CN106995492A - 蔗糖转运蛋白及其在调控植物雄性不育中的应用 - Google Patents
蔗糖转运蛋白及其在调控植物雄性不育中的应用 Download PDFInfo
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Abstract
本发明公开了一种蔗糖转运蛋白及其在调控植物雄性不育中的应用。本发明提供了一种蛋白质,获自黄瓜,命名为CsSUT1蛋白,是如下(a1)或(a2):(a1)由序列表中序列1所示的氨基酸序列组成的蛋白质;(a2)将序列表中序列1所示的氨基酸序列经过一个或几个氨基酸残基的取代和/或缺失和/或添加且与植物雄性育性相关的由序列1衍生的蛋白质。本发明还保护所述蛋白质的编码基因(CsSUT1基因)。本发明通过实验证明,下调CsSUT1基因的表达量会导致黄瓜植株雄性不育。本发明可以应用到黄瓜育种中,对提高育种效率有着积极的作用。
Description
技术领域
本发明涉及一种蔗糖转运蛋白及其在调控植物雄性不育中的应用。
背景技术
黄瓜属于葫芦科甜瓜属,为重要的园艺作物,在世界各地普遍栽培,是我国主栽蔬菜作物之一。糖是植物光合作用的主要产物,也是植物长距离运输和贮藏的主要形式,更是协调植物源库关系的重要信号分子。叶片制造的糖经过韧皮部长距离运输,到达并在花、果实、种子等库器官中卸出,其在库器官中卸出的方式、卸出的速度对产量和品质形成具有重要作用。对大多数植物来说,蔗糖是其光合产物的主要运输形式,少数作物以棉籽糖家族系列寡糖为主要运输形式。糖到达库器官以后会以多种方式卸出,分解或重新转运的过程需要各种转运蛋白和酶的参与,其代谢转运机制相当复杂。蔗糖转运蛋白作为运输蔗糖的工具,参与韧皮部糖卸载,在很大程度上调节着植株源库关系,调节着植株碳素分配。在黄瓜中研究蔗糖转运蛋白的作用及分子机理,有利于丰富和完善同化物源库运输细胞学路径理论体系,为实现黄瓜优质高产栽培提供理论参考。
植物雄性不育一直是近年来的研究热点,因为在杂交育种中雄性不育系可以极大地节省人工,节约时间,减少种子生产成本,著名的三系法杂交水稻育种就是应用了雄性不育系大大提高了种子生产效率。黄瓜的花属于雌雄异花同株,雌花包括下位的子房和上位的柱头及花瓣、萼片,雄花包括花药、花瓣、花萼。花器官在植株生长发育过程中属于非常强大的库,花从分化到开放的整个发育过程一般需要在数天之内完成,此过程需要大量的糖分供应,糖分供应不及时往往会造成花器官发育异常甚至败育。
发明内容
本发明的目的是提供一种蔗糖转运蛋白及其在调控植物雄性不育中的应用。
本发明提供的蛋白质,获自黄瓜,命名为CsSUT1蛋白,是如下(a1)或(a2):
(a1)由序列表中序列1所示的氨基酸序列组成的蛋白质;
(a2)将序列表中序列1所示的氨基酸序列经过一个或几个氨基酸残基的取代和/或缺失和/或添加且与植物雄性育性相关的由序列1衍生的蛋白质。
为了使(a1)中的CsSUT1蛋白便于纯化和检测,可在由序列表中序列1所示的氨基酸序列组成的蛋白质的氨基末端或羧基末端连接上如表1所示的标签。
表1标签的序列
标签 | 残基 | 序列 |
Poly-Arg | 5-6(通常为5个) | RRRRR |
Poly-His | 2-10(通常为6个) | HHHHHH |
FLAG | 8 | DYKDDDDK |
Strep-tag II | 8 | WSHPQFEK |
c-myc | 10 | EQKLISEEDL |
上述(a2)中的CsSUT1蛋白可人工合成,也可先合成其编码基因,再进行生物表达得到。上述(a2)中的CsSUT1蛋白的编码基因可通过将序列表中序列2所示的DNA序列中缺失一个或几个氨基酸残基的密码子,和/或进行一个或几个碱基对的错义突变,和/或在其5′端和/或3′端连上表1所示的标签的编码序列得到。
编码所述CsSUT1蛋白的基因(CsSUT1基因)也属于本发明的保护范围。
所述CsSUT1基因为如下(b1)-(b3)中任一所述的DNA分子:
(b1)编码区如序列表中序列2所示的DNA分子;
(b2)在严格条件下与(b1)限定的DNA序列杂交且编码与植物雄性育性相关蛋白的DNA分子;
(b3)与(b1)或(b2)限定的DNA序列具有90%以上同源性且编码与植物雄性育性相关蛋白的DNA分子。
上述严格条件可为用0.1×SSPE(或0.1×SSC),0.1%SDS的溶液,在DNA或者RNA杂交实验中65℃下杂交并洗膜。
含有所述CsSUT1基因的重组表达载体、表达盒、转基因细胞系或重组菌也属于本发明的保护范围。
本发明还保护CsSUT1蛋白或CsSUT1基因在调控植物雄性育性中的应用。
本发明还保护CsSUT1蛋白或CsSUT1基因在植物育种中的应用。
