基于连锁作图及转录组分析的山葡萄性别分化相关基因筛选

The dioecious Vitis amurensis is a distinctive fruit tree species native to China. The sex differentiation study of Vitis amurensis is of great importance in gene exploring and molecular assisted breeding. In this project, we plan to launch research in two strategies——linkage mapping and transcriptom analysis. Firstly, a mapping population with separating sexes will be used to construct genetic maps containing the sex locus. SSR markers and sex (as a morphological marker) will be applied for map construction. Meanwhile, based on the successful sex conversion, we use male flower (converted vs non-converted) for transcriptome high-throughput sequencing, aiming to obtain differential expression genes. Integrating the sex location information and transcriptom results, sex-linked expression genes could be defined as candidate genes closely related to sex differentiation. Then with single plants of different sexes, candidate genes will be verified by semi-quantitative RT-PCR technique. We’ll also analyze the tissue specificity and spatio-temporal expression profile of the candidate genes. Finally, genes closely related to sex differentiation will be defined. This study is beneficial for uncovering the sex differentiation mechanism in Vitis amurensis, and is also of great significance in marker assisted selection in grapevine breeding in the future.

山葡萄是原产我国的特色果树，为雌雄异株，研究其性别分化对深度挖掘其优良基因及山葡萄分子辅助育种具有重要意义。本项目拟采用连锁作图与转录组分析两种策略开展研究，首先以雌雄花型分离的杂交群体为试材，采用SSR分子标记结合性别表型这一形态学标记，构建含有性别位点的遗传图谱；同时在成功诱导雄株性别转换的基础上，以转换前后的雄花为试材，进行转录组高通量测序，得到二者间的差异表达基因。综合性别位点的定位信息及转录组分析结果，获得与性别位点紧密连锁的差异表达基因——即性别分化候选基因。再采用半定量RT-PCR及RT-qPCR等技术对候选基因进行单株验证，及组织特异性分析和时空表达分析，最终明确与山葡萄性别分化密切相关的基因，研究结果不仅为后期山葡萄性别分化机理研究奠定基础，而且对今后葡萄育种中的标记辅助选择具有重要意义。

山葡萄是原产我国的特色果树，为雌雄异株，研究其性别分化对深度挖掘其优良基因及山葡萄分子辅助育种具有重要意义。本项目以花型分离的山葡萄杂交群体为试材，集群分离分析策略结合基因组重测序技术，实现了山葡萄性别连锁群构建及性别位点的精细定位，将性别位点定位在了物理距离为30kb的两个分子标记之间，该研究结果对今后深入挖掘山葡萄性别决定基因奠定了坚实的基础，而且对葡萄育种中的标记辅助选择具有重要的现实意义。以山葡萄雄株性别转换前后的花蕾为试材开展了细胞学及转录组学研究。细胞学研究表明山葡萄雄株雌蕊的败育可能是四核胚囊形成之前珠心细胞的程序性死亡所导致，该研究结果不仅为山葡萄性别转换机理提供了较直观的细胞学依据，而且对今后其他葡萄属植物性别决定相关研究奠定了细胞学基础。通过转录组测序筛选得到了与山葡萄性别分化相关的基因，这些基因大多与植物激素信号转导相关，我们采用半定量及定量PCR技术明确了部分关键基因的表达模式，该研究结果为今后系统地开展与山葡萄性别分化相关的RNA组学、蛋白质组学乃至多组学联合分析奠定了基础。