ser Ser Met Met Tyr Ala His Pro Ser Met Ser Pro Leu
115 120 125
Gln Gln Gln Gln Ala Ala His Gly Gln Leu Gly Met Ala Ser Gly Gly
130 135 140
Gly Gly Gly Thr Thr Ser Gly Phe Asn Ile Leu His Gly Glu Ala Ser
145 150 155 160
Met Gly Gly Ala Gly Gly Ala Cys Ala Gly Asn Asn Met Met Asn Ala
165 170 175
Gly Met Phe Ser Gly Phe Gly Arg Ser Gly Ser Gly Ala Lys Glu Gly
180 185 190
Ser Thr Ser Leu Ser Val Asp Val Arg Gly Gly Thr Ser Ser Gly Ala
195 200 205
Gln Ser Gly Asp Gly Glu Tyr Leu Lys Ala Gly Thr Glu Glu Glu Gly
210 215 220
Ser
225
<210>73
<211>561
<212>DNA
<213>甘蔗
<400>73
atgcagcagc cgatgcccat gcagccgcag gcgccggaga tgaccccggc cgccggaatc 60
accacggagc agatccaaaa gtatctggat gagaataagc agcttatttt ggctattttg 120
gaaaatcaga acctaggaaa attggcagaa tgtgctcagt atcaatcaca acttcagaag 180
aacctcttgt atctcgctgc aatcgcagat gcccaaccac agactgctgt aagccgccct 240
cagatggcgc cgcctggtgc attgcctgga gtagggcagt acatgtcaca ggtgcctatg 300
ttcccaccga ggacacctct aacaccccag cagatgcagg agcagcaact tcagcagcag 360
caggctcagc tgctaaattt cagtggccta atggttgcta gacctggcat ggtcaacggc 420
atgcctcagt ccattcaagt tcagcaagct cagccaccac cagcagggaa caaacaggat 480
gctggtgggg tcgcctcgga gccctcgggc attgagaacc acaggagcac tggtggtgat 540
aatgatggtg gaagcgacta g 561
<210>74
<211>186
<212>PRT
<213>甘蔗
<400>74
Met Gln Gln Pro Met Pro Met Gln Pro Gln Ala Pro Glu Met Thr Pro
1 5 10 15
Ala Ala Gly Ile Thr Thr Glu Gln Ile Gln Lys Tyr Leu Asp Glu Asn
20 25 30
Lys Gln Leu Ile Leu Ala Ile Leu Glu Asn Gln Asn Leu Gly Lys Leu
35 40 45
Ala Glu Cys Ala Gln Tyr Gln Ser Gln Leu Gln Lys Asn Leu Leu Tyr
50 55 60
Leu Ala Ala Ile Ala Asp Ala Gln Pro Gln Thr Ala Val Ser Arg Pro
65 70 75 80
Gln Met Ala Pro Pro Gly Ala Leu Pro Gly Val Gly Gln Tyr Met Ser
85 90 95
Gln Val Pro Met Phe Pro Pro Arg Thr Pro Leu Thr Pro Gln Gln Met
100 105 110
Gln Glu Gln Gln Leu Gln Gln Gln Gln Ala Gln Leu Leu Asn Phe Ser
115 120 125
Gly Leu Met Val Ala Arg Pro Gly Met Val Asn Gly Met Pro Gln Ser
130 135 140
Ile Gln Val Gln Gln Ala Gln Pro Pro Pro Ala Gly Asn Lys Gln Asp
145 150 155 160
Ala Gly Gly Val Ala Ser Glu Pro Ser Gly Ile Glu Asn His Arg Ser
165 170 175
Thr Gly Gly Asp Asn Asp Gly Gly Ser Asp
180 185
<210>75
<211>642
<212>DNA
<213>甘蔗
<400>75
atgcagcagc agatgcccat gccgccggcg cccgctgcgg cggcggcgcc cccggcggcc 60
ggcatcacca ccgagcagat ccaaaagtat ttggacgaaa ataagcaact tattttggcc 120
atcctggaaa atcagaactt aggaaagttg gctgaatgtg ctcagtatca agctcaactt 180
caaaagaacc tcttgtacct ggctgcgatt gctgatgccc aaccccagcc accacaaaac 240
cctgcaggtc gccctcagat gatgcaacct ggtatagtgc caggtgcggg gcattacatg 300
tcacaagtac caatgttccc tccaagaact ccattaaccc cacagcagat gcaagagcag 360
cagcagcaac agcttcagca gcagcaagcg caggctctta cattccctgg acagatggtc 420
atgagaccag ctaccatcaa cggcatacag cagcctatgc aagctgaccc tgcccgggca 480
