肠炎沙门菌调控基因hilD与hilA的体外表达规律研究

Analysis of the in vitro expression characteristics of regulatory genes hilD and hilA from Salmonella enterica serovar Enteritidis

利用原核表达载体pCold I构建肠炎沙门菌转录调控蛋白HilD和HilA的重组表达菌,并采用Western-blot技术检测两者在体外培养条件下的表达规律。结果表明:成功构建了重组表达大肠埃希菌(E.coli)BL21(pCold I-hilD)和E.coli BL21(pCold I-hilA),经IPTG诱导得到了重组表达的rHis-HilD和rHis-HilA蛋白。将重组表达的蛋白免疫BALB/c小鼠,制备蛋白的多克隆抗体,Western-blot进一步证明了抗体与蛋白可发生特异性反应。肠炎沙门菌C50041在LB培养条件下,HilD与HilA蛋白表现出不同的表达规律,从而为体外研究肠炎沙门菌SPI1与SPI2效应蛋白的表达和功能研究奠定了基础。

The prokaryotic expression plasmids pCold I was used to construct recombinant expression bacteria expressing two regulators hilDand hilAfromSalmonella enterica serovar Enteritidis,and western blot analysis was applied in detecting the expression pattern of both regulators under the culture condition of LB medium.The result showed that the recombinant expression bacteria E.coli BL21（pCold I-hilD）and E.coli BL21（pCold I-hilA）was successfully constructed.When the bacteria were induced by IPTG,the recombinant proteins were collected,purified,and then inoculated into BALB/c to induce the production of polyclonal antibodies,which were identified by western blot analysis.The Western blot result revealed the different expression pattern of both regulators in vitro,which could be used to study the function of effector proteins belonging to SPI1 and SPI2 in vitro.