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目的 在大肠杆菌中的高表达人 Era和人 Era C端蛋白 .方法 PCR扩增人 era基因 (h- era)的全长 c DNA和 h-era c DNA的 C端区域基因 . h- era c DNA克隆到 (His) 6融合表达载体 p RSET- C中 ,构建融合表达质粒并转化大肠杆菌BL 2 1(DE3) ,经 IPTG诱导表达 (His) 6 - h- Era融合蛋白 ;h- erac DNA的 C端区域基因克隆到非融合表达载体 p DH中 PL 启动子下游 ,转化大肠杆菌 TAP10 6 ,42℃热诱导表达人 Era C端蛋白 (h- Era- C) . SDS- PAGE电泳、凝胶薄层扫描检测蛋白的表达 .结果 表达的 (His) 6 - h- Era融合蛋白产物占全菌总蛋白的 80 % ;人 Era C端蛋白占全菌总蛋白的 40 % .结论 人 Era蛋白和 Era C端蛋白在大肠杆菌中获得了高表达 .
Objective To overexpress human Era and human Era C-terminal protein in E.coli.METHODS The full-length c DNA of human-era gene (h-era) and the C-terminal region of h-era c DNA were amplified by polymerase chain reaction (His) 6 fusion expression vector p RSET-C to construct a fusion expression plasmid and transformed into E. coli BL 2 1 (DE3), induced by IPTG expression of (His) 6 - h- Era fusion protein; h- erac DNA C-terminal region was cloned into the downstream of the PL promoter in the non-fusion expression vector p DH and transformed into E. coli TAP10 6. The human Era C-terminal protein (h-Era-C) was induced by heat at 42 ° C. SDS-PAGE electrophoresis, gel (His) 6 - h - Era fusion protein product accounted for 80% of the total bacterial total protein and human Era C - terminal protein accounted for 40% of the total bacterial total protein.Conclusion Human Era protein and Era C-terminal protein is highly expressed in E. coli.