茶树中EGCG-O-甲基转移酶基因的转录调控机制研究

项目名称： 茶树中EGCG-O-甲基转移酶基因的转录调控机制研究

项目编号： No.31470692

项目类型： 面上项目

立项/批准年度： 2015

项目学科： 农业科学

项目作者： 黄建安

作者单位： 湖南农业大学

项目金额： 88万元

中文摘要： 甲基化儿茶素是茶树重要的次生代谢产物，儿茶素-O-甲基转移酶(EGCG-O-甲基转移酶, ECG-O-甲基转移酶, EGC-O-甲基转移酶)是各种甲基化儿茶素合成中的关键酶。由于富含甲基化儿茶素的茶树资源稀少，甲基化儿茶素的生物合成关键酶基因的功能及其转录调控机理尚未明晰。本项目拟以前期研究获得的富含甲基化EGCG的茶树资源为材料，研究EGCG-O-甲基转移酶(CsOMT)在基因、蛋白和酶活性水平对甲基化EGCG合成的影响；进一步克隆CsOMT基因的启动子，采用酵母单杂筛库、凝胶阻滞分析和体内瞬时表达等DNA-蛋白质互作技术筛选、鉴定调控CsOMT基因的关键转录因子；同时挑选3-4个关键转录因子在茶树中进行RNA干扰，明确其在甲基化EGCG生物合成的具体功能，最终阐明甲基化EGCG生物合成关键酶基因CsOMT的转录调控机制，为高甲基化EGCG的茶树种质资源创新提供理论依据。

中文关键词： 茶树；EGCG-O-甲基转移酶；转录调控

英文摘要： O-methylated catechins are the most important secondary metabolites in tea plant (Camellia sinensis). Catechins-O-methyltransferase (EGCG- O-methyltransferase, ECG- O-methyltransferase, EGC- O-methyltransferase) is the key enzyme to synthesize O-methylated catechins. The function and transcriptional regulation mechanism of Catechins-O- methyltransferase is largely unknown due to the scarce tea resources with high O-methylated catechin content. The effects of EGCG-O-methyltransferase (CsOMT) in gene, protein and enzyme levels on the synthesis O-methylated EGCG were studied using different cultivar leaves with high O-methylated EGCG content. The promoter sequence of CsOMT gene was cloned by genome walking. Transcription factors that regulate CsOMT gene were obtained by Yeast one hybrid library screening, and further identified by electrophoretic mobility shift and effector-reporter assays. Three or four key transcription factors were selected and their functions were verified by RNA interference in tea plant. These results will elucidate the transcriptional regulation mechanism of CsOMT which provide theoretical support for innovation of tea cultivar resources with high O-methylated EGCG content.

英文关键词： tea plant;EGCG-O-methyltransferase;transcriptional regulation