gcggagctgc aacaaccacc acctatccca gctgacgggc gagtaagcaa gcagcaggac 540
acaacggctg gcgtgagctc agagccttct gccaatgaga gccacaagac cacaactgga 600
gcagatagtg aggcaggtgg tgacgtggcg gagaaatcct aa 642
<210>76
<211>213
<212>PRT
<213>甘蔗
<400>76
Met Gln Gln Gln Met Pro Met Pro Pro Ala Pro Ala Ala Ala Ala Ala
1 5 10 15
Pro Pro Ala Ala Gly Ile Thr Thr Glu Gln Ile Gln Lys Tyr Leu Asp
20 25 30
Glu Asn Lys Gln Leu Ile Leu Ala Ile Leu Glu Asn Gln Asn Leu Gly
35 40 45
Lys Leu Ala Glu Cys Ala Gln Tyr Gln Ala Gln Leu Gln Lys Asn Leu
50 55 60
Leu Tyr Leu Ala Ala Ile Ala Asp Ala Gln Pro Gln Pro Pro Gln Asn
65 70 75 80
Pro Ala Gly Arg Pro Gln Met Met Gln Pro Gly Ile Val Pro Gly Ala
85 90 95
Gly His Tyr Met Ser Gln Val Pro Met Phe Pro Pro Arg Thr Pro Leu
100 105 110
Thr Pro Gln Gln Met Gln Glu Gln Gln Gln Gln Gln Leu Gln Gln Gln
115 120 125
Gln Ala Gln Ala Leu Thr Phe Pro Gly Gln Met Val Met Arg Pro Ala
130 135 140
Thr Ile Asn Gly Ile Gln Gln Pro Met Gln Ala Asp Pro Ala Arg Ala
145 150 155 160
Ala Glu Leu Gln Gln Pro Pro Pro Ile Pro Ala Asp Gly Arg Val Ser
165 170 175
Lys Gln Gln Asp Thr Thr Ala Gly Val Ser Ser Glu Pro Ser Ala Asn
180 185 190
Glu Ser His Lys Thr Thr Thr Gly Ala Asp Ser Glu Ala Gly Gly Asp
195 200 205
Val Ala Glu Lys Ser
210
<210>77
<211>645
<212>DNA
<213>马铃薯
<400>77
atgcagcagc acctgatgca gatgcagccc atgatggcag cttactatcc aacgaacgtc 60
actactgacc atattcaaca gtatttggat gagaacaaat cactcattct gaaaattgtt 120
gagagccaaa actcgggaaa actcagtgaa tgtgcagaga accaagctag gcttcagagg 180
aatctgatgt accttgctgc tattgctgat tcacaacctc agccttctag catgcattct 240
cagttctctt ctggtgggat gatgcagcca gggacacaca gttacctgca gcagcagcag 300
cagcaacaac aagcgcaaca aatggcaaca caacaactca tggctgcaag atcctcatca 360
atgctctatg gacaacaaca gcagcagcag cagcagtctc agttatcaca atttcaacaa 420
ggcttgcata gtagccaact tggcatgagt tctggcagtg gtggaagcac tggacttcat 480
cacatgcttc aaagtgaatc atcacctcat ggtggtggtt tctctcatga cttcggccgt 540
gcaaataagc aagacattgg gagtagtatg tctgctgaag ggcgcggcgg aagctcaggt 600
ggtgatggtg gtgagaatct ttatctgaaa gcttctgagg attga 645
<210>78
<211>214
<212>PRT
<213>马铃薯
<400>78
Met Gln Gln His Leu Met Gln Met Gln Pro Met Met Ala Ala Tyr Tyr
1 5 10 15
Pro Thr Asn Val Thr Thr Asp His Ile Gln Gln Tyr Leu Asp Glu Asn
20 25 30
Lys Ser Leu Ile Leu Lys Ile Val Glu Ser Gln Asn Ser Gly Lys Leu
35 40 45
Ser Glu Cys Ala Glu Asn Gln Ala Arg Leu Gln Arg Asn Leu Met Tyr
50 55 60
Leu Ala Ala Ile Ala Asp Ser Gln Pro Gln Pro Ser Ser Met His Ser
65 70 75 80
Gln Phe Ser Ser Gly Gly Met Met Gln Pro Gly Thr His Ser Tyr Leu
85 90 95
Gln Gln Gln Gln Gln Gln Gln Gln Ala Gln Gln Met Ala Thr Gln Gln
100 105 110
Leu Met Ala Ala Arg Ser Ser Ser Met Leu Tyr Gly Gln Gln Gln Gln
115 120 125
Gln Gln Gln Gln Ser Gln Leu Ser Gln Phe Gln Gln Gly Leu His Ser
130 135 140
Ser Gln Leu Gly Met Ser Ser Gly Ser Gly Gly Ser Thr Gly Leu His
