烟草细胞质雄性不育系及其保持系的花蕾差异蛋白质分析

摘要

为探讨烟草雄性不育的分子遗传机制,对烟草细胞质雄性不育系和保持系进行差异蛋白质组学研究.采用固相pH梯度SDS-PAGE双向电泳,对烟草细胞质雄性不育系MSK326及其保持系K326雄蕊原基分化期、四分体时期及单核时期花蕾蛋白质进行分离,经考马斯亮蓝染色,获得了分辨率和重复性较好的双向电泳图谱.使用PDQuest软件分析蛋白质图谱,利用基质辅助激光解吸电离飞行时间串联质谱分析,然后用Mascot软件对NCBInr数据库搜索.在分子量14.4～97.6 kD、等电点4～7线性范周内,检测到365个蛋白点,其中7个蛋白质点在保持系中出现,在不育系中缺失,被鉴定为核酮糖-1,5-二磷酸羧化酶/力氧酶、多酚氧化酶、Patatin homolog蛋白、PSI 9 kD蛋白4类蛋白,推测不育系MSK326雄性不育性可能与营养代谢紊乱、间接抑制雄性器官发育、养分供给不足、免疫能力低和能量转移受到抑制等有关.%To explore the molecular genetic mechanisms of cytoplasmic male sterility (CMS) in tobacco, we studied the differential proteins between the CMS line and its maintainer. Immobilized pH gradient two-dimensional gel electrophoresis (2-DE) technique was used to separate the protein spots while gels were stained with the Coomassie Blue G-250. The difference between protein maps of bud from cytoplasmic male-sterile line MSK326 and maintainer line K326 was analyzed with PDQuest image software. Matrix-assisted laser-adsorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) technique was used to obtain the peptide mass of differentially expressed protein spots. The Mascot software was used to search the protein database NCBlnr to identify those spots interested. A total of 365 protein spots were detected within Mr 14.4-97.6 kD and pH 4-7. Seven protein spots appeared in the protein map of maintainer line K326 but absent in that of CMS line MSK326. Which were identified as ribulose-l,5-bisphosphate carboxylase/oxygenase, polyphenol oxidase, patatin homolog, and PSI 9 kD protein. It was inferred that the male sterility of MSK326 might be related to energy metabolism turbulence, abnormality in male organ development, deficient nutrition supply and immunity, and inhibition of energy transfer.