‘红高’葡萄VvMYBA1启动子区转座子结构序列形成及其调控果皮着色机理研究

芽变育种是葡萄（果树）育种的重要方式之一。葡萄红色品种‘红高’由绿色‘意大利’芽变经选育而成。申请人前期发现‘红高’VvMYBA1启动子区插入了一段VvMYBA3启动子序列后形成了一个转座子结构序列，使其在该位点多了2个ABA响应元件，而‘意大利’没有。该转座子结构的形成原因与VvMYBA1及启动子区调控着色的机理尚不清楚。本项目拟做以下研究：（1）分析‘意大利’及其亲本、子代等品种的VvMYBA1编码区和启动子序列，明确该转座子结构序列的形成原因；（2）通过‘红高’VvMYBA1启动子截断功能验证试验，明确调控着色功能的关键位点；通过酵母单（双）杂交等技术，验证调控该过程的上游转录因子及其互作蛋白；用ABA处理具有调控着色功能的关键位点转基因番茄果实，明确VvMYBA1启动子区对ABA的响应。研究结果将为完善葡萄果皮色泽形成机理提供理论基础也为人工调控果实外观品质及色泽育种提供技术支撑。

The use of bud sports is one of the important methods in grape (fruit)breeding. The red grape 'Benitaka' was derived from a bud sport of the green 'Italia'. The applicant's previous study found that the promoter region of 'Benitaka' VvMYBA1 inserted a VvMYBA3 promoter sequence to form a transposon-like structure, resulting in the increase of two ABA response elements in 'Benitaka' at this site but not in 'Italia'. Why this transposon-like structure is formed and how VvMYBA1 and its promoter region to regulate color are still unclear. Based on previous findings, this project plans to do the following: (1) By analyzing the coding region and promoter sequence of 'Italia' and its parents, progeny and cultivars with the same genetic background, to further clarify the formation of this transposon-like structure; (2) By verifying function of the 'Benitaka' VvMYBA1 promoter truncation, to confirm the key functional sites that regulate the skin coloration; through yeast one-hybrid system, transcriptome sequencing, yeast two-hybrid system, and other technologies to verify the key upstream transcription factors and proteins that may interact with these factors; to obtain transgenic tomato plants that carry VvMYBA1 variants with key functional sites in the promoter region, and treated tomato fruits with ABA to clarify the response of the VvMYBA1 promoter region to ABA application. The outcomes of this study are expected to provide a theoretical foundations for the improved understanding of grape skin color formation and provide technical support for artificially regulating the appearance of fruit and breeding for fruit cultivar with well coloration.