145 150 155 160
His Met Leu Gln Ser Glu Ser Ser Pro His Gly Gly Gly Phe Ser His
165 170 175
Asp Phe Gly Arg Ala Asn Lys Gln Asp Ile Gly Ser Ser Met Ser Ala
180 185 190
Glu Gly Arg Gly Gly Ser Ser Gly Gly Asp Gly Gly Glu Asn Leu Tyr
195 200 205
Leu Lys Ala Ser Glu Asp
210
<210>79
<211>645
<212>DNA
<213>两色蜀黍(Sorghum bicolor)
<400>79
atgcagcagc agatgcccat gccgccggcg cccgctgcgg cggcggcgac ggcgcccccg 60
gcggccggca tcaccaccga gcagatccag aagtatttgg acgaaaataa gcaacttatt 120
ttggccatcc tagaaaatca gaacttagga aagttggctg aatgtgctca gtatcaagct 180
caacttcaaa agaacctctt gtacctggct gcgattgctg atgcccaacc ccgaccaccg 240
caaaaccctg caggtcgccc tcagatgatg caacctggta tagtgccagg tgcagggcat 300
tacatgtcac aagtaccaat gttccctcca agaactccat taaccccaca gcaaatgcaa 360
gagcagcagc agcaacagct tcagcagcag caagcgcagg ctcttgcatt ccctgggcag 420
atggtcatga gaccagctac catcaacggc atgcagcagc ctatgcaggc tgaccctgcc 480
cgggcagcgg agctgcaaca gccagcatct gtcccagccg acgggcgagt aagcaagcag 540
gacacagcgg ctggggtgag ctcagagcct tctgccaatg agagccacaa gaccacaacc 600
ggagcagata gtgaggcagg tggagacgtg gcggagaaat cctaa 645
<210>80
<211>214
<212>PRT
<213>两色蜀黍
<400>80
Met Gln Gln Gln Met Pro Met Pro Pro Ala Pro Ala Ala Ala Ala Ala
1 5 10 15
Thr Ala Pro Pro Ala Ala Gly Ile Thr Thr Glu Gln Ile Gln Lys Tyr
20 25 30
Leu Asp Glu Asn Lys Gln Leu Ile Leu Ala Ile Leu Glu Asn Gln Asn
35 40 45
Leu Gly Lys Leu Ala Glu Cys Ala Gln Tyr Gln Ala Gln Leu Gln Lys
50 55 60
Asn Leu Leu Tyr Leu Ala Ala Ile Ala Asp Ala Gln Pro Arg Pro Pro
65 70 75 80
Gln Asn Pro Ala Gly Arg Pro Gln Met Met Gln Pro Gly Ile Val Pro
85 90 95
Gly Ala Gly His Tyr Met Ser Gln Val Pro Met Phe Pro Pro Arg Thr
100 105 110
Pro Leu Thr Pro Gln Gln Met Gln Glu Gln Gln Gln Gln Gln Leu Gln
115 120 125
Gln Gln Gln Ala Gln Ala Leu Ala Phe Pro Gly Gln Met Val Met Arg
130 135 140
Pro Ala Thr Ile Asn Gly Met Gln Gln Pro Met Gln Ala Asp Pro Ala
145 150 155 160
Arg Ala Ala Glu Leu Gln Gln Pro Ala Ser Val Pro Ala Asp Gly Arg
165 170 175
Val Ser Lys Gln Asp Thr Ala Ala Gly Val Ser Ser Glu Pro Ser Ala
180 185 190
Asn Glu Ser His Lys Thr Thr Thr Gly Ala Asp Ser Glu Ala Gly Gly
195 200 205
Asp Val Ala Glu Lys Ser
210
<210>81
<211>558
<212>DNA
<213>普通小麦(Triticum aestivum)
<400>81
atgcagcaag cgatgcccat gccgccggcg gcggcggcgc cggggatgcc tccgtctgct 60
ggcctcagca ccgagcagat ccaaaagtac ctggatgaaa ataagcaact aattttggct 120
atcttggaaa atcagaacct gggaaagttg gcggaatgtg ctcagtatca agctcagctt 180
cagaagaatc ttttgtattt ggctgcaatc gctgatactc agccacagac cactgtaagc 240
cgtcctcaga tggcaccacc tagtgcatcc ccaggggcag ggcattacat gtcacaggtg 300
ccaatgttcc ctccgaggac ccctctaacg cctcagcaga tgcaggagca gcaactacag 360
cagcaacagg ctcagatgct tccgtttgct ggtcaaatgg ttgcgagacc tggggctgtc 420
aatggcatgc ctcaggcccc tcaagttgaa ccagcctatg cagcaggtgg ggccagttct 480
