CN112226459A - 一种普通野生稻粒型相关编码基因及其应用 - Google Patents
一种普通野生稻粒型相关编码基因及其应用 Download PDFInfo
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- CN112226459A CN112226459A CN202011101224.3A CN202011101224A CN112226459A CN 112226459 A CN112226459 A CN 112226459A CN 202011101224 A CN202011101224 A CN 202011101224A CN 112226459 A CN112226459 A CN 112226459A
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Abstract
本发明公开了一种普通野生稻粒型相关编码基因LTG5在调控水稻粒型中的应用，所述普通野生稻粒型相关编码基因LTG5来源于稻属普通野生稻(Oryzarufipogon Griff.)的Y12株系的DNA序列；所述调控水稻粒型是指调控水稻粒型变短、变窄。本发明首次鉴定了与普通野生稻粒型相关基因LTG5，普通野生稻粒型相关基因LTG5在功能增强或表达量上升的条件下会得到粒型普通野生稻，证明普通野生稻粒型相关基因蛋白或其蛋白在控制普通野生稻粒型中发挥重要作用；本发明获得的LTG5基因表达升高的转基因水稻，作为新的水稻种质材料，可用于研究粒型水稻机理和发现更多的调控普通野生稻粒型发育的基因。
Description
技术领域
本发明涉及基因工程技术领域，特别涉及一种普通野生稻粒型相关编码基因及其应用。
背景技术
水稻是人类最重要的粮食作物之一，世界上以稻米为主食的人口约占50％。亚洲地区对稻米的需求量逐年增加，由于经济快速发展、人口压力增大、工业用地剧增、耕地面积减少，水稻生产面临巨大的压力。而消费市场对优质水稻提出了更高的需求，对优质稻米的外观有着更细化的要求。因此对水稻优质化育种提供粒型基因资源，是科学家们的主要研究方向。水稻粒型与生产、品质、产量有着密切的关系。
已克隆的GS3基因包含5个外显子，编码一个由232个氨基酸组成的跨膜蛋白，对粒重起负调控作用。控制粒宽的GW2编码一个环型E3泛素连接酶，位于细胞质中，通过将其底物锚定到蛋白酶体进行降解，从而负调节细胞的分裂，最后控制谷粒的宽度。影响粒宽的GW5编码一个144氨基酸组成的核定位蛋白，该蛋白包含一个核定位信号和一个富精氨酸区域，通过泛素蛋白酶体途径调节粒宽和粒重。Yu等利用关联分析克隆了OsLG3,OsLG3b，OsLG3是粒长的正调控因子；OsLG3b编码MADS-box转录因子1(OsMADS1)，过表达OsLG3b基因能够使水稻谷粒变长。Ma等克隆了负调控粒径的OsSNB，Yuan等克隆了OsSPL18，通过积极调控DEP1的表达来影响细胞增殖来调控小穗外壳的发育，粒型基因的克隆在水稻品种改良中起着重要作用。
当今水稻优质化育种的主要问题是基因来源范围狭窄，导致水稻的优良基因选择范围不大。如何在野生稻里挖掘新的优良基因显得尤为关键。当广西普通野生稻中蕴藏着丰富的基因，挖掘普通野生稻的有利基因，从普通稻中发掘并鉴定粒型新资源，深入探究调控粒型的分子机制，将有助于水稻育种家在育种过程中为分子设计育种改良水稻优质提供理论依据，最终实现高效育种。
公开于该背景技术部分的信息仅仅旨在增加对本发明的总体背景的理解，而不应当被视为承认或以任何形式暗示该信息构成已为本领域一般技术人员所公知的现有技术。
发明内容
