两个WD40转录因子对银杏类黄酮生物合成调控的研究

项目名称： 两个WD40转录因子对银杏类黄酮生物合成调控的研究

项目编号： No.31270717

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： 程水源

作者单位： 武汉轻工大学

项目金额： 86万元

中文摘要： 申请者前期利用cDNA-AFLP 结合RACE技术筛选出2 个参与调控银杏类黄酮合成的WD40类转录因子GbWD40-1 和GbWD40-2基因cDNA 全长。在此基础上，拟采用荧光定量PCR 技术与HPLC，分析叶片发育过程中这2个基因表达量与类黄酮含量变化的相互关系，并对其表达产物进行亚细胞定位分析；通过凝胶阻滞结合染色质免疫沉淀分析WD40与相关酶基因启动子结合的顺式元件特点，筛选WD40所调控的靶基因；利用酵母双杂交与免疫共沉淀分离验证与WD40的互作蛋白；进一步构建其正义与反义表达载体转化银杏愈伤组织，通过类黄酮含量、转录因子WD40基因以及相关结构基因表达谱分析，解析2个WD40的作用途径与调控网络，阐明WD40调控银杏类黄酮生物合成的分子机制。

中文关键词： 银杏；转录因子；GbWD40-1；GbWD40-2；GbWD40-3

英文摘要： In our previous work, We have isolated tow partial cDNA full lengthes of GbWD40-1 and GbWD40-2 genes from Ginkgo biloba, which two transcription factors are potentially involved in flavonoid biosynthesis pathway. The present research is to analyse the time and spatial expression profiles of these two genes by real-time PCR. In addition, We are going to determine the flavonoid content by HPLC method and reveal the correlative relation between the expression level of the two WD40 genes and flavonoid content. The EMSA and chromatin immunoprecipitation assay will be conducted for screening the target genes of the two WD40 transcription factors. Also, the combinational experiments of two yeast hybrid and Co-Immunoprecipitation will be carried out for isolation the interaction proteins of the two WD40 proteins. Furthermore, the sense and antisense plant expression vectors of the two WD40 genes will be constructed and be transferred into G. biloba calluses by Agrobacterium-mediated system. Afterwards, We will investigate the function of the two WD40 genes in regulating flavonoid biosynthesis and transcript level of related genes through overexpression and suppression experiments of the two WD40 genes in transgenic G. biloba callus lines. These results will provide theory model to unveil molecular regulation mechanism o

英文关键词： Ginkgo biloba；Transcription factor；GbWD40-1；GbWD40-2；GbWD40-3