gagccttctg gcactgagag ccacaggagc actggtgccg ataatgacgg ggggagcggc 540
tgggctgatc agtcctaa 558
<210>82
<211>185
<212>PRT
<213>普通小麦
<400>82
Met Gln Gln Ala Met Pro Met Pro Pro Ala Ala Ala Ala Pro Gly Met
1 5 10 15
Pro Pro Ser Ala Gly Leu Ser Thr Glu Gln Ile Gln Lys Tyr Leu Asp
20 25 30
Glu Asn Lys Gln Leu Ile Leu Ala Ile Leu Glu Asn Gln Asn Leu Gly
35 40 45
Lys Leu Ala Glu Cys Ala Gln Tyr Gln Ala Gln Leu Gln Lys Asn Leu
50 55 60
Leu Tyr Leu Ala Ala Ile Ala Asp Thr Gln Pro Gln Thr Thr Val Ser
65 70 75 80
Arg Pro Gln Met Ala Pro Pro Ser Ala Ser Pro Gly Ala Gly His Tyr
85 90 95
Met Ser Gln Val Pro Met Phe Pro Pro Arg Thr Pro Leu Thr Pro Gln
100 105 110
Gln Met Gln Glu Gln Gln Leu Gln Gln Gln Gln Ala Gln Met Leu Pro
115 120 125
Phe Ala Gly Gln Met Val Ala Arg Pro Gly Ala Val Asn Gly Met Pro
130 135 140
Gln Ala Pro Gln Val Glu Pro Ala Tyr Ala Ala Gly Gly Ala Ser Ser
145 150 155 160
Glu Pro Ser Gly Thr Glu Ser His Arg Ser Thr Gly Ala Asp Asn Asp
165 170 175
Gly Gly Ser Gly Trp Ala Asp Gln Ser
180 185
<210>83
<211>603
<212>DNA
<213>普通小麦
<400>83
atgcagcagg cgatgtcctt gcccccggga gcggtcggcg cggtgtcctc gccggccggc 60
atcaccaccg agcagatcca aaagtatttg gatgaaaata agcaacttat tttggccatc 120
cttgaaaatc agaacctagg aaagttggct gaatgtgctc agtatcaagc tcaactccaa 180
aagaatctct tgtatctagc tgctatcgcg gatgcccaac caccacagaa ccctacaagt 240
caccctcaga tggtgcagcc tggtagtatg caaggtgcag ggcattacat gtcacaagta 300
ccaatgttcc ctccaagaac gcctttaacc ccacagcaga tgcaagagca gcagcaccag 360
cagcttcagc agcagcaagc ccaggccctt tctttccccg cccaggtggt catgagacca 420
ggcaccgtca acggcatgca gcagcctatg caagcagccg gcgacctcca gccagcagca 480
gcacctggag ggagcaagca ggacgccgca gtggctgggg ccagctcgga accatctggc 540
accaagagcc acaagaacgc gggagcagag gaggtgggcg ctgatgtagc agaacaatcc 600
taa 603
<210>84
<211>200
<212>PRT
<213>普通小麦
<400>84
Met Gln Gln Ala Met Ser Leu Pro Pro Gly Ala Val Gly Ala Val Ser
1 5 10 15
Ser Pro Ala Gly Ile Thr Thr Glu Gln Ile Gln Lys Tyr Leu Asp Glu
20 25 30
Asn Lys Gln Leu Ile Leu Ala Ile Leu Glu Asn Gln Asn Leu Gly Lys
35 40 45
Leu Ala Glu Cys Ala Gln Tyr Gln Ala Gln Leu Gln Lys Asn Leu Leu
50 55 60
Tyr Leu Ala Ala Ile Ala Asp Ala Gln Pro Pro Gln Asn Pro Thr Ser
65 70 75 80
His Pro Gln Met Val Gln Pro Gly Ser Met Gln Gly Ala Gly His Tyr
85 90 95
Met Ser Gln Val Pro Met Phe Pro Pro Arg Thr Pro Leu Thr Pro Gln
100 105 110
Gln Met Gln Glu Gln Gln His Gln Gln Leu Gln Gln Gln Gln Ala Gln
115 120 125
Ala Leu Ser Phe Pro Ala Gln Val Val Met Arg Pro Gly Thr Val Asn
130 135 140
Gly Met Gln Gln Pro Met Gln Ala Ala Gly Asp Leu Gln Pro Ala Ala
145 150 155 160
Ala Pro Gly Gly Ser Lys Gln Asp Ala Ala Val Ala Gly Ala Ser Ser
165 170 175
Glu Pro Ser Gly Thr Lys Ser His Lys Asn Ala Gly Ala Glu Glu Val
180 185 190
Gly Ala Asp Val Ala Glu Gln Ser
195 200
<210>85
