Cloning and expression of the specific genes from Yersinia pestis and analysis of their antigenicity / 中国地方病学杂志

ABSTRACT

Objective To clone and express specific genes (YP01089,psi.ymt) from Yersinia pestis in Escherichia Coli(E.coli)and to analyze the antigenicity of these recombinant proteins.Methods The target genes were amplified by polymerose chmn reaction(PCR).The amplified products were ligated with pET-30a(+) vector after purification and cut by two different restriction enzymes,then these recombinant plasmids were transfefred into the host cells of E.coli BL21(DE3) strain.The target genes were successfully expressed following induction with Isopropyithio-β-D-galactoside(IPTG),and the target proteins were purified by the method of affinity chromatography.Sodium dodecyl sulfate-polyaerylamide gel eleetrophoresis(SDS-PAGE)and Westem blot were used to detect the expressed recombinant protein.Results Three recombinant plasmids were finally constructed. rYP01089,rPst and rYmt were expressed stably and effectively in E.coli thmngh optimizing the induction condition. The Western blot analysis indicated that rPst was capable of binding with positive sernm of phgue.The purity of rest was up to 95%in this stuay.Conclusions This work indicates that the genes of Yersinia pestis are able to be efficiently expressed in the prokaryotie protein expression system.The immune characteristic of rPst is sensitive and specific,80 this study has settled a foundation for developing a new type diagnostic reagent of plague.