【作者】 钟欢；

【导师】 刘曙东；

【作者基本信息】 西北农林科技大学 ， 作物遗传育种， 2012， 硕士

【摘要】 小麦雄性不育的研究对于杂种优势的利用具有重要意义，本研究以TR1376A不育系为材料，对该不育材料的育性遗传、花粉败育以及花粉显微镜结构进行研究，并采用了SSR和AFLP两种分子标记对转基因雄性不育基因进行定位，构建转P_SAG12-IPT基因小麦不育系育性基因的遗传连锁图谱，进一步探索小麦雄性不育的遗传机理。结果表明：1.用TR1376A不育株与9个正常可育品种杂交F1都分离出1/2的可育株和1/2的不育株，经卡方检验符合1：1的比例，表明其不育性受一对显性核不育基因控制，所有不育株都是杂合基因型。2.采用TTC和碘-碘化钾染色（IKI）两种染色方法检验花粉活性，同可育花粉相比，不育花粉染色较浅，且在同一放大倍数视野下，不育植株的产生的花粉数量明显少于可育植株。由此可知，不育植株同可育植株相比，花粉量较少且花粉大多数发育不良或者败育。3.采用1049对SSR引物对育性池进行初步筛选，其中10对引物在育性池间存在差异，它们分别是：Xbarc129、Xbarc232、Xbarc380、Xgdm98、Xcfd13、Xcfd29、Xcfd46、Xcfd76、Xcfd188、Xwmc28。后经含有170个单株的大群体对以上多态性引物进行验证，其中3对SSR引物多态性稳定，它们分别是： Xgdm98、Xcfd188和Xcdf76，该3对SSR引物均位于小麦6D染色体上。4.采用AFLP反应体系和程序，由BSA法建立的育性池筛选已合成的512对AFLP引物组合，经初步筛选，30对引物组合在育性池间存在差异，后经小群体和170个单株大群体检验后，其中3对引物组合存在较稳定的多态性，它们分别是：P-GCA/M-GAC、P-GTT/M-GTG、P-TGT/M-GGG，但引物组合P-GCA/M-GAC和P-TGT/M-GGG同目的基因之间的距离大于40cM，只有P-GTT/M-GTG引物组合同目的基因之间连锁紧密。5.用MapDraw V2.1绘制遗传连锁图谱，可知微卫星位点Xgdm98、Xcfd188和Xcdf76与目的基因之间的遗传距离分别为19.2cM、7.7cM、9.5cM，且分布在目的基因的两侧，AFLP标记位于目的基因和SSR标记之间，同目的基因之间的遗传距离为4.7cM。4个标记的发掘，为分离克隆小麦不育基因奠定了基础。更多还原

【Abstract】 The study on male sterility of wheat plays an important role on utilization of heterosis. Inorder to further explore wheat genetic mechanism of male sterility, iIn this paper, we studiedthe male sterile plant TR1376A from following aspects: fertility genetic, pollen abortion andpollen microscope structures, and molecular marker technology. This study used SSR andAFLP techniques to map the male sterility gene and construct the genetic linkage map ofmale-sterility wheat TR1376A. The results showed that:1. F1hybrids of male sterile plant TR1376A and9male male fertile varieties gave1:1segregation of male fertility to sterility, indicating that the male sterility of TR1376A washeritable and controlled by a single dominant gene, all infertility is heterozygous plantgenotype.2. The pollen activity was investigated by2,3,5-triphenyltetrazolium chloride (TTC)and IKI respectively. Compared with fertile pollen, infertility pollen was dyed less shallowly.Under the same magnification field view of microscope, the number of infertility pollen isless, which contrasted with fertile poolen.The data indicated that infertility pollen number wasfewer, and most of them were pooly developed or aborted.3. Using fertile and sterile DNA bulks to screen1049SSR primers, the result showed thatthere existed polymorphism in10primers. They were Xbarc129, Xbarc232, Xbarc380,Xgdm98, Xcfd13, Xcfd29, Xcfd46, Xcfd76, Xcfd188and Xwmc28. Subsequently, using apopulation of170individuals to verify the primers that showed polymorphism,3SSR primers,Xgdm98, Xcfd188and Xcdf76showed stable polymorphism and all located in chromosome6D of wheat.4. Using fertile and sterile DNA bulks to screen512AFLP primer combinations, theresult showed that there existed polymorphism in30primers. Then, a small population andusing a population of170individuals to verify the primers that showed polymorphism,3primers, P-GCA/M-GAC, P-GTT/M-GTG, P-TGT/M-GGG，showed stable polymorphism.But the genetic distance between primer combinations P-GCA/M-GAC, P-TGT/M-GGG andthe target gene were larger than40cM, so that only primer combination P-GTT/M-GTG was closely linked to target gene.5. Using MapDraw V2.1drew genetic linkage map, the genetic distance betweenmicrosatellite locis, Xgdm98, Xcfd188, Xcfd76and target gene were19.2cM,7.7cM,9.5cMrespectively. They were distributed on both sides of target gene. AFLP markers locatedbetween SSR markers and target gene, the distance between AFLP markers and target genewas4.7cM. The identification of the four markers will lay the foundation for the isolationand cloning sterile gene of wheat.更多还原