所述植物育种是为了选育雄性不育的植物。
本发明还保护一种培育转基因植物的方法,包括如下步骤:抑制目的植物中CsSUT1基因的表达,得到雄性不育的转基因植物。
所述方法中,所述“抑制目的植物中CsSUT1基因的表达”是通过干扰载体实现的。
所述干扰载体可通过Ti质粒、Ri质粒、植物病毒载体、直接DNA转化、显微注射、电导、农杆菌介导等常规生物学方法转化到目的植物中。
所述干扰载体具体可为含有干扰片段的重组表达载体。
所述干扰片段包括区段甲和区段乙。所述区段甲和所述区段乙为反向互补序列。所述区段甲的序列如序列表的序列3所示。所述干扰片段具体可为序列表的序列4所示。
所述干扰载体具体可为在载体pFGC1008的多克隆位点中插入了序列表的序列4所示的双链DNA分子得到的重组表达载体。
所述干扰载体具体可为将载体pFGC1008的AscI和SpeI酶切位点之间的小片段取代为了序列表的序列4所示的DNA分子得到的重组表达载体。
本发明还保护一种培育雄性不育植物的方法,包括如下步骤:抑制目的植物中CsSUT1蛋白的活性和/或表达量,得到雄性不育的植物。
以上任一所述目的植物具体可为双子叶植物。所述双子叶植物可为葫芦科植物。所述葫芦科植物可为黄瓜属植物。所述黄瓜属植物具体可为黄瓜,例如新泰密刺黄瓜。
本发明还保护以上任一所述方法在植物育种中的应用。
所述植物育种是为了选育雄性不育的植物。
本发明还保护一种特异DNA分子,包括区段甲和区段乙。所述区段甲和所述区段乙为反向互补序列。所述区段甲的序列如序列表的序列3所示。所述特异DNA分子如序列表的序列4所示。
本发明还保护一种干扰载体,是将所述特异DNA分子导入表达载体得到的。
所述干扰载体具体可为在载体pFGC1008的多克隆位点中插入了序列表的序列4所示的双链DNA分子得到的重组表达载体。
所述干扰载体具体可为将载体pFGC1008的AscI和SpeI酶切位点之间的小片段取代为了序列表的序列4所示的DNA分子得到的重组表达载体。
本发明还保护所述特异DNA分子或所述干扰载体在植物育种中的应用。
所述植物育种是为了选育雄性不育的植物。
以上任一所述植物具体可为双子叶植物。所述双子叶植物可为葫芦科植物。所述葫芦科植物可为黄瓜属植物。所述黄瓜属植物具体可为黄瓜,例如新泰密刺黄瓜。
以上任一所述雄性不育具体可表现为雄花无法生长和/或雄花花药表面无花粉散出和/或雄花花粉无活力。
本发明发现了一种黄瓜蔗糖转运蛋白CsSUT1及其编码基因,下调CsSUT1基因的表达量会导致黄瓜植株雄性不育。本发明可以应用到黄瓜育种中,对提高育种效率有着积极的作用。
附图说明
图1为黄瓜不同组织部位CsSUT1基因表达量分析。
图2为干扰载体pFGC1008-RNAi-CsSUT1部分原件示意图。
图3为转基因干扰植株鉴定结果。
图4为干扰植株(RNAi)雄花生长速度观察结果。
图5为干扰植株(RNAi)花粉粒染色观察结果。
具体实施方式
以下的实施例便于更好地理解本发明,但并不限定本发明。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试验材料,如无特殊说明,均为自常规生化试剂商店购买得到的。以下实施例中的定量试验,均设置三次重复实验,结果取平均值。
新泰密刺黄瓜:参考文献:Cheng J,Wang Z,Yao F,et a1.Down-RegulatingCsHT1,a Cucumber Pollen-Specific Hexose Transporter,Inhibits PollenGermination,Tube Growth,and Seed Development.[J].Plant Physiology,2015,168(2):635-47.;公众可以从中国农业大学获得。
载体pFGC1008:参考文献:Chen CH,Yin S,Liu XW,,Liu B,Yang S,Xue SD,CaiYL,Black K,Liu HL,Dong MM,Zhang YQ,Zhao BY,Ren HZ(2016)The WD-Repeat ProteinCsTTG1 Regulates Fruit Wart Formation through Interaction with theHomeodomain-Leucine Zipper I Protein Mict.Plant Physiology 171:1156-1168;公众可以从中国农业大学获得。
