苦瓜(Momordica CharantiaL.)MADS-box基因BAG启动子的克隆分析及表达载体的构建  

Cloning and Analysis of the Promoter of Mads-Box Gene "BAG" In Bitter Melon and Construction of Expression Vector

作　　者：杨满业[1] 赵茂俊[2] 唐琳[1] 徐莺[1] 王胜华[1] 邢杰[1] 陈放[1] 

机构地区：[1]四川大学生命科学学院,成都610064 [2]四川农业大学生命科学与理学院,四川雅安625000

出　　处：《四川大学学报（自然科学版）》2003年第4期783-786,共4页Journal of Sichuan University(Natural Science Edition)

基　　金：国家自然科学基金(30270090);教育部博士点基金;四川省重点科研项目基金

摘　　要：作者提取苦瓜基因组总DNA,构建了DNA步移文库,并根据已克隆并公布的苦瓜MADS box基因BAG的基因序列设计引物,通过染色体步移技术克隆出BAG基因起始密码子上游调控序列BAGP.对BAGP的鉴定和分析表明其具备大多数高等植物启动子的保守元件,预测它对BAG基因的表达具有一定的作用.为鉴定BAG基因的基本启动子元件,将基因5′侧翼序列做缺失片段分析,利用PCR方法从BAGP中得到3个大小不等两端带有HindⅢ,BamHⅠ酶切位点的片段BAGP1,BAGP2和BAGP3,定向插入载体pMGFP4(pBI221改建,报告基因为GFP)中,取代原有的CaMV35S启动子,构建了由驱动报告基因GFP的植物表达载体BAGPV1,BAGPV2和BAGPV3.One pairs of specific nested PCR primers were designed according to the pubilished sequence of MADSbox gene 'BAG' in bitter melon. The promoter of Madsbox gene (BAGP) with a length of 664 bp was cloned by DNA walking technology. The sequence of BAGP was analyzed by software DNA Tools 5.1 and CallMat, which showed that it has most of conservative elements of higher plants' promoter. In order to establish a foundation for further study of BAGP function in promoting BAG gene specific expressions in higher plant reproductive organs, three BAGP absence sequences BAGP1(664bp),BAGP2(564bp)and BAGP3(485bp)were obtained by PCR and inserted into another vector pMGFP4(with GFP report gene) to take place of CaMV35S promoter to get three expression vector BAGPV1,BAGPV2 and BAGPV3. 

关 键 词：苦瓜 启动子 MADS-BOX基因 DNA步移技术 表达载体 PCR扩增 

分 类 号：Q785[生物学—分子生物学] Q786