本发明针对上述技术问题，提供一种普通野生稻粒型相关编码基因及其应用，旨在得到一种粒型相关的普通野生稻。
为实现上述目的，本发明提供的技术方案如下：
一种普通野生稻粒型相关编码基因，名称为LTG5，来源于稻属普通野生稻(Oryzarufipogon Griff.)的Y12株系的DNA序列，LTG5在水稻粒型中的应用。
其中，所述普通野生稻粒型相关编码基因LTG5为a)或b)：
a)cDNA序列如SEQ ID No.1所示的基因序列，由1419个碱基组成；
b)将SEQ ID No.1所示的基因序列经过一个或几个碱基的取代和/或缺失和/或添加得到的具有普通野生稻粒型相关基因编码的LTG5蛋白活性的由a)衍生的蛋白质。
其中，与所述普通野生稻粒型相关基因LTG5相关的生物材料，在培育粒型普通野生稻的转基因水稻中的应用；所述与普通野生稻粒型相关基因LTG5相关的生物材料为下述A1)至A20)中的任一种：
A1)核酸分子1；所述核酸分子1为编码所述普通野生稻粒型相关基因LTG5的核酸分子；
A2)含有A1)所述核酸分子1的表达盒；
A3)含有A1)所述核酸分子1的重组载体；
A4)含有A2)所述表达盒的重组载体；
A5)含有A1)所述核酸分子1的重组微生物；
A6)含有A2)所述表达盒的重组微生物；
A7)含有A3)所述重组载体的重组微生物；
A8)含有A4)所述重组载体的重组微生物；
A9)含有A1)所述核酸分子1的转基因植物细胞系；
A10)含有A2)所述表达盒的转基因植物细胞系；
A11)含有A3)所述重组载体的转基因植物细胞系；
A12)含有A4)所述重组载体的转基因植物细胞系；
A13)含有A1)所述核酸分子1的转基因植物组织；
A14)含有A2)所述表达盒的转基因植物组织；
A15)含有A3)所述重组载体的转基因植物组织；
A16)含有A4)所述重组载体的转基因植物组织；
A17)含有A1)所述核酸分子1的转基因植物器官；
A18)含有A2)所述表达盒的转基因植物器官；
A19)含有A3)所述重组载体的转基因植物器官；
A20)含有A4)所述重组载体的转基因植物器官。
上述与所述普通野生稻粒型相关基因LTG5相关的生物材料在调控水稻粒型中的应用，所述核酸分子可以是DNA，如cDNA、基因组DNA或重组DNA；所述核酸分子也可以是RNA，如mRNA或hnRNA等。
上述与所述普通野生稻粒型相关基因LTG5相关的生物材料中，所述表达盒是指能够在宿主细胞中表达相应蛋白质的DNA，该DNA不但可包括启动相关基因转录的启动子，还可包括终止相关基因转录的终止子，如A2)所述的含有编码所述普通野生稻粒型相关蛋白LTG5的核酸分子的表达盒，是指能够在宿主细胞中表达所述普通野生稻粒型相关基因LTG5的DNA。
构建含有所述LTG5基因表达盒的重组载体的现有的植物表达载体，为pET-28a、pCAMBIA2301、pSP72、pROKII、pBin438、pCAMBIA1302、pCAMBIA1301、pCAMBIA1300、pBI121、pCAMBIA1391-Xa或pCAMBIA1391-Xb(CAMBIA公司)等中的一种。
上述生物材料中，A5)-A8)中任一所述重组微生物或B5)-B8)中任一所述重组微生物具体可为细菌、酵母、藻或真菌。其中，细菌可来自埃希氏菌属(Escherichia)、欧文氏菌(Erwinia)、根癌农杆菌属(Agrobacterium)、黄杆菌属(Flavobacterium)、产碱菌属(Alcaligenes)、假单胞菌属(Pseudomonas)、芽胞杆菌属(Bacillus)等。
上述生物材料中，A9)-A12)中任一所述的转基因细胞系，A13)-A16)中任一所述的转基因植物组织，A17)-A20)中任一所述的转基因植物器官不包括植物的繁殖材料。
