rhIL-6表达载体的构建及其在大肠杆菌中的高效表达  

Construction of Recombinant Human IL- 6 Expression Vector and Its High Level Expression in E. coli

作　　者：田志刚[1] 刘杰[1] 孙汭[1] 田彤[1] 张捷[1] 明雨[1] 张建华[1] 

机构地区：[1]山东省医学科学院山东肿瘤生物治疗研究中心,济南250001

出　　处：《中国生化药物杂志》1994年第4期235-238,共4页Chinese Journal of Biochemical Pharmaceutics

摘　　要：通过PCR技术扩增出约0.5kb的IL-6基因片段,并且采用含有P_RP_L串联启动子及SD区的高效表达载体pBV₂20,经DNA重组技术,成功地构建了rhIL-6的基因工程菌E.coli DH₅a/pBV₂20IL-6。所表达的IL-6经SDS-PAGE证明约17.5kD,表达量占菌体总蛋白的30%以上。以IL-6依赖细胞株MH₆0·BSF₂检测,表达产物粗提物1:100复性后效价可达到5×10⁸u/ml,每升发酵菌液可获10¹2u IL-6粗品。Human IL-6 cDNA fragment (0. 5kb) was amplified through PCR method, inserted into EcoR I /BamH I site of pBV220 expression vector which contains PRPL promoters and SD sequence. After transformation and screening the E. coli DH5α in Amp+/LB medium, rhIL-6/pBV220 positive clone ratio was as high as 80%. The expression level of rhIL-6 was more than 30% of the total bacterial protein in SDS-PAGE, molecular weight was 17. 5kD.The biological activity of renatured bacterial extracts(1 : 100 dilution) was 5×108 u/ml on MH60 ·BSF2 cell lines, that is, 1012u rhIL-6/L bacterial fermentation.

关 键 词：高效表达 DNA 重组 白细胞介素6 基因工程 

分 类 号：R394.8[医药卫生—医学遗传学] R392－33[医药卫生—基础医学]