马MxA基因真核载体表达重组质粒的构建与鉴定

Construction and Identification on Expression Recombinant Plasmid of MxA Gene Eukarya Vector in Horse

从马外周血单核细胞中提取RNA后,采用逆转录聚合酶链式反应(RT-PCR)方法,获得了MxAcDNA。扩增的目的基因片段经琼脂糖凝胶电泳回收,与pGEM-T Easy载体连接,转入大肠杆菌中筛选鉴定重组子。将质粒经酶切鉴定后,把酶切下的目的基因MxA cDNA进一步克隆到真核表达载体pCI-neo中,经PCR及序列鉴定,证实插入载体pCI中的片段为含有目的基因的核苷酸序列。真核表达重组质粒pCI-neo MxA cDNA的构建成功,为进一步研究马MxA蛋白的抗病毒作用奠定了基础。

MxA cDNA was obtained using RT-PCR methods after extracting RNA from horse external circle blood.The am-plified destinative gene fragment was connected with pGEM-T Easy vector and transformed to Escherichia coli to select recon.After the modification and identification of the plasmid,the destinative gene MxA cDNA was cloned into eukarya ex-pression vector pCI-neo.Through PCR and sequence analysis,the fragment in the vector pCI-neo was proved to be the nu-cleotide sequence containing the destinative gene.Eukarya expression recombinant plasmid pCI-neo MxA cDNA was success-fully constructed;it laid the foundation for the further study of MxA protein in horse.