蓖麻性别表现差异的分子机理

In this research, the differences of sex expression were analyzed on the DNA and transcriptome levels in castor bean. In order to understand wherther there exist the DNA mutation, insertion or modification between the pistillate line and monoecious line for castor bean, the genomic discrepancies between them were assayed by general molecular markers, retrotransposon-based molecular markers and methylation sensitive amplified polymorphis（MSAP）method; To comprehend the divergences of transcriptome expression between female line and monoecious line, pistillate flower and staminate flower, respectively, and to find some candidate genes relate to the development of pistillate/staminate flower or genes referring the sex differentiation or decision, the transcriptome sequencing of pistillate flower and staminate flower, apical leaf buds (which before change into flower bud) and squaring inflorescences of female line and monoecist, were conducted using the new, high throughput RNA-Seq technology; Base on the transcriptome analysis, to know those genes acts in the development of female/male flower or sex differentiation, the real-time expression of some important candidate genes were performed in different tissues and organs, and in situ hybridization of those genes were carried out in the racemes of different sex type, female/male flowers and apical buds of different development stage. It was worthy for future researches of sex change mechanism and sex genetic regulation, and significant for realizing artificial sex regulation, pistillate line preservation and hybrid breeding in cator bean.

本项目将从DNA和转录表达水平对蓖麻性别表现差异进行分子水平的研究分析。利用常用分子标记、反转录转座子标记和甲基化敏感多态性技术（MSAP）对蓖麻雌性系和其相应的两性系进行基因组水平上的差异分析，明确其是否存在DNA水平上的突变、插入或修饰；利用新一代高通量转录组测序技术（RNA-Seq）对蓖麻雌花和雄花、雌性系和两性系的花芽分化前顶芽及刚现蕾花序进行转录组测序分析，明确雌花与雄花、雌性系与两性系之间的转录表达差异，寻找雌、雄花发育相关基因或性别分化、决定相关基因；在此基础上，对重要相关基因在不同组织器官中的表达进行定量分析，并对其在不同时期顶芽、雌花和雄花以及不同性别花序中的表达情况进行原位杂交表达分析，明确其在雌、雄花发育和性别分化中的作用。本项目研究可为深入研究蓖麻性别变化机理及遗传调控奠定基础，同时对实现人工性别调控、雌性材料的保持及杂交育种具有重要意义。

蓖麻（Ricinus communist L.）属重要工业油源作物，是世界十大油料作物之一。蓖麻产量、不育系（单雌材料）的保持和杂交种选育都与蓖麻性别表现密切相关，蓖麻为大戟科雌雄异花同株植物，性别表现复杂多样，加强对蓖麻性别表现分子水平的研究，了解蓖麻性别表现的分子机制，对深入认识植物性别调控及生殖器官发育等具有基础理论研究价值，同时对蓖麻增产、性别调控及杂种优势利用等具有生产实践应用价值。本研究对不同来源的单雌材料进行了雌株率的调查分析；利用SSR及MSAP甲基化标记对蓖麻雌性系“JXBM0705P”和两性系“JXBM0705M”进行了DNA水平上的差异分析，但并未找到与性状相关的差异片段。并对蓖麻雌性系和两性系花芽分化前的顶芽和刚现蕾花序，以及雌、雄花进行转录组测序及差异表达基因的分析与验证；同时对蓖麻雌性系和两性系三个不同时期的顶芽进行数字表达谱分析、差异基因聚类分析及表达验证，并对雌性系和两性系三个不同时期的激素含量变化进行测定，明确了顶芽不同发育时期的转录表达水平差异及激素含量变化，通过对这些差异基因的功能注释及KEGG分析，找到了激素应答、合成和运输，转录因子、信号传导，授粉、花粉管生长、性别分化等与性别发育相关的基因，为明确蓖麻性别表现机制、深入研究蓖麻性别变化及遗传调控奠定基础。