目的 通过基因工程技术获得重组结核分枝杆菌19-kDa蛋白。方法 应用PCR技术扩增结核分枝杆菌H37 Rv19-kDa DNA序列;以质粒pET24b为表达载体,构建19-kDa重组质粒,然后转化大肠杆菌BL21(DE3);在异丙基硫代-13-D-半乳糖苷(IPTG)诱导下,分别对不同诱导时间的表达产物进行十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE),凝胶经考马斯亮蓝染色检测蛋白。结果重组质粒pET24b-19-kDa测序表明与报道的序列相同。它在大肠杆菌BL21(DE3)细胞内以可溶性形式表达。不同IPTG诱导时间实验表明重组结核分枝杆菌19-kDa蛋白诱导3~4h重组蛋白在大肠杆菌中的表达量最高。结论 pET24b-19-kDa大肠杆菌工程株可高表达结核分枝杆菌重组19-kDa蛋白。
Objective To obtain recombinant 19 - kDa protein from Mycobacterium tuberculosis by using gene engi- neering technology. Methods The gene coding 19 - kDa protein inserted into a expression vector pET24b, and then transferred E. coli BL21 ( DE3 ). The expressed product was identified by SDS - PAGE, Plasmid pET24b containingl9 - kDa gene was transferred into competent Escherichia coli BL21 ( DE3 ) and 19 - kDa gene was over expressed by the inducement of IPTG. Resuits The recombinant 19 - kDa protein was expressed in soluble form in E. coli. When the culture was induced for 3 - 4 hours, the recombinant 19 - KDa protein was produced in highest quantity . Conclusion Escherichia coli containing recombinant plasmid pET24b - 19 - kDa gene could express recombinant 19 - KDa protein in high level.