草莓匍匐茎发生调控基因的定位与鉴定

The propagation of strawberry is mainly based on runners. Mechanism of produce of strawberry runners is majorly focused on physiological aspect rather than molecular level. We have obtained a mutant with runners from woodland strawberry (Fragaria vesca) which do not have runners through chemomorphosis. Genetic analysis revealed that the phenotype of produce of strawberry runners was controlled by a single recessive gene (srp). SSR and InDel markers were developed according to the re-sequencing of parental genome and srp gene was initially delimited to 2.5cM genetic distance on chromosome 4. Based on these data, we will construct a large F2 population with more than 1000 recessive plants with mutant phenotype. SSR and InDel markers and whole genome re-sequencing BSA methods will be used to perform fine-mapping of the candidate gene. In addition, we will combine transcriptome data with gene mapping data to obtain the key candidate genes regulating the produce of strawberry runners. Candidate gene clone, overexpression and RNAi vectors will be conducted to investigate the function of candidate genes. Subcellular localization and spatial and temporal expression pattern will also be analyzed in this project. The aim of this project is to clone the gene regulating the produce of strawberry runners and lay a solid foundation for uncovering the molecular mechanism of the produce of strawberry runners.

匍匐茎繁殖是草莓繁殖的主要方式，有关草莓匍匐茎发生的机理，至今主要集中在生理层面，而分子层面的机理尚不清楚。我们从不抽生匍匐茎的二倍体森林草莓（Fragaria vesca）中人工诱变出抽生匍匐茎的突变体，遗传分离比例显示该突变性状由隐性单基因（srp）控制。基于杂交亲本基因组重测序数据，我们开发出了SSR和InDel标记，并初步将srp基因定位于4号染色体2.5cM范围内。基于上述研究基础，本项目拟构建隐性个体数大于1000的F2群体，采用分子标记（SSR、InDel）与全基因组重测序BSA相结合的方法进行srp基因的定位，同时结合转录组数据，筛选调控草莓匍匐茎发生的关键候选基因。克隆候选基因并构建其过量表达载体和RNAi载体，通过草莓转基因来鉴定候选基因功能。对候选基因表达产物进行亚细胞定位并分析其表达特性。本项目旨在克隆调控草莓匍匐茎发生基因，为揭示草莓匍匐茎发生分子机理奠定基础。

匍匐茎繁殖是草莓生产上的主要繁殖方式。本项目以二倍体森林草莓中不抽生匍匐茎的种质资源和人工诱变的抽生匍匐茎突变体为试材，揭示抽生匍匐茎的突变性状由隐性单基因（srp）控制。基于传统分子标记技术和全基因组重测序BSA方法均将srp基因定位于4号染色体上。全基因组重测序获得的SNP数据显示编码DELLA蛋白的FveRGA1基因上有存在导致FveRGA1蛋白质翻译提前终止的变异。构建了草莓RGA1基因的RNAi载体并分别对二倍体森林草莓和八倍体栽培草莓进行了遗传转化，无匍匐茎森林草莓种质资源的FveRGA1基因RNAi植株均产生匍匐茎，而有匍匐茎栽培草莓‘晶玉’的 FaRGA1基因RNAi植株持续抽生匍匐茎且不开花。抽生匍匐茎突变体与野生型茎尖生长点转录组数据显示D27（独脚金内酯合成关键基因）等基因显著差异表达，而在森林草莓中过量表达D27基因显著抑制新茎分枝形成。无匍匐茎森林草莓植株喷施赤霉素（GA3）后产生匍匐茎，其机制与一些激素相关基因的差异表达有关。本项目找到调控草莓匍匐茎发生的关键基因RGA1，为深入揭示草莓匍匐茎发生分子机理奠定重要基础。项目组完成了项目研究目标，培养3名博士研究生和6名硕士研究生，获得国家发明专利1项，发表论文5篇，其中SCI收录论文2篇。