油菜叶色基因BnaC.HO1的功能分析及其突变导致叶色变异的机理

项目名称： 油菜叶色基因BnaC.HO1的功能分析及其突变导致叶色变异的机理

项目编号： No.31501340

项目类型： 青年科学基金项目

立项/批准年度： 2016

项目学科： 食品科学、农学基础与作物学

项目作者： 祝利霞

作者单位： 华中农业大学

项目金额： 20万元

中文摘要： 叶色突变体是植物中突变频率较高且易于鉴定的突变性状，造成叶色变化的基因直接或间接影响着叶绿素代谢过程，本项目的研究材料是经EMS诱变获得油菜黄化突变体BnaC.ygl，该突变体叶绿素合成减少，叶绿体发育不良，该黄化表型是BnaC.HO1基因的缺失造成。BnaC.HO1与拟南芥中血红素加氧酶（HO1）同源，HO1参与四吡咯代谢的血红素分支，主要催化血红素b分解。我们拟以该基因为研究对象进行以下工作：1.通过qPCR分析及酶活分析解析该基因及其同源基因的表达模式及功能；2.鉴定出BnaC.HO1基因启动子区一个起主要作用的顺式作用元件，通过酵母单杂技术获得与该顺式作用元件结合的转录因子并进行功能分析；3.通过酵母双杂技术筛选与其互作的蛋白，并对互作基因进行功能解析；4. 利用转录组等分析将该基因、转录因子及其互作基因整合到四吡咯代谢及叶绿发育的调控网络。最终揭示该突变体叶色变异机理

中文关键词： 甘蓝型油菜；黄叶突变体；血红素加氧酶基因；功能分析；分子机制

英文摘要： The mutation of leaf coloration is very obvious trait mutation and commonly ocurred. The genes caused leaf color variation affected the metabolic process of chlorophyll directly or indirectly. In our study, a chlorophyll-deficient mutant with yellow-green leaves of Brassica napus was obtained by treatment with EMS. The chlorophyll-deficient mutant had decreased total Chlorophyll content, contained less granal stacks and lacked granal membranes. By a map-based cloning approach with the candidate gene approach, the leaf coloration gene BnaC.HO1 was isolated. BnaC.HO1 gene was highly homogous to the HY1 that catalysed decomposition of heme b in phytochrome chromophore synthesis-the heme branch in tetrapyrrole metabolism of Arabidopsis thaliana. In further research, we will conduct several following investigations:1. The analysis of qPCR and Enzyme activity detection will be carried out to clarify the fuctions of the BnaC.HO1 gene and its homologous gene. 2. The cis-acting elements in the promoter region will be identified by the promoter truncated method. The transcription factors will be selected through the yeast one-hybrid technology and the functions of the transcription factors will be pointed out by transformation experiments. 3.The genes that interacted with the BnaC.HO1 gene will be screened out through the yeast two hybrid technology and the functions of these genes will be pointed out through the gene expression analysis、RNAi analysis in Brassica napus and mutants analysis in Arabidopsis thaliana. In addition, BnaC.HO1 gene, the transcription factors and the genes interacted with BnaC.HO1 will be integrated into the regulatory networks of the tetrapyrrole metabolism and chloroplast development via transcriptome sequencing analysis and genetic analysis. Finally, these results will bring to light the molecular mechanisms of the leaf color variation in chlorophyll-deficient mutant BnaC.ygl.

英文关键词： Brassica napus;chlorophyll-deficient mutant;BnaC.HO1 gene;Functional Analysis;Molecular Mechanisms