【作者】 姜伟；

【导师】 杨守萍；

【作者基本信息】 南京农业大学 ， 遗传学， 2010， 硕士

【摘要】 质核互作雄性不育在作物杂种优势利用中起着重要的作用,研究其遗传基础和机制具有重要的理论价值和实践意义。为揭示大豆质核互作雄性不育的分子机理,本研究开展了大豆质核互作雄性不育系与保持系基因差异表达分析及atp9基因RNA编辑研究,获得如下结果：1. cDNA-AFLP分析获得大量差异片段,包括特异表达片段和高表达片段；对特异表达片段TDFs进行分析,结果显示在不育系NJCMS1A和保持系NJCMS1B中特异表达片段TDFs主要集中出现在中花蕾期和大花蕾期,而在不育系NJCMS2A和保持系NJCMS2B中特异表达片段TDFs主要集中出现在小花蕾期和中花蕾期；对回收测序的55条特异表达片段TDFs进行分析,发现它们的功能主要涉及信号转导、电子传递和能量代谢、细胞壁合成与调控、蛋白质代谢、防卫和胁迫反应等；对其中12个特异表达片段TDFs进行荧光定量分析,结果显示它们的表达存在时空差异性。2.从保持系NJCMS1B中分离得到一个仅在大花蕾期特异表达的片段TDFs25,经网上预测该基因为DUF620, cDNA全长1308bp,编码436个氨基酸,是一个未知功能蛋白；半定量和荧光定量分析结果表明DUF620基因在不育系NJCMS1A和NJCMS2A的大花蕾时期表达量均极显著低于其相应保持系NJCMS1B和NJCMS2B,推测该基因可能与大豆质核互作雄性不育相关。3.序列分析显示以不育系NJCMS1A和保持系NJCMS1B的cDNA为模板扩增的atp9基因的序列一致,而以不育系NJCMS2A和保持系NJCMS2B的cDNA为模板扩增的ap9基因的序列存在差异；蛋白质跨膜结构预测结果显示不育系NJCMS2A和保持系NJCMS2B中ap9基因编码的蛋白的跨膜结构存在较大差异。至于ap9基因的RNA编辑与大豆质核互作雄性不育的关系有待进一步研究。更多还原

【Abstract】 Cytoplasmic-nuclear male sterility (CMS) plays an important role in the utilization of crop heterosis. It is of important significance on the theory and practice to study the genetic base and mechanism of CMS. To reveal the molecular mechanism of soybean cytoplasmic-nuclear male sterility, the gene differential expression and RNA editing of atp9 gene between soybean cytoplasmic-nuclear male sterile lines and their maintainer lines were studied in the present paper. The main results were as follows:1. A large number of differential expression bands including special expression bands(TDFs) and high expression bands between soybean cytoplasmic-nuclear male sterile lines NJCMS1A, NJCMS2A and their corresponding maintainer lines NJCMS1B, NJCMS2B were obtained by the cDNA-AFLP technology. The results of TDFs analysis showed that the TDFs from NJCMS1A and NJCMS1B were mainly appeared in the middle and large flower buds, while those from NJCMS2A and NJCMS2B mainly in the small and middle flower buds. Total 55 TDFs were recycled and sequenced. The BLAST analysis showed that the function of the proteins encoded by these 55 TDFs were mainly related to the signal transduction, the electron transport and energy metabolism, the cell wall synthesis and regulation, the protein metabolism, the defense and stress responses. QRT-PCR results showed that the expression of 12 TDFs had spatial and temporal differences.2. A special expression fragment TDFs25 was isolated from the large flower buds of NJCMS1B by cDNA-AFLP. The blast results showed that TDFs25 was high homology with the conservative region of the DUF620 gene, which was an unknown functional protein. The full-length cDNA of DUF620 was 1308bp and encoded 436 amino acids. The semi-quantitative and quantitative PCR results showed that the DUF620 gene expression in the large flower buds of NJCMSIA and NJCMS2A were significantly lower than those of NJCMS1B and NJCMS2B. It was inferred that DUF620 gene might be related to soybean cytoplasmic-nuclear male sterility. 3. The sequence analysis showed that the cDNA sequence of atp9 from NJCMS1A was the same as that from NJCMS1B, while the cDNA sequence of atp9 from NJCMS2A was different from that of NJCMS2B. The protein trans-membrane structure prediction showed that the atp9 protein structure of NJCMS2A was obviously different from that of NJCMS2B. The relation between atp9 and soybean cytoplasmic-nuclear male sterility needed to be further studied.更多还原