<211>672
<212>DNA
<213>葡萄(Vitis vinifera)
<400>85
atgcagcagc acctgatgca gatgcagccc atgatggcag cctattaccc cagcaacgtc 60
accactgatc acattcagca gtatcttgat gaaaacaagt cattgattct gaagattgtt 120
gagagccaga attcaggaaa attgactgaa tgtgcagaga accaggcaag actacagaga 180
aacctcatgt acctggctgc aattgctgat tctcaacccc aaccacccac catgcatgct 240
cagttccctc ctagtggcat tgttcagcca ggagctcact acatgcaaca ccaacaagct 300
caacaaatga caccacagtc gctcctggct gcacgctcct ccatgctgta cacccaacaa 360
ccattttcgg ccctgcaaca acaacaagcc atccatagcc agcttggcat gggctctggt 420
ggaagtgcag gacttcacat gctgcaaagc gaggggagta atccaggagg caatggaaca 480
ctggggactg gtgggtttcc tgatttcagc cgtggaactt ctggagaagg cctgcaggct 540
gcaggcaggg gaatggctgg tgggagcaag caagatatgg gaaatgcaga agggcgagga 600
gggaactcag gaggtcaggg tggggatgga ggtgagactc tttacttgaa agctgctgaa 660
gatgggaatt ga 672
<210>86
<211>223
<212>PRT
<213>葡萄
<400>86
Met Gln Gln His Leu Met Gln Met Gln Pro Met Met Ala Ala Tyr Tyr
1 5 10 15
Pro Ser Asn Val Thr Thr Asp His Ile Gln Gln Tyr Leu Asp Glu Asn
20 25 30
Lys Ser Leu Ile Leu Lys Ile Val Glu Ser Gln Asn Ser Gly Lys Leu
35 40 45
Thr Glu Cys Ala Glu Asn Gln Ala Arg Leu Gln Arg Asn Leu Met Tyr
50 55 60
Leu Ala Ala Ile Ala Asp Ser Gln Pro Gln Pro Pro Thr Met His Ala
65 70 75 80
Gln Phe Pro Pro Ser Gly Ile Val Gln Pro Gly Ala His Tyr Met Gln
85 90 95
His Gln Gln Ala Gln Gln Met Thr Pro Gln Ser Leu Leu Ala Ala Arg
100 105 110
Ser Ser Met Leu Tyr Thr Gln Gln Pro Phe Ser Ala Leu Gln Gln Gln
115 120 125
Gln Ala Ile His Ser Gln Leu Gly Met Gly Ser Gly Gly Ser Ala Gly
130 135 140
Leu His Met Leu Gln Ser Glu Gly Ser Asn Pro Gly Gly Asn Gly Thr
145 150 155 160
Leu Gly Thr Gly Gly Phe Pro Asp Phe Ser Arg Gly Thr Ser Gly Glu
165 170 175
Gly Leu Gln Ala Ala Gly Arg Gly Met Ala Gly Gly Ser Lys Gln Asp
180 185 190
Met Gly Asn Ala Glu Gly Arg Gly Gly Asn Ser Gly Gly Gln Gly Gly
195 200 205
Asp Gly Gly Glu Thr Leu Tyr Leu Lys Ala Ala Glu Asp Gly Asn
210 215 220
<210>87
<211>663
<212>DNA
<213>玉蜀黍
<400>87
atgcagcagc agatgcccat gccgccggcg cccgctgccg ccgcggcggc ggcgcccccg 60
gcggcaggca tcactaccga gcagatccag aagtatttgg acgaaaataa gcaacttatt 120
ttggccatcc tggaaaatca gaacttaggg aagttggctg aatgtgctca gtatcaagct 180
caacttcaaa agaacctctt gtacctggct gcgattgctg atgcccaacc ccagcctccg 240
caaaaccctg caggtcgccc tcagatgatg cagcctggta tagtgccagg tgcggggcat 300
tacatgtcac aagtaccaat gttccctcca agaaccccat taaccccaca gcagatgcag 360
gagcagcagc aacaacaaca gtttcagcag cagcagcagc aagtgcaggc tcttacattt 420
cctggacaga tggtcatgag accaggcacc atcaacggca tgcagcagca gcagcctatg 480
caggctgacc ctgcccgggc agcagcggag ctgcagcagg cagcacctat cccagctgac 540
gggcgaggaa gcaagcagga caccgcgggt ggggcgagct cagagccttc tgccaatgag 600
agccacaaga gcgccaccgg agcagatacc gaggcaggtg gcgacgtggc cgagaaatcc 660
taa 663
<210>88
<211>220
