人TRAIL基因原核表达载体的构建及鉴定

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。

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

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.