SmTCP7a正调控茄子抗青枯病机理

TCPs（Teosinte Branched1/Cycloidea/Proliferating cell factors）transcriptional factor effect the plant defense by regulation the hormone synthesis. Our previous study suggested that the SmTCP7a positive regulate the bacterial wilt resistance of eggplant. The defense reaction may involve in salicylic acid but the mechanism is unknown. To reveal the mechanism of SmTCP7a regulation bacterial wilt resistance, the research contents are as follows: the expression pattern and biochemistry was analyzed to analysis the transcriptional activity; After the SmTCP7a was edited by CRISPR/cas9 in resistance eggplant and over-expressed in susceptible eggplant, the R. solanacearum was inoculated. The disease index, SA content and RNA-seq were analyzed to confirm the function of SmTCP7a and effection on the SA signal pathway. The CRISPR/cas9 mutant eggplant was treated by SA content, and the disease index were analyzed. The role of SmTCP7a in the upstream of the SA synthesis pathway was analyzed.The correlation analysis between Chip-seq and RNA-seq was conducted to screen the target gene which regulate the SA synthesis. Based on the above research，the SmTCP7a function and the mechanism of resistance to bacterial wilt were identified and will provide a basis for the generation of bacterial wilt resistant eggplant by genetic engineering and breeding.

TCP（Teosinte Branched1/Cycloidea/Proliferating cell factors）转录因子通过调控植物激素合成影响植物抗性。前期证明SmTCP7a正调控茄子青枯病抗性且可能与水杨酸相关，但其作用机理未知。本项目进行以下研究：对SmTCP7a进行表达模分析，确定其转录活性；超表达和沉默SmTCP7a后，接种R.solanacearum进行病情指数和水杨酸含量等分析，确定SmTCP7a正调控抗青枯病功能及其对水杨酸的影响；利用SA喷施SmTCP7a沉默植株，调查病情指数，探究SmTCP7a是否处于SA合成途径上游；利用Chip-Seq和RNA-seq数据关联分析，筛选SmTCP7a下游与水杨酸合成相关基因，解析SmTCP7a调控水杨酸合成机理。通过上述研究从SmTCP7a调控水杨酸合成的角度解析其抗病机理，为通过基因工程创制抗青枯病材料提供理论基础。

青枯病是危害茄子产业的主要病害之一，而抗病品种的选育是防控青枯病最有效的手段。筛选调控茄子抗青枯病的关键基因，并且分析其调控机理，对选育茄子抗青枯病新品种有着重要的意义。TCP（Teosinte Branched1/Cycloidea/Proliferating cell factors）转录因子是植物特有转录因子，其广泛的参与调控植物的生长发育、响应植物的生物和非生物胁迫。在我们的前期研究中，我们通过酵母双杂交筛选到一个与SmEDS1互作的蛋白，其具有典型的TCP结构域。. 在本项目中，SmTCP7a转录因子受到病原菌R.Solanacearum、水杨酸和茉莉酸的诱导，并且在抗病材料的表达量高于感病材料。在烟草叶片中进行瞬时表达，结果表明SmTCP7a定位于细胞核中，这与其转录因子的功能相符。但是其与SmEDS1互作后，其亚细胞位置由细胞和变为细胞核与细胞质共存。利用酵母双杂交和双荧光素酶分析其转录活性，结果表明SmTCP7a具有转录激活活性。在感病材料中超表达SmTCP7a和抗病材料VIGS后接种R.Solanacearum，研究结果表明SmTCP7a正调控茄子青枯病抗性。为进一步分析其抗青枯病分子机理，我们制备SmTCP7a多克隆抗体进行chip实验。Chip-seq实验结果表明，接种R.Solanacearum后，R48h 样本中鉴定到的peaks数量高于对照R0h。但是大部分的peaks位于基因间隔区域。利用Diffbind R包和位置关系分析差异peaks，获得的候选基因的大部分功能未知。RNA-seq分析结果表明oxidation-reduction、plant hormone signal transduction等植物信号通路参与了茄子调控青枯病抗性。RNA-seq和chip-seq联合分析仅仅鉴定到了1个reticulon-like基因。EMSA实验结果表明SmTCP7a能够与Smechr0300595、Smechr0103296和Smechr0702191的启动子结合。但是进一步的实验结果表明Smechr0300595、Smechr0103296和Smechr0702191基因可能未参与正调控茄子青枯病抗性。本研究探究了SmTCP7a调控茄子青枯病的功能，为通过转基因技术创制茄子抗青枯病材料提供理论基础。