人TRAIL细胞外段基因在大肠杆菌中的表达及其包涵体的复性  

Expression of Extracellular Domain Gene of Human TRAIL in E.coli and Re-naturalization of Inclusion Body

作　　者：田生和[1] 全家妩[1] 

机构地区：[1]武汉生物制品研究所,武汉430060

出　　处：《中国生物制品学杂志》2005年第2期114-116,共3页Chinese Journal of Biologicals

摘　　要：目的　在大肠杆菌中表达人肿瘤坏死因子相关凋亡诱导配体细胞外段 (sTRAIL)基因 ,并研究其包涵体的复性。方法　应用RT- PCR法从正常人外周血单核细胞中获得sTRAIL的编码基因 ,将其克隆到原核表达载体pBV2 2 0 ,于大肠杆菌DH5α中通过温控诱导表达 ;将包涵体进行变性和复性处理后 ,采用离子交换层析纯化表达产物 ,并经Dotblot及MTT试验鉴定。结果　通过温度诱导 ,目的基因主要以包涵体形式高效表达 ,包涵体蛋白经梯度透析法复性可获得具有抗肿瘤活性的重组人sTRAIL。结论　已获得大量纯化重组人sTRAIL ,为开发新型抗肿瘤制剂奠定基础。Objective To express the extracellular domain gen e of human tumor necrosis factor-related apoptosis inducing ligand (sTRAIL) in E.coli and study the re-naturalization of inclusion body.Methods Amplify the gene encoding sTRAIL from normal human peripheral blood monon uclear cells by RT-PCR,clone into prokaryotic expression vector pBV220 and tran sform to E.coli for expression under induction of temperature.The expressed product existing in a form of inclusion body was de-naturalized,re-naturalized and purified by ion exchange chromatography,then identified by dot blot and MTT test.Results sTRAIL was highly expressed in a form of incl usion body and showed anti-tumor activity after re-naturalization by gradient dialysis.Conclusion A large quantity of purified recombinant hu man sTRAIL was obtained.It laid a foundation of developing novel anti-tumor dru gs. [

关 键 词：TRAIL 包涵体 复性 高效表达 细胞外 目的基因 大肠杆菌 诱导表达 纯化 离子交换层析 

分 类 号：Q786[生物学—分子生物学] R378[医药卫生—病原生物学]