玉米CMS-C核恢复基因Rf4的克隆及育性恢复机理的解析

项目名称： 玉米CMS-C核恢复基因Rf4的克隆及育性恢复机理的解析

项目编号： No.31471504

项目类型： 面上项目

立项/批准年度： 2015

项目学科： 食品科学、农学基础与作物学

项目作者： 薛亚东

作者单位： 河南农业大学

项目金额： 88万元

中文摘要： 细胞质雄性不育（CMS）是作物杂种优势利用和开展核-质互作分子生物学基础研究的重要性状。课题组前期利用图位克隆及关联分析策略，将玉米CMS-C育性恢复基因Rf4的候选基因Rf4_T03精细定位在Chr8短臂60Kb的区段内，并完成了其DNA及cDNA全长序列的克隆。生物信息学分析发现Rf4_T03编码的蛋白不含PPR模体，无靶向线粒体的信号肽，表明Rf4可能是一种新类型的CMS恢复基因。基于此，本研究拟在获得带有自身启动子的Rf4_T03正义链和35S驱动的反义链转基因植株的基础上，开展表型鉴定、Northern Blotting、原位杂交、免疫组化分析、花药转录组分析，Y2H筛选与Rf4_P03的互作蛋白等研究，以期阐明Rf4基因表达的调控网络及育性恢复的分子机制。研究结果不仅可为完善植物CMS育性机理提供理论依据，并将为玉米CMS-C不育化制种技术的应用提供理论支撑。

中文关键词： 玉米；细胞质雄性不育；恢复基因；转录组；调控网络

英文摘要： Cytoplasmic male sterility in plants is an important trait for its application to crop breeding and heterosis and for fundamental research of crosstalk between nucleus and mitochondrion. A candidate gene for restorer of fertility in CMS-C maize was conditioned into a 60k nt region in the short arm of chromosome 8 by map-based cloning combined with association analysis. Bioinformatic analysis showed that RF4 lacks signal peptides for mitochondrial localization and doesn't contain any PPR motif, which indicates that RF4 has a different mechanism of fertility restoration from those proteins with PPR motifs. Based on previous results, a new construct driven by the promoter of Rf4 gene itself and a RNAi construct will be developed and introduced into maize plant for further confirming the restoring ability of Rf4 gene. The expression pattern will be analyzed by northern blot, in situ hybridization; the cytological localizations by immunohistochemistry. Yeast two-hybrid will be applied to screen the potential protein(s) interacting with RF4. Combining the results from transcriptome analysis, the regulation network of Rf4 gene will be finally identified. The functional investigation of Rf4 gene will bring new insights into the biology of CMS and boost the application of maize CMS-C in its hybrid production.

英文关键词： maize;CMS;restoring gene;transcriptome;regulation network