毛尖紫萼藓GH394基因克隆、表达载体的构建  

Grimmia pilifera P.Beauv GH394 Gene Cloning and Expression Carrier Construction

作　　者：张百成[1] 沙伟[1] 宋璐[1] 

机构地区：[1]齐齐哈尔大学生命科学与农林学院,黑龙江齐齐哈尔161006

出　　处：《北方园艺》2012年第12期113-116,共4页Northern Horticulture

基　　金：国家自然科学基金资助项目(31070180);齐齐哈尔大学研究生创新科研资助项目(YJSCX2010-010X)

摘　　要：利用PCR技术,以毛尖紫萼藓总RNA为模板,扩增出上、下游分别加入BamHI、SacⅠ酶切位点的GH394(657 bp)基因CDS区全长序列,采用pMD18-T Vector、pRI 101-AN Vector构建了该基因克隆、表达载体,并将重组载体质粒转化到大肠杆菌(Escherichia coli)DH 5α和根癌农杆菌(Agrobacterium tumefaciens)工程菌株GV3101中,并选用X-gal、IPTG和利福平(Rifampicin,Rif)、卡那霉素(Kanamyein,Km)筛选阳性菌株。结果表明:已成功构建pMD-GH394克隆载体和pRI 101-AN-GH394表达载体并将重组质粒转入目的菌株中;该试验为后续实现毛尖紫萼藓GH394基因抗旱预期功能的验证,奠定了良好的试验基础。Using the Grimmia pilifera P.Beauv of totle RNA to template,amplified GH394（657 bp） gene in the full sequence of CDS area which is upstream and downstream adding the restriction site was BamHⅠand SacⅠ,using pMD18-T Vector and pRI 101-AN Vector constructed with Cloning vector and Expression vector for this gene,then, recombinant plasmid transformation to Escherichia coli and Agrobacterium tumefaciens,using X-gal,IPTG and Rif, Kanamycin to screening of positive strains.The results showed that constructed with pMD-GH394 Cloning vector and pRI 101-AN-GH394 Expression vector were succeed,and recombinant plasmid transformation to objective strains.The experimental provided an good essential for realization of Grimmia pilifera P.Beauv GH394 gene expected drought functional verification.

关 键 词：毛尖紫萼藓 抗旱 T-PCR 基因克隆 载体构建 

分 类 号：Q348[生物学—遗传学]