鼠伤寒沙门氏菌STM1697蛋白的原核表达及纯化

Prokaryotic expression and preliminary purification of STM1697 protein of Salmonella typhimurium

目的原核表达并纯化沙门氏菌毒性相关蛋白STM1697,为其功能研究奠定基础。方法以鼠伤寒沙门氏菌(ATCC14028)基因组为模板,PCR扩增STM1697基因,双酶切后连接到原核表达载体pGL01中。将阳性重组子转化入大肠埃希菌BL21(DE3)以IPTG诱导,表达产物经镍离子亲和柱进行纯化,并用SDS-PAGE分析鉴定蛋白及纯度。结果成功将STM1697基因克隆到原核表达载体pGL01中。STM1697蛋白在大肠埃希菌BL21(DE3)以可溶形式表达,经镍离子亲和柱纯化后得到的蛋白纯度>95%。结论伤寒沙门氏菌STM1697蛋白在大肠埃希菌中可稳定表达,为其功能研究奠定了基础。

Objectives To express and purify the toxic protein STMJ697 of Salmonella typhimurium in order to provide a foundation for study of its function. Methods Using the S. typhimurium （ATCC14028） genome as a template, the STM1697 gene was amplified with PCR. After double enzyme digestion, the gene was ligated into the prokaryotic expression vector pGL01. The positive recombinant plasmid was transformed into Escherichia coli BL21 （DE3） and its expression was induced with IPTG. The expressed product was purified with a nickel ion affinity column. The protein was identified and its purity was determined using SDS-PAGE analysis. Results The STM1697 gene was cloned into the prokaryotic expression vector pGL01. STM1697 protein was expressed in E. coli BL21 （DE3） in soluble form. After purification with a nickel ion affinity column, the purity of the protein was 95 %. Conclusion STM1697 protein of S. typhimuriurn can be stably expressed in E. coli. This finding lays the foundation for study of its function.