<212>PRT
<213>玉蜀黍
<400>88
Met Gln Gln Gln Met Pro Met Pro Pro Ala Pro Ala Ala Ala Ala Ala
1 5 10 15
Ala Ala Pro Pro Ala Ala Gly Ile Thr Thr Glu Gln Ile Gln Lys Tyr
20 25 30
Leu Asp Glu Asn Lys Gln Leu Ile Leu Ala Ile Leu Glu Asn Gln Asn
35 40 45
Leu Gly Lys Leu Ala Glu Cys Ala Gln Tyr Gln Ala Gln Leu Gln Lys
50 55 60
Asn Leu Leu Tyr Leu Ala Ala Ile Ala Asp Ala Gln Pro Gln Pro Pro
65 70 75 80
Gln Asn Pro Ala Gly Arg Pro Gln Met Met Gln Pro Gly Ile Val Pro
85 90 95
Gly Ala Gly His Tyr Met Ser Gln Val Pro Met Phe Pro Pro Arg Thr
100 105 110
Pro Leu Thr Pro Gln Gln Met Gln Glu Gln Gln Gln Gln Gln Gln Phe
115 120 125
Gln Gln Gln Gln Gln Gln Val Gln Ala Leu Thr Phe Pro Gly Gln Met
130 135 140
Val Met Arg Pro Gly Thr Ile Asn Gly Met Gln Gln Gln Gln Pro Met
145 150 155 160
Gln Ala Asp Pro Ala Arg Ala Ala Ala Glu Leu Gln Gln Ala Ala Pro
165 170 175
Ile Pro Ala Asp Gly Arg Gly Ser Lys Gln Asp Thr Ala Gly Gly Ala
180 185 190
Ser Ser Glu Pro Ser Ala Asn Glu Ser His Lys Ser Ala Thr Gly Ala
195 200 205
Asp Thr Glu Ala Gly Gly Asp Val Ala Glu Lys Ser
210 215 220
<210>89
<211>2193
<212>DNA
<213>稻
<400>89
aatccgaaaa gtttctgcac cgttttcacc ccctaactaa caatataggg aacgtgtgct 60
aaatataaaa tgagacctta tatatgtagc gctgataact agaactatgc aagaaaaact 120
catccaccta ctttagtggc aatcgggcta aataaaaaag agtcgctaca ctagtttcgt 180
tttccttagt aattaagtgg gaaaatgaaa tcattattgc ttagaatata cgttcacatc 240
tctgtcatga agttaaatta ttcgaggtag ccataattgt catcaaactc ttcttgaata 300
aaaaaatctt tctagctgaa ctcaatgggt aaagagagag atttttttta aaaaaataga 360
atgaagatat tctgaacgta ttggcaaaga tttaaacata taattatata attttatagt 420
ttgtgcattc gtcatatcgc acatcattaa ggacatgtct tactccatcc caatttttat 480
ttagtaatta aagacaattg acttattttt attatttatc ttttttcgat tagatgcaag 540
gtacttacgc acacactttg tgctcatgtg catgtgtgag tgcacctcct caatacacgt 600
tcaactagca acacatctct aatatcactc gcctatttaa tacatttagg tagcaatatc 660
tgaattcaag cactccacca tcaccagacc acttttaata atatctaaaa tacaaaaaat 720
aattttacag aatagcatga aaagtatgaa acgaactatt taggtttttc acatacaaaa 780
aaaaaaagaa ttttgctcgt gcgcgagcgc caatctccca tattgggcac acaggcaaca 840
acagagtggc tgcccacaga acaacccaca aaaaacgatg atctaacgga ggacagcaag 900
tccgcaacaa ccttttaaca gcaggctttg cggccaggag agaggaggag aggcaaagaa 960
aaccaagcat cctcctcctc ccatctataa attcctcccc ccttttcccc tctctatata 1020
ggaggcatcc aagccaagaa gagggagagc accaaggaca cgcgactagc agaagccgag 1080
cgaccgcctt cttcgatcca tatcttccgg tcgagttctt ggtcgatctc ttccctcctc l140
cacctcctcc tcacagggta tgtgcccttc ggttgttctt ggatttattg ttctaggttg 1200
tgtagtacgg gcgttgatgt taggaaaggg gatctgtatc tgtgatgatt cctgttcttg 1260
gatttgggat agaggggttc ttgatgttgc atgttatcgg ttcggtttga ttagtagtat 1320
ggttttcaat cgtctggaga gctctatgga aatgaaatgg tttagggtac ggaatcttgc 1380
gattttgtga gtaccttttg tttgaggtaa aatcagagca ccggtgattt tgcttggtgt 1440
