摘要
将质粒pSB166中包含ED35s启动子、Omega元件及TNOS终止子的一段核苷酸序列定向克隆到质粒pCAMBIA1303,构建了中间表达载体pWR306;以中国野生葡萄华东葡萄白河35-1cDNA为模板,通过PCR扩增出葡萄芪合成酶基因(STS)、醛脱氢酶基因(ALDH),与pGEM-TEasy克隆载体连接,获得重组质粒pGEM-TEasy-STS和pGEM-TEasy-ALDH;双酶切重组质粒及表达载体pWR306,将STS、ALDH基因片段与线性表达载体pWR306进行定向连接,构建了葡萄芪合成酶基因及醛脱氢酶基因的植物表达载体pWR-STS、pWR-ALDH,并用改进冻融法导入农杆菌GV3101。
A nucleotide sequence of plasmid pSB166 carrying promotor ED35s,Omega TMV translation enhancer and nopaline synthase terminator was specifically cloned into plasmid pCAMBIA1303 to construct intermediate vector pWR306;stilbene synthase gene and aldehyde dehydrogenase gene were amplified by PCR with the cDNA template of Vitis pesudoreticulata Baihe35-1 and then linked to cloning vector pGEM-T easy to obtain recombinant plasmids pGEM-TEasy-STS and pGEM-Teasy-ALDH;the recombinant plasmid from double-enzyme digestion,and expression vector pWR306 linked the segments of stilbene synthase gene and aldehydehyde dehydrogenase gene with linear expression vector pWR306 to construct the plant expression vectors of stilbene synthase gene and aldehydehyde dehydrogenase gene,pWR-STS and pWR-ALDH,which were introduced into Agrobacterium strain GV3101 by means of the improved freezing-thawing method.
出处
《西北植物学报》
CAS
CSCD
北大核心
2005年第5期851-857,共7页
Acta Botanica Boreali-Occidentalia Sinica
基金
国家转基因植物研究与产业化开发专项(JY03A20-02)
国家自然科学基金资助项目(30170653)
关键词
中国野生葡萄
葡萄芪合成酶基因
葡萄醛脱氢酶基因
植物表达载体
wild grape species in China
stilbene synthase gene
aldehydehyde dehydrogenase gene
plant expression vector