本发明所提供的一种培育粒型的转基因水稻的方法，将SEQ ID No.1所示的普通野生稻粒型相关基因LTG5构建过表达载体导入水稻中，获得LTG5基因过表达的转基因水稻。
其中，所述水稻为籼稻测253。
其中，所述过表达载体为重组表达载体PMDC32或载体pCAMBIA1301；将表达载体PMDC32或载体pCAMBIA1301的Asc I和PacI识别位点间的序列替换为SEQ ID No.1所示的DNA序列。
其中，所述重组表达载体PMDC32可通过使用农杆菌介导、Ti质粒，植物病毒载体，直接DNA转化，微注射，电穿孔等常规生物技术方法导入植物细胞或组织。
其中，所述方法还包括从导入SEQ ID No.1所示DNA序列的受体水稻中筛选出普通野生稻粒型相关基因LTG5表达量升高的水稻，得到所述LTG5基因表达水平升高的转基因水稻的步骤。
其中，所述转基因水稻理解为不仅包含将所述基因转化受体水稻得到的第一代转基因水稻，也包括其子代。对于转基因水稻，可以在该物种中繁殖该基因，也可用常规育种技术将该基因转移进入相同物种的其它品种，特别包括商业品种中。所述转基因水稻包括种子、愈伤组织、完整植株和细胞。
本发明将如SEQ ID No.1所示DNA序列的普通野生稻粒型相关编码基因LTG5导入水稻中，获得LTG5基因表达水平提高的转基因水稻。表达水平高于受体亲本水稻，而且该过表达转基因水稻出现了粒型的表现。
与现有技术相比，本发明具有如下有益效果：
本发明首次鉴定了与普通野生稻粒型相关基因LTG5，普通野生稻粒型相关基因LTG5在功能增强或表达量上升的条件下会得到粒型普通野生稻，证明普通野生稻粒型相关基因蛋白或其蛋白在控制普通野生稻粒型中发挥重要作用；本发明不仅为进一步阐明普通野生稻粒型的分子机理提供基础，而且为水稻育种提供新的基因资源和育种资源。本发明获得的LTG5基因表达升高的转基因水稻，作为新的水稻种质材料，可用于研究粒型水稻机理和发现更多的调控普通野生稻粒型发育的基因。
附图说明
图1为普通野生稻粒型相关基因LTG5表达水平升高的过表达转基因水稻的LTG5基因的转录水平的检测；其中，Ce253代表受体亲本籼稻测253植株，OE-LTG5-1、OE-LTG5b、OE-LTG5c代表转入重组载体PMDC32-LTG5的过表达阳性植株OE-LTG5独立转化事件。
图2为普通野生稻粒型相关基因LTG5表达水平升高的过表达转基因水稻的表型观察；其中，Ce253代表受体亲本籼稻测253植株，OE-LTG5-1、OE-LTG5b、OE-LTG5c代表转入重组载体PMDC32-LTG5的过表达阳性植株OE-LTG5独立转化事件。
具体实施方式
下面结合附图具体实施方式进行详细描述，但应当理解本发明的保护范围并不受具体实施方式的限制。
下述实施例中的实验方法，如无特殊说明，均为常规方法。
下述实施例中所用的材料、试剂等，如无特殊说明，均可从商业途径得到。
下述实施例中的水稻籼稻测253(也称为野生型水稻，简称Ce253)，公众可从广西壮族自治区农业科学院水稻研究所获得该生物材料，该生物材料只为重复本发明的相关实验所用，不可作为其它用途使用。
下述实施例中所用表达载体PMDC32为市售所得；公众也可从中广西壮族自治区农业科学院水稻研究所获得该生物材料，该生物材料只为重复本发明的相关实验所用，不可作为其它用途使用。