农杆菌LBA4404:参考文献:Cheng J,Wang Z,Yao F,et al.Down-RegulatingCsHT1,a Cucumber Pollen-Specific Hexose Transporter,Inhibits PollenGermination,Tube Growth,and Seed Development.[J].Plant Physiology,2015,168(2):635-47.;公众可以从中国农业大学获得。
MS固体培养基:MS培养基4.43g,蔗糖30g,植物凝胶2.5g,补水至1L,调pH 5.7-5.8。
MS分化培养基:MS培养基4.43g,蔗糖30g,植物凝胶2.5g,6-BA 0.5mg,ABA 1mg,补水至1L,pH 5.7-5.8。
MS培养基:北京西美杰科技有限公司,货号:M519。
实施例1、CsSUT1蛋白及其编码基因的获得
对各品种黄瓜基因组进行序列分析、区段截取和功能验证,从新泰密刺黄瓜中发现一种蔗糖转运蛋白,将其命名为CsSUT1蛋白,如序列表的序列1所示。将编码CsSUT1蛋白的基因命名CsSUT1基因,如序列表的序列2所示。
实施例2、黄瓜不同组织部位CsSUT1基因表达量分析
分别取长至初瓜期的新泰密刺黄瓜的根(R)、茎(S)、幼叶(YL)、成熟叶(ML)、雄花(MF)、雌花(FF)和果实(F)。
提取上述材料的总RNA,并合成第一链cDNA,采用qRT-PCR的方法检测CsSUT1基因的表达情况(以Tubllin基因为内参基因),采用引物YSUT1-F和引物YSUT1-R组成的引物对检测CsSUT1基因的表达,采用引物TUB-F和引物TUB-R组成的引物对检测Tubllin基因的表达。
YSUT1-F:5’-CGTGGTTACAAAGGTTGCTGAG-3’;
YSUT1-R:5’-GCGGATACGATGAACTGTGGA-3’;
TUB-F:5’-ACGCTGTTGGTGGTGGTAC-3’;
TUB-R:5’-AGAGGGGTAAACAGTGAATC-3’。
结果如图1所示。结果表明,CsSUT1基因在黄瓜的雄花中特异性表达,其他部位较低或检测不到。
实施例3、转基因植株的获得
一、干扰载体的构建
1、提取新泰密刺黄瓜的总RNA,并反转录为cDNA,以cDNA为模板,采用CsSUT1-RNAi-1F和CsSUT1-RNAi-1R组成的引物对进行PCR扩增,回收PCR扩增产物。
CsSUT1-RNAi-1F:5’-AGGCGCGCCATCGGTTGGTTCCCATTTATCAT-3’;
CsSUT1-RNAi-1R:5’-ATTTAAATCCCCAGCACGAACACCAA-3’。
CsSUT1-RNAi-1F和CsSUT1-RNAi-1R中,下划线分别标注AscI和SwaI酶切位点。
2、用限制性内切酶AscI和SwaI双酶切步骤1的PCR扩增产物,回收酶切产物。
3、用限制性内切酶AscI和SwaI双酶切载体pFGC1008,回收约10907bp的载体骨架。
4、将步骤2的酶切产物和步骤3的载体骨架连接,得到重组载体pFGC1008-RNAi-1。根据测序结果,对重组载体pFGC1008-RNAi-1进行结构描述如下:将载体pFGC1008的AscI和SwaI酶切位点之间的小片段取代为了序列表中序列3所示的DNA分子。
5、提取新泰密刺黄瓜的总RNA,并反转录为cDNA,以cDNA为模板,采用CsSUT1-RNAi-2F和CsSUT1-RNAi-2R组成的引物对进行PCR扩增,回收PCR扩增产物。
CsSUT1-RNAi-2F:5’-GACTAGTATCGGTTGGTTCCCATTTATCAT-3’;
CsSUT1-RNAi-2R:5’-CGCGGATCCCCCCAGCACGAACACCAA-3’。
CsSUT1-RNAi-2F和CsSUT1-RNAi-2R中,下划线分别标注SpeI和BamHI酶切位点。
6、用限制性内切酶SpeI和BamHI双酶切步骤5的PCR扩增产物,回收酶切产物。
7、用限制性内切酶SpeI和BamHI双酶切步骤4得到的重组载体pFGC1008-RNAi-1,回收约11190bp的载体骨架。
8、将步骤6的酶切产物和步骤7的载体骨架连接,得到干扰载体pFGC1008-RNAi-CsSUT1。根据测序结果,对干扰载体pFGC1008-RNAi-CsSUT1进行结构描述如下:将载体pFGC1008的AscI和SpeI酶切位点之间的小片段取代为了序列表中序列4所示的DNA分子。干扰载体pFGC1008-RNAi-CsSUT1部分原件示意图如图2所示。
二、转基因干扰植株的获得
1、将步骤一得到的干扰载体pFGC1008-RNAi-CsSUT1导入农杆菌LBA4404,得到重组农杆菌。
