甘蓝型油菜菘油CMS系的线粒体基因组结构及不育机制

Novel cytoplasmic male sterile (CMS) lines of rapeseed can be produced by replacing the nucleus through interspecific hybridization, or mitochondrial recombination resulting from somatic cell fusion. By recurrent backcrossing the somatic hybrid between Brassica napus cv. Huashuang 3 and Isatis indigotica with parental B. napus, new inap CMS line which expressed carpelloid stamens was obtained. Previous analysis revealed that the line contained the recombinant mitochondrial genomic compositions with both parental mtDNAs. In this project, the mitochondrial genomes of inap CMS line will be sequenced to reveal the parental origin of mtDNA for the new line and predict the CMS-associated orf. Gene expressions in the CMS and maintainer should be studied by RNA-seq analysis at different stages of flower bud development to elucidate the molecular mechanism of carpelloid stamens and to explore how the nuclear-encoded flower genes were regulated by mitochondrial retrograde signals.

通过种间核置换或体细胞融合实现的线粒体基因重组可产生新型的油菜细胞质雄性不育(CMS)系统。我们前期通过甘蓝型油菜和菘蓝的族间体细胞杂种与甘蓝型油菜多代回交，获得雄蕊心皮化的“菘油”雄性不育系（inap CMS），初步结果表明双亲的线粒体基因组发生了重组。本项目拟进行inap CMS的线粒体基因组测序、揭示不育系的重组线粒体成分、预测不育相关orf；转录组测序分析CMS和保持系的花蕾在不同发育阶段的基因表达，阐明雄蕊心皮化发生的遗传机制，探讨线粒体反向调控核编码的花发育基因的表达，为该新不育系的利用奠定基础。

细胞质雄性不育是油菜杂种优势利用的主要途径之一，通过远缘杂交进行种间核置换可产生新的细胞质雄性不育。我们通过体细胞杂交、系统选育将药用植物菘蓝的异源胞质导入甘蓝型油菜，培育新的甘蓝型油菜“菘油”不育系。“菘油”CMS败育始于雄蕊原基分化时期，雄蕊发生心皮化，不育彻底、稳定。本项目通过线粒体基因组测序，揭示了菘油CMS的线粒体组成，由菘蓝和甘蓝型油菜重组而来，大部分是菘蓝mtDNA序列；预测并克隆了不育基因orf291，其位于rps7的上游，与之共转录。ORF291在不育系中花蕾中积累，但在恢复系中表达量降低。不育基因orf291源自菘蓝，由短片段的重复序列介导的mtDNA重组而产生的。结合转录组和蛋白组测序，在不育系中很多早期涉及雄蕊发育基因都下调表达，如 AP3、PI、SPL基因等；涉及能量代谢和线粒体结构蛋白也受到影响，导致线粒体结构和功能异常。本研究为甘蓝型油菜“菘油”CMS不育机理解析和油菜杂交种生产提供科学基础。