下述实施例中的农杆菌为根癌农杆菌EHA105(Agrobacterium tumefaciensEHA105)(New Agrobacterium helper plasmids for gene transfer toplants.Hood,ElizabethE；Gelvin,StantonB；Melchers,LeoS；Hoekema,Andre.Trans genic research,2(4):p.208-218(1993))为市售所得；公众也可从广西壮族自治区农业科学院水稻研究所获得该生物材料，该生物材料只为重复本发明的相关实验所用，不可作为其它用途使用。
实施例1
普通野生稻粒型相关基因LTG5过表达载体的构建
1、LTG5基因的获得
以普通野生稻Y12(Oryzarufipogon Griff.)的DNA为模板，用如下引物primer1和primer2进行PCR扩增获得目的基因：
primer1：5'CGGGGTACCATGACGACAAAGACCTTT 3'；
primer2：5'CTTAATTAATCAGCTGCGAACTCCATT 3'。
将PCR产物回收纯化后连接入Zero(购买自北京全式金公司)测序载体，转化DH5α感受态细胞，挑选阳性克隆后，进行测序。
测序结果表明，扩增得到的PCR产物序列如SEQ ID No.1所示的核苷酸序列，长度为1490bp，命名为LTG5基因，LTG5基因编码的蛋白质的氨基酸序列如SEQ ID No.2所示。
2、普通野生稻粒型相关基因LTG5过表达载体(重组表达载体OE-LTG5)的构建
1)用primer1和primer2扩增野生稻cDNA，获得LTG5基因的序列，并连接到重组载体Zero-LTG5，得到了Zero-LTG5的阳性克隆，用限制性内切酶AscI和PacI酶切重组载体Zero-LTG5获得LTG5片段，连接重组成OE-LTG5；
2)用限制性内切酶Asc I和PacI酶切表达载体PMDC32，得到线性表达载体PMDC32，回收该线性片段；将步骤1)中得到的片段1OE-LTG5采用同源重组定向克隆的方法整合到线性表达载体PMDC32上(具体方法参考PMDC32使用说明书)，得到同源重组产物1，再将同源重组产物1转入DH5α感受态细胞，37℃培养过夜，得到重组载体OE-LTG5；
3)对步骤2)所得重组载体OE-LTG5进行测序，结果表明该重组载体OE-LTG5是在表达载体PMDC32的Asc I酶切位点正向插入了如SEQ ID No.1所示的核苷酸序列，即成功将PMDC32的Asc I和PacI识别位点(识别序列)间的DNA序列替换为如SEQ ID No.1所示的DNA序列。
实施例2
培育普通野生稻粒型相关基因LTG5表达水平升高的过表达LTG5转基因植株及转基因植株的鉴定
一、培育普通野生稻粒型相关基因LTG5表达水平升高的过表达LTG5转基因植株，将重组载体OE-LTG5通过根癌农杆菌EHA105介导转化籼稻测253粳稻，具体方法如下：
1、将实施例1所得重组载体OE-LTG5用热激法导入根癌农杆菌EHA105中得到含有重组载体OE-LTG5的重组根癌农杆菌EHA105；将含有重组载体OE-LTG5的重组根癌农杆菌EHA105在28℃培养16h，收集菌体；采用含有浓度为100μM乙酰丁香酮的N6液体培养基(Sigma，产品目录号为C1416)将菌体进行稀释，得到稀释菌液，稀释菌液的OD600≈0.5；
2、将培养至一个月的水稻成熟胚胚性愈伤组织与步骤1的稀释菌液混合侵染30min，采用滤纸吸干菌液后转入N6固体共培养培养基中，在24℃共培养3d，得到共培养处理后的愈伤组织；
3、将步骤2的共培养处理后的愈伤组织接种在含有质量浓度为150mg/L潮霉素的N6固体筛选培养基(向N6固体培养基中加入潮霉素得到N6固体筛选培养基，N6固体筛选培养基中潮霉素的质量浓度为150mg/L)上进行第一次筛选；