2、取步骤1得到的重组农杆菌,接种至含有25μg/ml利福平和100μg/ml氯霉素的YEB液体培养基中,120rpm、28℃培养至菌液OD600nm达到0.6-0.8;将菌液5000rpm离心5min收集菌体沉淀,将菌体沉淀用1/2MS液体培养基洗涤2次后,重新悬浮于1/2MS液体培养基中,调整菌液OD600nm为0.2-0.3。
3、取新泰密刺黄瓜种子,播种于MS固体培养基中28℃恒温暗培养至种子萌发2天后进行后续实验。
4、取步骤3萌发2天后的黄瓜幼嫩子叶,去除生长点后将每片子叶均匀分成2块,只保留靠近生长点一半的部分。
5、将步骤4处理后的黄瓜子叶浸没在步骤2得到的农杆菌菌液中侵染15分钟,然后用灭菌的滤纸吸干多余菌液,将黄瓜子叶接种至MS分化培养基中,黑暗条件28℃培养2天后收获外植体。
6、将步骤5得到的外植体接种至含有500mg/L羧苄青霉素的MS分化培养基上,25℃、2000LX光照强度(14h光照/10h黑暗)培养,待抗性芽长至1cm时,将其切下移入含有200mg/L羧苄青霉素的MS固体培养基中诱导生根,获得T0代植株,待植株根系发育好后,移入盛有无菌土的花盆中,覆膜保温保湿,28℃、2000LX光照强度(12h光照/12h黑暗)培养,10天后除去覆膜,14天后定植于日光温室,进行常规田间管理。
7、取步骤6的T0代植株的叶片,提取总RNA并反转录为cDNA,以eDNA为模板,采用引物35S2-F和35S2-R进行PCR鉴定。
35S2-F:5’-TGGTTAGAGAGGCTTACGCAGCAGGTC-3’;
35S2-R:5’-CCATCTTTGGGACCACTGTCGGCA-3’。
将PCR产物进行电泳,如果得到特异性条带,则待测植株鉴定为阳性。
设置采用新泰密刺黄瓜(野生型)cDNA为模板的野生型对照(WT)。
设置采用水为模板的阴性对照(-)。
设置采用干扰载体pFGC1008-RNAi-CsSUT1为模板的阳性对照(+)。
鉴定结果如图3所示。
8、取步骤7鉴定为阳性的T0代植株的叶片,提取总RNA,采用实施例2中的方法检测CsSUT1基因的表达情况。
检测结果如图3所示。结果表明,RNAi 14中CsSUT1基因的表达量下降最显著。
9、将T0代RNAi 14植株自交,得到T1代植株。取若干RNAi 14株系的T1代植株,按照步骤7和步骤8的方法进行鉴定和表达量检测。
结果如图3所示。结果表明,T1代植株中,RNAi 14-1中CsSUT1基因的表达量下降最显著。
三、转空载体植株的获得
采用载体pFGC1008代替干扰载体pFGC1008-RNAi-CsSUT1,按照步骤二进行操作,得到转空载体植株。
实施例4、转基因干扰植株的表型观察
待测植株为:新泰密刺黄瓜(WT)、实施例3构建的T1代转基因干扰植株(RNAi14-1、RNAi 14-6和RNAi 14-8)和实施例3构建的T1代转空载体植株。
1、选取待测植株同一节位、库强相似、大小相同的早期雄花进行连续多天的观察。
结果如图4所示。结果表明干扰植株(RNAi)的雄花生长速度明显慢于野生型(WT),雄花发育后期甚至生长停滞,无法开放,然后变黄萎蔫凋落。转空载体植株表型与野生型(WT)相同。
2、取待测植株开花当天的雄花,去掉花瓣萼片,在体式显微镜下观察花药表面的花粉散出情况。
结果如图5A所示。结果表明,野生型(WT)花药表面有大量花粉粒,而干扰植株(RNAi)花药表面无花粉散出。转空载体植株表型与野生型(WT)相同。
3、取待测植株开花当天雄花花药,摩擦载玻片使花粉散落到载玻片上,滴一滴碘-碘化钾溶液,染色5min,然后盖上盖玻片在显微镜下观察。
结果如图5B所示。结果表明,野生型(WT)花粉大量散出且被染成蓝黑色,活力强,干扰植株(RNAi)花药摩擦后几乎无花粉散出,仅有的1-2颗花粉粒也不能被碘-碘化钾染成蓝色,没有花粉活力。说明干扰植株(RNAi)是完全雄性不育的。转空载体植株表型与野生型(WT)相同。
<110> 中国农业大学
<120> 蔗糖转运蛋白及其在调控植物雄性不育中的应用
<160> 4
<210> 1
<211> 495
<212> PRT
<213> 黄瓜(Cucumis sativus L.)
<400> 1
Met Glu His Gly Gly Val Val Ser Lys Gly Met Ala Ser Asp Pro Ser
1 5 10 15
Ser Ser Tyr Gln Lys Ile Ile Ile Val Ala Ala Ile Ala Ala Gly Val
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Gln Phe Gly Trp Ala Leu Gln Leu Ser Leu Leu Thr Pro Tyr Val Gln
35 40 45