aataaaagta cggttgtttg gtcctcgatt ctggtagtga tgcttctcga tttgacgaag 1500
ctatcctttg tttattccct attgaacaaa aataatccaa ctttgaagac ggtcccgttg 1560
atgagattga atgattgatt cttaagcctg tccaaaattt cgcagctggc ttgtttagat 1620
acagtagtcc ccatcacgaa attcatggaa acagttataa tcctcaggaa caggggattc 1680
cctgttcttc cgatttgctt tagtcccaga attttttttc ccaaatatct taaaaagtca 1740
ctttctggtt cagttcaatg aattgattgc tacaaataat gcttttatag cgttatccta 1800
gctgtagttc agttaatagg taatacccct atagtttagt caggagaaga acttatccga 1860
tttctgatct ccatttttaa ttatatgaaa tgaactgtag cataagcagt attcatttgg 1920
attatttttt ttattagctc tcaccccttc attattctga gctgaaagtc tggcatgaac 1980
tgtcctcaat tttgttttca aattcacatc gattatctat gcattatcct cttgtatcta 2040
cctgtagaag tttctttttg gttattcctt gactgcttga ttacagaaag aaatttatga 2100
agctgtaatc gggatagtta tactgcttgt tcttatgatt catttccttt gtgcagttct 2160
tggtgtagct tgccactttc accagcaaag ttc 2193
<210>90
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<212>PRT
<213>人工序列
<220>
<223>盒I
<220>
<221>MISC_FEATURE
<222>(3)..(3)
<223>Xaa可以是Gln或Lys
<220>
<221>MISC_FEATURE
<222>(4)..(4)
<223>Xaa可以是Tyr、Met、Phe或His中任一
<220>
<221>MISC_FEATURE
<222>(6)..(6)
<223>Xaa可以是Asp或Glu
<220>
<221>MISC_FEATURE
<222>(7)..(7)
<223>Xaa可以是Glu或Asp
<220>
<221>MISC_FEATURE
<222>(9)..(9)
<223>Xaa可以是Lys或Asn
<220>
<221>MISC_FEATURE
<222>(10)..(10)
<223>Xaa可以是任何氨基酸
<400>90
Ile Gln Xaa Xaa Leu Asp Xaa Asn Xaa Xaa Leu Ile
1 5 10
<210>9l
<211>10
<212>PRT
<213>人工序列
<220>
<223>盒II
<220>
<221>MISC_FEATURE
<222>(3)..(3)
<223>Xaa可以是Met、Leu或Val中任一
<220>
<221>misc_feature
<222>(7)..(7)
<223>Xaa可以是任何天然存在的氨基酸
<220>
<221>MISC_FEATURE
<222>(8)..(8)
<223>Xaa可以是Ala或Thr
<400>91
Asn Leu Xaa Tyr Leu Ala Xaa Ile Ala Asp
1 5 10
<210>92
<211>53
<212>DNA
<213>人工序列
<220>
<223>引物:prm06681
<400>92
ggggacaagt ttgtacaaaa aagcaggctt aaacaatgca acagcacctg atg 53
<210>93
<211>50
<212>DNA
<213>人工序列
<220>
<223>引物:prm06682
<400>93
ggggaccact ttgtacaaga aagctgggtc atcattaaga ttccttgtgc 50
<210>94
<211>53
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<213>人工序列
<220>
<223>引物:prm06685
<400>94
ggggacaagt ttgtacaaaa aagcaggctt aaacaatgca gcagcagcag tct 53
<210>95
<211>50
<212>DNA
<213>人工序列
<220>
<223>引物:prm06686
<400>95
ggggaccact ttgtacaaga aagctgggtt ctttggatcc ttttcacttg 50
<210>96
<211>55
<212>DNA
<213>人工序列
<220>
<223>引物:prm06683
<400>96
ggggacaagt ttgtacaaaa aagcaggctt aaacaatgca gcaatctcca cagat 55
<210>97
<211>52
<212>DNA
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<220>
<223>引物:prm06684
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ggggaccact ttgtacaaga aagctgggtt cctctatttc attttccttc ag 52
Claims (35)
1.增加相对于相应野生型植物的植物产率的方法,包括调节植物中滑膜肉瘤转位(SYT)多肽或其同源物编码核酸的表达,和任选地选择产率增加的植物,其中所述SYT多肽或其同源物从N末端到C末端包含:(i)与SEQID NO:2的SNH结构域具有至少40%序列同一性的SNH结构域;和(ii)Met富含结构域;和(iii)QG富含结构域。
