芪合酶基因的克隆与植物表达载体的构建  

Cloning of Stilbene Synthase Gene and Construction of Its Plant Expression Vector

作　　者：谭琳[1,2] 郑学勤[2] 康由发[2] 施江[2] 

机构地区：[1]海南医学院生化教研室 [2]中国热带农业科学院生物技术国家重点实验室,海口571101

出　　处：《热带作物学报》2003年第2期42-45,共4页Chinese Journal of Tropical Crops

摘　　要：为获得含白藜芦醇且具有保健作用的转基因番茄,进行了芪合酶基因克隆、植物表达载体构建及导入受体细胞的研究。从葡萄2个品种———雷司令和粉红玫瑰的叶片中提取基因组DNA,并以其为模板,经PCR扩增芪合酶基因,各获得一长约1.6Kb的片段,将这2个片段分别连接到pGEM-Teasy和pUCm-T上进行测序,1个片段长为1622bp(命名为S1),另一个片段长为1617bp(命名为S2)。然后将S1正向插入pEV2的果实特异性启动子TFP2和NOS终止子,构建了植物表达载体pEV2S1;将S2正向插入植物表达载体pBI121的35S启动子和NOS终止子之间,构建了植物表达载体pBS2。将重组体pEV2S1和pBS2转化大肠杆菌E.coliXL1,并对重组体进行了筛选和鉴定。Total DNA was extracted from leaves of V.vinefera cvs.Leisiling and pink rose.A specific DNA fragment of about 1.6kb long was obtained by PCR with the total DNA served as template.The PCR products were cloned into pGEM-Teasy vector and pUCm-T vector respectively and named pS1and pS2respectively after their sequences were confirmed.The stilbene synthase genes were then excised from the plasmids by BamHI and SacI digestion and integrated into a binary vector,pBI121,and a fruit-specific vector,pEV2,from which theβ-glucuronidase(GUS)gene sequence had been removed by the same digestion,to prepare a35S promoter-stilbene synthase -nopaline synthase polyadenylation site construct and a fruit -specific promoter -stilbene synthase -nopaline synthase polyadenylation site construct.The recombinant plasmids were named pBS2and pEV2S1,respectively.

关 键 词：芪合酶基因 基因克隆 植物表达载体 转基因番茄 白藜芦醇 基因转化 

分 类 号：S641.2[农业科学—蔬菜学]