Gln Leu Gly Val Ser His Thr Trp Ser Ala Phe Ile Trp Leu Cys Gly
50 55 60
Pro Leu Ser Gly Leu Ile Val Gln Pro Thr Val Gly Tyr Tyr Ser Asp
65 70 75 80
Arg Cys Thr Ser Arg Phe Gly Arg Arg Arg Pro Phe Ile Val Ala Gly
85 90 95
Ser Thr Phe Val Ala Thr Ala Val Phe Leu Ile Gly Phe Ala Ala Asp
100 105 110
Ile Gly His Ala Val Gly Asp Pro Leu Asn Lys Pro Thr Lys Pro Arg
115 120 125
Ala Val Ala Ile Phe Val Val Gly Phe Trp Val Leu Asp Val Ala Asn
130 135 140
Asn Met Leu Gln Gly Pro Cys Arg Ala Leu Leu Ala Asp Met Ser Cys
145 150 155 160
Asn Asn His Lys Lys Met Arg Met Ala Asn Gly Phe Phe Ser Phe Phe
165 170 175
Met Gly Val Gly Asn Val Leu Gly Tyr Ala Ala Gly Ser Tyr Asn Lys
180 185 190
Leu Tyr Lys Phe Leu Pro Phe Thr Leu Thr Lys Ala Cys Asp Ser Tyr
195 200 205
Cys Ala Asn Leu Lys Thr Cys Phe Leu Ile Asp Ile Val Phe Leu Leu
210 215 220
Leu Val Thr Thr Phe Ala Val Leu Met Val Ser Glu Asn Gln Phe Asp
225 230 235 240
Pro Leu Glu Ile Asp Glu Glu Ala Thr Pro Phe Phe Gly Lys Leu Phe
245 250 255
Gly Ala Leu Lys Lys Leu Glu Lys Pro Met Trp Leu Leu Leu Leu Val
260 265 270
Thr Ala Leu Asn Trp Ile Gly Trp Phe Pro Phe Ile Met Tyr Asp Thr
275 280 285
Asp Trp Met Gly Leu Glu Val Tyr Gly Gly Lys Pro Lys Gly Ser Pro
290 295 300
Glu Glu Val Lys Phe Tyr Asp Leu Gly Val Arg Ala Gly Ala Leu Gly
305 310 315 320
Leu Met Val Asn Ser Phe Val Leu Gly Phe Ser Ala Leu Gly Ile Glu
325 330 335
Pro Ile Ser Arg Ile Leu Gly Gly Leu Arg Trp Trp Trp Gly Ile Val
340 345 350
Asn Ile Ile Phe Thr Val Cys Met Gly Ser Thr Val Val Val Thr Lys
355 360 365
Val Ala Glu Arg Trp Arg Ser Val Asn Gly Leu Arg Pro Pro Pro Leu
370 375 380
Asn Val Arg Ala Gly Ala Phe Ser Ile Phe Ala Ile Leu Gly Ile Pro
385 390 395 400
Leu Ser Val Thr Phe Ser Val Pro Phe Ala Leu Ala Ser Ile Phe Ser
405 410 415
Ser Glu Ser Asp Ala Gly Gln Gly Leu Ser Leu Gly Ile Leu Asn Leu
420 425 430
Phe Ile Val Ile Pro Gln Phe Ile Val Ser Ala Val Ser Gly Pro Leu
435 440 445
Asp Ala Ala Phe Gly Gly Gly Asn Leu Pro Ala Phe Val Met Gly Gly
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Ile Ala Ser Phe Ala Ser Ala Met Cys Ala Met Phe Val Leu Pro Asp
