人TRAIL基因原核表达载体的构建及鉴定  

Construction and Identification of Prokaryotic Expression Vector of Human TNF-related Apoptosis Inducing Ligand

作　　者：刘转转[1] 郝林[2] 范涛[2] 贺厚光[2] 尤红娟[1] 汤仁仙[1] 韩从辉[2] 

机构地区：[1]徐州医学院病原生物与免疫学教研室,江苏徐州221004 [2]徐州市中心医院泌尿外科,江苏徐州221009

出　　处：《现代生物医学进展》2011年第7期1220-1223,共4页Progress in Modern Biomedicine

基　　金：国家自然科学基金项目(30973443);徐州市科技计划项目(XM08C055)

摘　　要：目的:克隆肿瘤坏死因子相关凋亡诱导配体(TRAIL)基因片段(114～281氨基酸残基)并构建原核表达载体。方法:取健康人外周血提取总RNA,设计合成引物并引入EcoR I和Xho I酶切位点,RT-PCR扩增TRAIL基因的胞外区片段,克隆入原核表达载体pGEX-6P-1中,经双酶切、PCR及测序鉴定阳性克隆。结果:从外周血cDNA中扩增出501 bp的目的片段,测序结果证实成功构建重组质粒pGEX-6P-1/TRAIL。结论:成功构建TRAIL基因的原核表达载体pGEX-6P-1/TRAIL,为肿瘤细胞的凋亡研究提供理论依据。Objective：To clone and construct the prokaryotic expression vector of the TNF-related apoptosis inducing ligand gene（amino acids 114～281）.Methods：RNA was extracted from the peripheral blood of health adult.A pair of primers with restriction site EcoR I/Xho I was designed.The extracellular domain region of TRAIL gene was amplified by RT-PCR and then cloned into the prokary-otic expression vector pGEX-6P-1.The recombinants were confirmed by EcoR I/Xho I digestion,PCR,and DNA sequencing.Result：The extracellular domain region with a molecular size of 501 bp of TRAIL gene was amplified successfully.DNA sequencing showed that the recombinant plasmid was successfully constructed.Conclusion：A prokaryotic expression vector pGEX-6P-1/TRAIL was con-structed successfully,which provides theory for tumor apoptosis research.

关 键 词：TRAIL RT-PCR 克隆 

分 类 号：Q75[生物学—分子生物学] Q78