HSV-tk基因逆转录病毒重组体的构建与DNA序列分析

目的　构建含有单纯疱疹病毒Ⅰ型胸苷激酶 (HSV1 tk)基因的逆转录病毒重组载体pLXSN TK。方法设计一对寡核苷酸引物 ,用PCR方法从质粒pHSV10 6中特异扩增HSV tk基因片段 ( 1168bp) ,分别用BamHI和Eco RI酶切后 ,定向连接到质粒pLXSN中 ,转化宿主菌TG1,分别用上述内切酶 ,PCR和DNA测序鉴定重组质粒。结果　酶切鉴定所切下的片段和PCR扩增的片段大小均与预计相符 ,测序结果与文献报道序列及预计结果一致 ,证实符合表达框架。结论　成功构建了HSV tk嵌合重组质粒pLXSN TK。

Construction of the Retroviral Recombinant Plasmid Vector Encoding Thymidine Kinase Gene of the Herpes Simplex Virus and DNA Sequencing

Objective To construct the retroviral recombinant plasmid vector encoding TK gene of the herpes simplex virus.Methods\ The specific primers were designed and synthesized to amplify the DNA fragment (PCR method) of HSV1 tk gene from the plasmid pHSV106. The length of the fragment was 1168bp.After digested by appropriate restriction endoenzymes(BamHI and EcoRI),the fragment of HSV1 tk gene was cloned into pLXSN according to its special orientation.Then the recombinant plasmid was identified by digesting of endoenzymes,PCR and DNA sequencing.Results\ The fragments digested by endoenzymes and amplified by PCR method were as large as the predicted results.DNA sequencing was the same with the sequence reported on the literature.Conclusion\ The retroviral recombinant plasmid pLXSN TK was successfully constructed.