基于dTph1转座子插入的矮牵牛花发育突变体分子遗传机理解析

Petunia is a flowering herbaceous perennial plant belonging to the Solanaceae family. It not only plays an important role in beautifying the environment because of its high ornamental value, but also has important research value as a model plant for the current molecular biology studies. In this project, in order to uncover the molecular mechanisms of some important traits in petunia such as flower development, we intend to establish a mutant library by continuous self-pollination of the W138 strain which has a high copy of active dTph1 transposon, and then obtain a batch of petunia mutants. Moreover, the molecular mechanisms of two new flower mutants of petunia will be investigated. First, the mutant genes will be isolated by transposon display technology, and then the gene expression patterns and functions will be analyzed. On the other hand, the interactive proteins of the mutant genes will be investigated by yeast two-hybrid screening, and then verified through BiFC (bimolecular fluorescence complementation); the upstream regulatory factors of the genes are identified by yeast one-hybrid screening. The relationship between the isolated genes and other genes controlling floral initiation and development including some known genes such as ALF and DOT will be understood after these studies. After the project, not only can the first domestic petunia mutant library be established, which will play important roles on gene isolation and identification, but also some unknown new genes, even new genetic pathway that regulated flower initiation and development, will be discovered.

矮牵牛属于茄科多年生草本花卉，不仅具有极高的观赏价值，在美化环境中发挥着重要作用，而且逐渐成为当前分子生物学研究的模式材料，具有重要的科研价值。本项目以解析矮牵牛花发育等重要性状的形成机理为目的，拟采用具有多拷贝、高活性dTph1转座子的W138株系为材料，通过多代自交创建较大规模的转座子插入突变体库，筛选得到一批突变体；同时，对前期研究中筛选到的两个新的矮牵牛花发育突变体进行深入分析，一方面通过转座子显示技术分离其突变基因，对基因的表达模式和功能进行系统研究，另一方面通过酵母杂交技术筛选其互作蛋白和上游调控因子，并通过BiFC等方法进行验证，明确突变基因与已知花发育相关基因的调控关系及分子作用机制。项目完成后，不仅可以建立国内首个矮牵牛突变体库，得到一批富有研究价值的突变体材料，为相关基因的鉴定和利用奠定基础，而且有望揭示调控植物花发育的新基因，乃至新的调控路径，具有十分重要的科学意义

矮牵牛为茄科多年生草本植物，具有极高的观赏和科研价值。本项目以具有多拷贝、高活性dTph1 转座子的W138 株系为材料，通过多代自交创建了较大规模的转座子插入突变体库，同时筛选得到一批突变体，为解析矮牵牛株型、花朵大小、花发育等重要性状的形成机理提供了材料。同时，对前期研究中筛选得到的两个矮牵牛花发育突变体进行了研究，明确了其突变基因。另外，我们从全基因组水平，对矮牵牛miR156/157和SPL基因家族进行了研究。一方面鉴定了矮牵牛基因组中miR156/157和SPL基因家族成员，明确了PhSPLs基因的结构特点、起源及系统进化关系，以及miR156/157和PhSPLs的时空表达特点；另一方面，通过转基因等方法揭示了miR156/157以及4个矮牵牛miR156/157直接下游靶基因PhSPL2, PhSPL9a, PhSPL9b 和PhSPL9c的功能，包括在拟南芥和矮牵牛中超表PhMIR0157a前体和4个PhSPLs基因、干涉矮牵牛miR156/157和PhSPLs基因的功能。同时构建了矮牵牛花发育过程中cDNA酵母表达文库，筛选了与PhSPL9b互作的蛋白。本项目研究结果为深入解析矮牵牛花发育的分子机理奠定了良好的基础，为miR156/157和SPLs的功能进化研究增添了新的认识，并且为植物花器官大小的调控提供了新的途径。