465 470 475 480
Pro Pro Pro Gln Ser Asp Val Ser Leu Thr Met Gly Gly Gly His
485 490 495
<210> 2
<211> 1488
<212> DNA
<213> 黄瓜(Cucumis sativus L.)
<400> 2
atggagcatg gaggtgttgt gtcgaagggg atggcgtcag acccttcaag ttcgtaccaa 60
aaaataataa tagttgcagc tattgcagcc ggagtccaat ttggttgggc tctacagctt 120
tcattgctaa caccttacgt ccaacaactt ggagtttcac atacatggtc cgctttcata 180
tggctttgcg gaccattatc aggacttatt gtgcaaccca ctgtcgggta ctacagtgat 240
cgctgcacct ctaggtttgg tcgtcgtcgg ccgttcattg ttgctggatc tacttttgtg 300
gcaactgcag ttttcctcat tggctttgct gcagacattg ggcatgcagt gggtgatccg 360
cttaacaaac ccacaaaacc aagagctgtt gccatctttg tggttggatt ttgggttctt 420
gatgttgcta acaacatgct ccaaggccct tgtagagctc tgttggcaga tatgtcatgt 480
aacaaccaca agaagatgag aatggccaat ggatttttct cattcttcat gggggtaggg 540
aatgttttgg ggtacgcagc tgggtcctat aacaaactct acaaattcct tcccttcaca 600
ctaaccaaag catgtgacag ttactgtgcc aacctcaaaa catgcttctt gatcgacatt 660
gtattccttc tcctcgttac cacgttcgct gtgttgatgg tgagcgagaa ccaatttgac 720
ccattggaga ttgacgaaga agcaacaccc tttttcggga aattgtttgg agcactcaag 780
aagttggaga agccaatgtg gcttttgttg ctggtaacag ccttgaactg gatcggttgg 840
ttcccattta tcatgtacga tactgactgg atgggtttgg aagtgtacgg aggaaagcca 900
aaggggagtc ctgaagaagt caagttctat gaccttggtg ttcgtgctgg ggcccttggg 960
ttgatggtaa actcatttgt tttgggattt tcagctttgg gaattgagcc aataagtcgt 1020
attttaggag gcctcagatg gtggtgggga attgttaaca ttatatttac agtttgcatg 1080
ggatccaccg tcgtggttac aaaggttgct gagcgttgga ggtccgttaa tggtttgcgt 1140
cctcctccac taaacgtcag ggcaggagca ttttcgatct ttgcgatatt gggtattcca 1200
ttgtcagtca cttttagtgt tccatttgcg cttgcgtcca tcttttcttc agaatcggat 1260
gcgggtcaag gcctctcctt gggaattctc aacctcttca ttgtcattcc acagttcatc 1320
gtatccgcag ttagtgggcc attagatgct gctttcggag gaggaaactt acccgcattc 1380
gtaatgggtg gaattgcttc ttttgcaagt gcaatgtgtg caatgttcgt cctccccgat 1440
ccaccacctc aatccgatgt ctccttaaca atgggcggtg gtcattga 1488
<210> 3
<211> 121
<212> DNA
<213> 黄瓜(Cucumis sativus L.)
