Prohibitin基因的shRNA真核表达载体的构建及鉴定

Construction and identification of shRNA eukaryotic expression vectors targeting to prohibitin gene

目的:构建靶向prohibitin基因的shRNA真核表达载体.方法:根据prohibitin mRNA序列,利用Ambion网上设计软件设计两条针对该基因不同靶点的shRNA干扰序列,克隆到线性化的质粒载体上,并进行酶切鉴定和测序.结果:酶切结果表明构建的shRNA已插入载体,经测序证明与设计一致.结论:成功构建两个靶向prohibitin基因的shRNA真核表达载体,为下一步研究prohibitin基因在肿瘤细胞中的作用和后续的的体内外实验奠定了基础.

AIM： To construct 2 shRNA eukaryotic expression vectors targeting to prohibitin gene. METHODS： Finding the sequence of human prohibitin gene in Pnbmed, the oliganucleotides of shRNA were designed by the software of Ambion Company then synthesized and directionally cloned into pGenesill. 1 vector, Then the recombinant vectors were extracted and confirmed by restriction enzyme and sequencing analysis. RESULTS： The restriction enzyme analyses demonstrated that shRNA had been inserted into vectors, and sequencing analyses demonstrated that their sequences were the same as the design. CONCLUSION： We have successfully constructed shRNA eukaryotic expression vectors targeting to prohibitin gene. It laid the foundations for the further study on the function of prohibitin gene in tumor cell and the future RNAi experimental research in vivo and in vitro.