人THAP11慢病毒载体的构建及表达研究  

Establishment of a recombinant lentivirus containing human thanatos-associated protein 11 gene

作　　者：黄先红[1] 李长燕[2] 詹轶群[2] 王治东[2] 许望翔[2] 杨晓明[2] 汪思应[1] 

机构地区：[1]安徽医科大学病理生理学教研室,合肥230032 [2]军事医学科学院放射与辐射医学研究所,北京100850

出　　处：《安徽医科大学学报》2010年第5期605-609,共5页Acta Universitatis Medicinalis Anhui

基　　金：北京市自然基金重点项目(编号:07G0802);国家自然科学基金重点项目(编号:30630035);国家自然科学基金面上项目(编号:30870958)

摘　　要：目的构建人死亡相关蛋白11(THAP11)基因的慢病毒载体,并建立其慢病毒表达系统。方法 PCR方法获得人THAP11基因,限制性内切酶酶切和基因重组构建慢病毒载体质粒pBPLV-THAP11-myc,并通过转染HEK293细胞观察绿色荧光蛋白(GFP)的表达以及Westernblot检测其表达。在脂质体介导下将重组质粒与包装质粒pLP1和pLP2、包膜质粒pLP/VSVG共转染293FT细胞包装产生慢病毒。结果构建的质粒经PCR,酶切及测序鉴定正确;该质粒与包装质粒共转染293FT细胞获取的5.5×106TU/ml慢病毒滴度。结论成功构建了人THAP11基因慢病毒载体质粒THAP11-myc-pBPLV,并建立了其慢病毒表达系统,为后续的应用研究奠定了基础。Objective To construction of lentiviral vector containing thanatos-associated protein 11 gene and establishment of an expression system. Methods The coding region of THAP11,amplified by polymerase chain reaction, was cloned into pBPLV vector by restriction enzymes. The recombinant vector was transfected transiently into HEK293 cell and the expression of THAP11 was investigated by Western blot and fluorescent microscopy. The pB-PLV-THAP11-myc plasmid was cotransfected with three packaging plasmids pLP1,pLP2,pLP/VSVG into 293FT cells to produce a replication-in-competent lentivirus. Results The lentivirus carring THAP11 gene was successfully packaged and the titer of lentivirus grossly obtained for 48 hours after cotransfection was 5. 5 × 106 transducing units（ TU） /ml. Conclusion The lentiviral expression system of thanatos-associated protein 11 gene establishes successfully.

关 键 词：质粒 遗传载体 

分 类 号：R394.3[医药卫生—医学遗传学] R373.9[医药卫生—基础医学]