4、在第一次筛选开始的第16天挑取健康愈伤组织转入含有质量浓度为200mg/L潮霉素的N6固体筛选培养基(向N6固体培养基中加入潮霉素得到N6固体筛选培养基，N6固体筛选培养基中潮霉素的质量浓度为200mg/L)上进行第二次筛选，每15天继代一次，共继代1次；
5、挑取步骤4获得的抗性愈伤组织转入含有质量浓度为150mg/L潮霉素的分化培养基上(分化培养基：6-BA 2mg，NAA 0.2mg，N6 4g，水解酪蛋白1g，肌醇0.1g，蔗糖25g，山梨醇2.4g，琼脂粉7g，去离子水1L)进行分化，在24℃培养45d(此时植株地上部分高度约为15cm)，打开瓶口炼苗3天，然后移栽至温室栽培，即为转OE-LTG5植株(T0代)。
二、普通野生稻粒型相关基因LTG5表达水平升高的OE-LTG5转基因植株的PCR鉴定提取
获得的转OE-LTG5植株的T0代幼苗和受体亲本水稻籼稻测253植株的幼苗(简称为Ce253)的基因组DNA，并采用引物
primer1：5'CGGGGTACCATGACGACAAAGACCTTT 3'；
primer2：5'CTTAATTAATCAGCTGCGAACTCCATT 3'；进行PCR分子检测鉴定阳性苗，得到1490bp PCR产物的植株为阳性苗，取其中三株分别标记为OE-LTG5a、OE-LTG5b、OE-LTG5c。
三、普通野生稻粒型相关基因LTG5表达水平升高的过表达转基因植株的LTG5基因表达水平的鉴定：
分别提取上述“二”步骤操作得到的OE-LTG5a、OE-LTG5b、OE-LTG5c植株和受体亲本水稻籼稻测253植株(简称为Ce253)叶片的RNA，设定内参为Actin，利用内参引物Actin-F和Actin-R，以及LTG5基因特异性定量引物LTG5-qRT-F和LTG5-qRT-R进行荧光定量PCR反应来检测不同转基因植株LTG5基因的表达水平的变化，上述引物如下：
Actin-F：5’-ATTTGGCACCACACATTCTAC-3’；
Actin-R：5’-ATAACCTTCGTAGATTGGGACT-3’；
LTG5-RT-F：5'CCCCGCCTACTTCTTCTTTC 3'
LTG5-RT-R:5'CGCCGCCTTATCCATCTC 3'
结果表明(图1)，在转入重组载体P OE-LTG5的阳性植株中LTG5基因的表达水平均比对照(Ce253)的LTG5基因的表达水平的显著上升。
四、普通野生稻粒型相关基因LTG5表达水平上升的LTG5过表达转基因植株的表型鉴定
分别将上述“二”步骤操作得到的OE-LTG5a、OE-LTG5b、OE-LTG5c和受体亲本水稻籼稻测253植株种植并收获T2代种子，对种子使用万深SC-G考种仪测量种子的粒宽数据，统计粒宽数据。观察结果如图2，与受体亲本水稻测253种子相比，OE-LTG5植株的种子粒宽比测253明显变小，并达到显著差异，从而证明了LTG5基因参与控制粒型普通野生水稻形成过程，即该LTG5基因为普通野生水稻粒型基因。
前述对本发明的具体示例性实施方案的描述是为了说明和例证的目的。这些描述并非想将本发明限定为所公开的精确形式，并且很显然，根据上述教导，可以进行很多改变和变化。对示例性实施例进行选择和描述的目的在于解释本发明的特定原理及其实际应用，从而使得本领域的技术人员能够实现并利用本发明的各种不同的示例性实施方案以及各种不同的选择和改变。本发明的范围意在由权利要求书及其等同形式所限定。
序列表
<110> 广西壮族自治区农业科学院
<120> 一种普通野生稻粒型相关基因及其应用
<130> JC
<160> 2
<170> SIPOSequenceListing 1.0
<210> 1
<211> 1419
<212> DNA
<213> 水稻(Oryza sativa L.)