2.根据权利要求1的方法,其中所述SNH结构域包含图2中显示为黑色的残基。
3.根据权利要求1的方法,其中所述SNH结构域由SEQ ID NO:1所代表。
4.根据权利要求1到3中任一项的方法,其中所述SYT多肽或其同源物还包含如下一个或多个序列:(i)SEQ ID NO:90;(ii)SEQ ID NO:91;(iii)位于SNH结构域之前的N末端的Met富含结构域。
5.根据权利要求1到4中任一项的方法,其中通过优选地在编码SYT多肽或其同源物的基因座引入遗传修饰来实现所述表达的调节。
6.根据权利要求5的方法,其中通过T-DNA激活、TILLING、定点诱变或定向进化中任一实现所述遗传修饰。
7.增加相对于相应野生型植物的产率、特别是种子产率的方法,包括在植物、植物部分或植物细胞中引入和表达SYT核酸或其变体。
8.根据权利要求7的方法,其中所述变体是SYT核酸的部分或能够与SYT核酸杂交的序列,所述部分或杂交序列编码多肽,所述多肽从N末端到C末端包含:(i)与SEQ ID NO:2的SNH结构域具有至少40%序列同一性的SNH结构域;和(ii)Met富含结构域;和(iii)QG富含结构域。
9.根据权利要求7的方法,其中所述SNH结构域包含图2中显示为黑色的残基。
10.根据权利要求7的方法,其中所述SNH结构域由SEQ ID NO:1所代表。
11.根据权利要求7到10中任一项的方法,其中所述SYT多肽或其同源物还包含如下一个或多个序列:(i)SEQ ID NO:90;(ii)SEQ ID NO:91;(iii)位于SNH结构域之前的N末端的Met富含结构域。
12.根据权利要求7到11中任一项的方法,其中所述SYT核酸或其变体在植物中过表达。
13.根据权利要求7到12中任一项的方法,其中所述SYT核酸或其变体是植物来源的,优选来自双子叶植物,还优选来自十字花科,更加优选核酸来自拟南芥。
14.根据权利要求7到13中任一项的方法,其中所述变体编码SEQ ID NO:4、SEQ ID NO:6和SEQ ID NO:8的SYT蛋白质的直系同源物或旁系同源物。
15.根据权利要求7到14任一项的方法,其中所述SYT核酸或其变体有效连接于组成型启动子。
16.根据权利要求15的方法,其中所述组成型启动子是植物来源的,优选来自于单子叶植物。
17.根据权利要求15或16的方法,其中所述组成型启动子是GOS2启动子。
18.根据权利要求1到17中任一项的方法,其中所述增加的产率是增加的种子产率。
19.根据权利要求1到18中任一项的方法,其中所述增加的产率是增加的种子产率和/或增加的TKW。
20.可根据权利要求1到19中任一项的方法获得的植物、植物部分或植物细胞。
21.构建体,含有:
(i)SYT核酸或其变体,
(ii)能够驱动(a)的核酸序列表达的一个或多个控制序列;和任选的
(iii)转录终止序列。
22.根据权利要求21的构建体,其中所述控制序列是来源于单子叶植物的组成型启动子。
23.根据权利要求22的构建体,其中所述组成型启动子是GOS2启动子。
24.根据权利要求23的构建体,其中所述GOS2启动子由SEQID NO:89所代表。
25.由根据权利要求21到24中任一项的构建体转化的植物、植物部分或植物细胞。
26.产生产率增加、特别是种子产率增加的转基因植物、优选转基因单子叶植物的方法,该方法包括:
(i)在植物或植物细胞中引入和表达编码SYT核酸或其变体;
(ii)在促进植物生长和发育的条件下培养植物细胞。
27.根据权利要求26的方法,包括通过杂交由培养步骤(ii)获得的植物,来产生一个或多个后续世代的植物或其包括种子的部分。
28.产率增加、特别是种子产率增加的转基因植物或其部分,其通过将SYT核酸或其变体引入所述植物或植物部分产生,所述产率增加是相对于相应野生型植物而言的。
29.根据权利要求20、25或28的转基因植物,其中所述植物是单子叶植物,如甘蔗,或者其中所述植物是谷类,如稻、玉米、小麦、大麦、粟、黑麦、燕麦或高粱。
30.根据权利要求20、25、28或29中任一项的植物的可收获部分。
31.根据权利要求30的植物可收获部分,其中所述可收获部分是种子。
32.从根据权利要求2 9的植物,和/或从根据权利要求30或31的植物可收获部分衍生、优选直接衍生的产品。
33.SYT核酸/基因或其变体、或者SYT多肽或其同源物、或者根据权利要求21到24中任一项的载体在相对于相应野生型植物提高产率、特别是种子产率中的用途。
34.根据权利要求33的用途,其中所述种子产率是增加的种子总产率和增加的TKW。
35.SYT核酸/基因或其变体、或者SYT多肽或其同源物作为分子标记的用途。
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Cited By (1)
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CN103517988A (zh) * | 2011-05-09 | 2014-01-15 | 巴斯夫植物科学有限公司 | 具有增强的产量相关性状的植物及其制备方法 |
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NZ556605A (en) | 2010-03-26 |
KR101356311B1 (ko) | 2014-02-06 |
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EP2298920A1 (en) | 2011-03-23 |
PH12013501782A1 (en) | 2015-12-02 |
US8426683B2 (en) | 2013-04-23 |
WO2006079655A3 (en) | 2006-11-23 |
RU2463351C2 (ru) | 2012-10-10 |
EP1844151A2 (en) | 2007-10-17 |
CA2595672A1 (en) | 2006-08-03 |
KR20070111458A (ko) | 2007-11-21 |
BRPI0607211A2 (pt) | 2009-12-22 |
US20130269049A1 (en) | 2013-10-10 |
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