弓形虫昆山分离株P30蛋白的纯化、复性和鉴定

目的表达并纯化弓形虫昆山分离株膜蛋白P30。方法将构建的pET28b-P30重组质粒转入大肠埃希菌BL21,挑重组菌于LB培养基中增菌,IPTG诱导其表达,然后纯化包涵体,用尿素溶解包涵体,Ni2 螯合柱纯化产物,以透析法复性蛋白,并用SDS-PAGE和Western blot进行鉴定。结果大肠埃希菌表达的P30蛋白以包涵体的形式存在,P30蛋白经Ni2 螯合柱纯化后纯度大于95%。经Western blot鉴定,复性蛋白能被弓形虫感染的鼠血清识别。结论得到了纯度高的膜蛋白P30。

Purification, renaturation and identification of P30 antigen of Toxoplasma gondii Kunshan strain

Objective To express and purify the P30 antigen of Toxoplasma gondii Kunshan strain.Methods The recombinant plasmid pET28b-P30 was transformed into Escherichia coli BL21,and the recombinant bacteria was incubated in LB culture and induced by IPTG.The inclusion bodies were lysed in urea.After purified by Ni2 chelating column,the solubilized fusion protein was renaturated and identified by the sera from mice infected with Toxoplasma.Results The P30 protein was expressed in E.coli as an inclusion body and it's purity was above 95% after purification.The renaturated protein can be recognized by sera from mice infected with T.gondii.Conclusion The P30 protein with high purity and good performance after renaturation was obtained.