<400> 1
atgacgacaa acacgtttgt gctgttcccg tcgctgggcg tcggccacct gaaccccatg 60
gtggagctag ccaagcacct gcgccgccgc ggcctcggcg tcatcatcgc ggtgatcgat 120
ccgcccaaca acgacgccat gtcggccgac gcgatggcgc gcctcgccgc ggccaaccct 180
tccgtcacgt tccgcatcct gccggcgccg gccagcccgg accccggcgc gcaccatgtc 240
aagcgcaacc tcgacacgct ccggctcgcc aaccccgtgc tccgcgagtt cctccgctcc 300
ctgcccgccg tcgacgcact cctgctcgac atgttctgcg tcgacgcgct cgacgtcgcg 360
gccgagctcg ccatccccgc ctacttcttc tttccctccc cggccagcgt cctcgccgtc 420
ttttcccacc tcccgtatta ctaccgcaac gcgccgtcgt tgagggagat ggataaggcg 480
gcgctcatcc gatttcccgg cattccgccg atccgcaacg tcgacatgct ggccacggtg 540
aaggacaagg agagcgagac gaccaagatc aggttgtacc agttcaagcg gatgatggaa 600
gggaagggcg tgctggtgaa tagcttcgac tggctggagc ccaaggccct gaaagcgctc 660
gccgccggtg tctgcgtgcc cgacatgccc aagcccagag tctacttaat cgggccactg 720
gtcgacgccg gcaagaagat agggagcggc gccgagaggc acgcgtgcct cccgtggctt 780
gacgcccagc cgcggcggag cgtcgtgttc ctctgcttcg gcagccaggg cgcgttcccg 840
gcggcgcagc tgaaggagtt agctcatggg ctggagagct ccggccaccg attcctgtgg 900
accgtgagga gcccaccgga ggagcagtcc acatcaccgg agccggacct ggagcggctg 960
cttccggcgg ggttcttgga gaggacgaag ggcagaggca tggtggtcaa gaactgggtg 1020
ccacaggcgg aggtggtgca gcacgaggcg gtaggcgcgt tcgtgacgca ctgcgggtgg 1080
aactcgacgc tggaggcgat catgtcggcg ctgccgatga tatgctggcc gctgtacgcg 1140
gagcaggcga tgaacaaggt gatcatggtg gaggagatga agatcgccgt gtcgctcgac 1200
gggtacgagg agggagggtt ggtgaaggcc gaggaagtgg agacgaaggt gaggctggtg 1260
atggagaccg aggaagggag aaagctcagg gagaaactgg tggagacgag ggacatggcg 1320
ttgaatgccg tcaaggatag tgggtcttct gaagtggcat ttgataagtt catgagagat 1380
ttggagaaga gcagattgga gaatggagtt cgcagctga 1419
<210> 2
<211> 472
<212> PRT
<213> 水稻(Oryza sativa L.)
<400> 2
Met Thr Thr Asn Thr Phe Val Leu Phe Pro Ser Leu Gly Val Gly His
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Gly Val Ile Ile Ala Val Ile Asp Pro Pro Asn Asn Asp Ala Met Ser
35 40 45
Ala Asp Ala Met Ala Arg Leu Ala Ala Ala Asn Pro Ser Val Thr Phe
50 55 60
Arg Ile Leu Pro Ala Pro Ala Ser Pro Asp Pro Gly Ala His His Val
65 70 75 80
Lys Arg Asn Leu Asp Thr Leu Arg Leu Ala Asn Pro Val Leu Arg Glu
85 90 95
Phe Leu Arg Ser Leu Pro Ala Val Asp Ala Leu Leu Leu Asp Met Phe
100 105 110
Cys Val Asp Ala Leu Asp Val Ala Ala Glu Leu Ala Ile Pro Ala Tyr
115 120 125
Phe Phe Phe Pro Ser Pro Ala Ser Val Leu Ala Val Phe Ser His Leu
130 135 140
Pro Tyr Tyr Tyr Arg Asn Ala Pro Ser Leu Arg Glu Met Asp Lys Ala
