基于启动子序列变异和DNA甲基化分析的红梨花青苷合成相关基因转录调控机制研究

项目名称： 基于启动子序列变异和DNA甲基化分析的红梨花青苷合成相关基因转录调控机制研究

项目编号： No.31272141

项目类型： 面上项目

立项/批准年度： 2013

项目学科： 农业科学

项目作者： 滕元文

作者单位： 浙江大学

项目金额： 80万元

中文摘要： 红梨着色模式多样，现有红梨着色机制的研究多集中在花青苷合成相关基因克隆，表达分析和功能鉴定的传统思路上，对这些基因转录调控机制研究极为薄弱，所涉及的研究材料也相对单一，难以从全局上认识不同着色类型红梨着色的分子调控机制。本研究从启动子序列变异和DNA甲基化分析两个角度系统研究红梨花青苷合成相关基因转录调控机制，拟以包括东方梨、西方梨和特殊类型等具有不同着色模式的典型红与非红梨为试材，分离其花青苷合成相关基因编码区和启动子，并对这些基因序列变异与色泽变异进行关联分析，找出与色泽变异相关的序列变异，再利用双荧光素酶检测、启动子删截、酵母双杂交、凝胶阻滞分析和转基因等实验手段对该序列进行功能鉴定分析，对于无法从基因序列变异角度解释的色泽变异现象，则对其进行DNA甲基化水平检测。研究成果有助于阐明红梨花青苷合成调控机制，同时可进一步丰富果实花青苷生物合成的分子机制的理论。

中文关键词： 红梨；花青苷；着色调控；DNA甲基化；miRNA

英文摘要： Red pears belonging to different species with different genetic backgrounds and coloration patterns may have the distinct transcriptional regulation for related anthocyanin biosynthesis genes compared with apples and grape. However, more attention was payed on the isolation, expression and function analysis of related genes in the study on molecular mechanism of coloration in red pears, few have carried out on the mechanism of transcriptional regulation. What's more, the limited and single material in the previous studies related to red pears would not be favorable to the comprehensive understanding of molecular regulation mechanism of different kinds of red pears. Therefore, in this project, typical red pears including Oriental pear, Occidental pear and some special types with different coloration patterns will be used to study transcriptional regulation for related anthocyanin biosynthesis genes. The promoter region will be isolated for analyzing the correlation between the sequence variations in the promoter region and peel phenotypes of red pears. The dual luciferase assay, promoter deletion analysis, electrophoretic mobility shift assay, yeast two-hybrid assay, transgenic technology will be further used for the analysis of their function. Methylation levels of these genes will also be investigated to unders

英文关键词： Red pear；Anthocyanin；Coloration regulation；DNA methylation；miRNA