MADS-box家族转录因子SlEBI对番茄花序分化发育的分子调控机理研究

Plant reproductive success and agricultural productivity are highly determined by the variable branch and flower number of inflorescence, however molecular mechanisms underlying inflorescence development are still poorly understood. The wild type tomato AC++ produces usually 5–8, most up to 20, opened flowers per inflorescence. We cloned a novel inflorescence development-related MADS-box gene SlEBI from tomato. Silencing of SlEBI resulted in extreme branched inflorescences with hundreds of flowers in tomato. Inflorescence-related genes ANANTHA (AN) and PISTILLATE (PI) were dramatically down-regulated in SlEBI-silenced lines, While FALSIFLORA (FA) was slightly down-regulated and SINGLE FLOWER TRUSS (SFT) was up-regulated in SlEBI-silenced lines. In this study, we'll construct the vector of SlEBI overexpression, transform wild type tomato AC++ and get overexpressed transgenic plants. Then the phenotypes related to inflorescence development will be analyzed to illuminate the function of SlEBI. The new inflorescence-related genes which are regulated by SlEBI will be cloned with RNA-Seq and yeast two-hybrid system to illuminate the mechanism of regulation of SlEBI on the inflorescence development in tomato. Set up the regulatory networks of the inflorescence development in tomato by crossing SlEBI-silenced transgenic plants with other tomato inflorescence mutants. The research work will shed more light on molecular mechanism in the development regulation of both definite inflorescence and indefinite inflorescence.

花序结构和花的数目是影响农作物单产的直接因素之一。花序可分为无限花序和有限花序。目前，对无限花序和有限花序发育差异及花序结构多样化的分子调控机理仍然缺乏了解。番茄每一花序的花数一般为5-8朵，多的20余朵。在前期研究中，克隆到一个调控番茄花序发育的关键基因SlEBI，沉默该基因导致番茄花序极度分支，每台花序着花数可多达350朵。研究表明SlEBI可调控AN、SFT、FA等花序发育相关基因的表达，是花序发育中较早的调控因子。本项目拟培育SlEBI超表达番茄株系，结合已获得的沉默株系，深入研究SlEBI基因对番茄花序发育的影响，明确该基因的生物学功能，阐明番茄SlEBI调控花序分化发育的作用机理；并将SlEBI的转基因材料与其他番茄花序发育突变体进行杂交，分析SlEBI与其他花序发育调控因子的上下游关系，建立番茄花序分化发育的调控网络。本项目对阐明花序发育的分子调控机制具有十分重要的理论意义。

花序结构和花的数目是影响农作物单产的直接因素之一。番茄每一花序的花数一般为5-8朵，多的20余朵，但关于番茄花序发育调控机制并不清楚。我们克隆到一个调控番茄花序发育的关键基因SlEBI，通过构建SlEBI单沉默及SlEBI-SlMADS1双沉默载体，遗传转化番茄子叶外植体，获得转基因番茄株系，发现SlEBI-SlMADS1双沉默株系的花序产生的花朵数量明显多于SlEBI单沉默株系，而SlEBI单沉默株系的花序产生的花朵数量明显多于野生型，经花序材料转录组测序分析，实时荧光定量PCR验证，扫描电镜分析，酵母双杂交分析，结果表明SlEBI与SlMADS1协同调控了番茄花序发育。此外，SlEBI-SlMADS1双沉默株系花的萼片明显比SlEBI单沉默株系大，SlEBI单沉默株系花萼又明显大于野生型，经石蜡切片，萼片发育相关基因的定量PCR检测，酵母双杂交分析，发现SlEBI与SlMADS1协同调控萼片发育。同时，我们构建了SlEBI超表达载体并获得靶基因超表达的番茄株系，发现SlEBI超表达植株叶片卷曲和小叶减少，花器官和花序结构生长缺陷，使果实产量下降，种子变小且不能正常萌发。经叶片极性调控及种子发育相关基因的表达检测以及生理生化检测，结果表明SlEBI对叶片极性发育和生殖发育有重要调控作用。该项目揭示了SlEBI的生理功能，为高产番茄新品种培育提供了理论基础。