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Ala Leu Ile Arg Phe Pro Gly Ile Pro Pro Ile Arg Asn Val Asp Met
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Leu Ala Thr Val Lys Asp Lys Glu Ser Glu Thr Thr Lys Ile Arg Leu
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Tyr Gln Phe Lys Arg Met Met Glu Gly Lys Gly Val Leu Val Asn Ser
195 200 205
Phe Asp Trp Leu Glu Pro Lys Ala Leu Lys Ala Leu Ala Ala Gly Val
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Cys Val Pro Asp Met Pro Lys Pro Arg Val Tyr Leu Ile Gly Pro Leu
225 230 235 240
Val Asp Ala Gly Lys Lys Ile Gly Ser Gly Ala Glu Arg His Ala Cys
245 250 255
Leu Pro Trp Leu Asp Ala Gln Pro Arg Arg Ser Val Val Phe Leu Cys
260 265 270
Phe Gly Ser Gln Gly Ala Phe Pro Ala Ala Gln Leu Lys Glu Leu Ala
275 280 285
His Gly Leu Glu Ser Ser Gly His Arg Phe Leu Trp Thr Val Arg Ser
290 295 300
Pro Pro Glu Glu Gln Ser Thr Ser Pro Glu Pro Asp Leu Glu Arg Leu
305 310 315 320
Leu Pro Ala Gly Phe Leu Glu Arg Thr Lys Gly Arg Gly Met Val Val
325 330 335
Lys Asn Trp Val Pro Gln Ala Glu Val Val Gln His Glu Ala Val Gly
340 345 350
Ala Phe Val Thr His Cys Gly Trp Asn Ser Thr Leu Glu Ala Ile Met
355 360 365
Ser Ala Leu Pro Met Ile Cys Trp Pro Leu Tyr Ala Glu Gln Ala Met
370 375 380
Asn Lys Val Ile Met Val Glu Glu Met Lys Ile Ala Val Ser Leu Asp
385 390 395 400
Gly Tyr Glu Glu Gly Gly Leu Val Lys Ala Glu Glu Val Glu Thr Lys
405 410 415
Val Arg Leu Val Met Glu Thr Glu Glu Gly Arg Lys Leu Arg Glu Lys
420 425 430
Leu Val Glu Thr Arg Asp Met Ala Leu Asn Ala Val Lys Asp Ser Gly
435 440 445
Ser Ser Glu Val Ala Phe Asp Lys Phe Met Arg Asp Leu Glu Lys Ser
450 455 460
Arg Leu Glu Asn Gly Val Arg Ser
465 470
Claims (7)
1.一种普通野生稻粒型相关编码基因LTG5在调控水稻粒型中的应用，其特征在于：所述普通野生稻粒型相关编码基因LTG5来源于稻属普通野生稻(Oryzarufipogon Griff.)的Y12株系的DNA序列；所述调控水稻粒型是指调控水稻粒型变短、变窄；
其中，所述普通野生稻粒型相关编码基因LTG5为a)或b)：
a)cDNA序列如SEQ ID No.1所示的基因序列，由1419个碱基组成；
b)将SEQ ID No.1所示的基因序列经过一个或几个碱基的取代和/或缺失和/或添加得到的具有普通野生稻粒型相关基因编码的LTG5蛋白活性的由a)衍生的蛋白质。
2.根据权利要求1所述的应用，其特征在于：与所述普通野生稻粒型相关基因LTG5相关的生物材料，在培育粒型普通野生稻的转基因水稻中的应用。
3.根据权利要求1所述应用，其特征在于：通过将SEQ ID No.1所示的普通野生稻粒型相关基因LTG5构建过表达载体导入水稻中，获得LTG5基因过表达的转基因水稻。
4.根据权利要求3所述应用，其特征在于：所述水稻为籼稻测253。
5.根据权利要求3所述应用，其特征在于：所述过表达载体为重组表达载体PMDC32或载体pCAMBIA1301；将表达载体PMDC32或载体pCAMBIA1301的Asc I和PacI识别位点间的序列替换为SEQ ID No.1所示的DNA序列。
6.根据权利要求5所述应用，其特征在于：所述重组表达载体PMDC32可通过使用农杆菌介导、Ti质粒，植物病毒载体，直接DNA转化，微注射，电穿孔等常规生物技术方法导入植物细胞或组织。
7.根据权利要求3所述应用，其特征在于：还包括从导入SEQ ID No.1所示DNA序列的受体水稻中筛选出普通野生稻粒型相关基因LTG5表达量升高的水稻，得到所述LTG5基因表达水平升高的转基因水稻的步骤。
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