<400> 3
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ggaaagccaa aggggagtcc tgaagaagtc aagttctatg accttggtgt tcgtgctggg 120
g 121
<210> 4
<211> 623
<212> DNA
<213> 人工序列
<220>
<223>
<400> 4
atcggttggt tcccatttat catgtacgat actgactgga tgggtttgga agtgtacgga 60
ggaaagccaa aggggagtcc tgaagaagtc aagttctatg accttggtgt tcgtgctggg 120
gatttaaatc cccagatgaa catggcatcg tggtgattga tgaaactgct gctgtcggct 180
ttaacctctc tttaggcatt ggtttcgaag cgggcaacaa gccgaaagaa ctgtacagcg 240
aagaggcagt caacggggaa actcagcaag cgcacttaca ggcgattaaa gagctgatag 300
cgcgtgacaa aaaccaccca agcgtggtga tgtggagtat tgccaacgaa ccggataccc 360
gtccgcaagg tgcacgggaa tatttcgcgc cactggcgga agcaacgcgt aaactcgacc 420
cgacgcgtcc gatcacctgc gtcaatgtaa tgttctgcga cgctcacacc gataccatca 480
gcgatctctt tgatggggat ccccccagca cgaacaccaa ggtcatagaa cttgacttct 540
tcaggactcc cctttggctt tcctccgtac acttccaaac ccatccagtc agtatcgtac 600
atgataaatg ggaaccaacc gat 623
Claims (10)
1.一种蛋白质,是如下(a1)或(a2):
(a1)由序列表中序列1所示的氨基酸序列组成的蛋白质;
(a2)将序列表中序列1所示的氨基酸序列经过一个或几个氨基酸残基的取代和/或缺失和/或添加且与植物雄性育性相关的由序列1衍生的蛋白质。
2.编码权利要求1所述蛋白质的基因。
3.如权利要求2所述的基因,其特征在于:所述基因为如下(b1)-(b3)中任一所述的DNA分子:
(b1)编码区如序列表中序列2所示的DNA分子;
(b2)在严格条件下与(b1)限定的DNA序列杂交且编码与植物雄性育性相关蛋白的DNA分子;
(b3)与(b1)或(b2)限定的DNA序列具有90%以上同源性且编码与植物雄性育性相关蛋白的DNA分子。
4.含有权利要求2或3所述基因的重组表达载体、表达盒、转基因细胞系或重组菌。
5.权利要求1所述蛋白质,或,权利要求2或3所述基因,在调控植物雄性育性中的应用。
6.一种培育转基因植物的方法,包括如下步骤:抑制目的植物中权利要求2或3所述基因的表达,得到雄性不育的转基因植物。
7.一种培育雄性不育植物的方法,包括如下步骤:抑制目的植物中权利要求1所述蛋白质的活性和/或表达量,得到雄性不育的植物。
8.一种特异DNA分子,包括区段甲和区段乙;所述区段甲和所述区段乙为反向互补序列;所述区段甲的序列如序列表中序列3所示。
9.一种干扰载体,是含有权利要求8所述特异DNA分子的重组表达载体。
10.权利要求1所述蛋白质,或,权利要求2或3所述基因,或,权利要求6或7所述的方法,或,权利要求8所述的特异DNA分子,或,权利要求9所述的干扰载体在植物育种中的应用。
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CN114656533A (zh) * | 2020-12-22 | 2022-06-24 | 北京市农林科学院 | 西瓜新型糖转运蛋白及其编码基因ClVST1和应用 |
CN114656533B (zh) * | 2020-12-22 | 2023-05-30 | 北京市农林科学院 | 西瓜新型糖转运蛋白及其编码基因ClVST1和应用 |
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