猪葡萄球菌脱落毒素ExhA的表达与纯化

Expression and purification of porcine Staphylococcus hyicus exfoliative toxin ExhA

为了研究猪葡萄球菌脱落毒素ExhA的生物学活性,试验以猪葡萄球菌的基因组DNA为模板,PCR扩增ExhA全基因序列,连接到pCold TF表达载体上,构建重组质粒pCold TF-ExhA,将重组质粒转入E.coil BL2（DE3）,IPTG低温诱导目的蛋白可溶性表达,重组表达的蛋白用Ni-IDA琼脂糖纯化树脂纯化,Western-blot及攻毒试验验证表达蛋白质的生物学功能。结果表明：成功构建了pCold TF-ExhA重组质粒,并且脱落毒素ExhA主要以可溶性形式表达,表达纯化的脱落毒素ExhA具有完整的生物学活性。说明表达的脱落毒素ExhA具有良好的抗原性。

To study the biological activity of Staphylococcus hyicus exfoliative toxin ExhA in pig, ExhA gene was amplified by PCR from genomic DNA of Staphylococcus hyicus and ligased to the pCold TF expression vector to construct recombinant plasraid pCold TF-ExhA. The recombinant plasmid was transformed into E. coil BL2 （ DE3）, then the target protein was soluble expressed by the induction of IPTG at low temperature. Next, the recombinant protein was purified by NI-1DA resin column. Western-blot and challenge experiments were carried out to verify the biological function of expressed proteins. The results showed that the recombinant plasmid pCold TF-ExhA was constructed and the ExhA protein was purified successfully,which has completed biological activity. The results indicated that the expressed protein has good antigenicity.