Expression and identification of Yersinia pestis fusion protein YPO1996.

Objective To construct the recombinant plasmid pET32a-ypo1996 based on Yersinia pestis comparative proteome and express the protein rYPO1996 in order to develop a new method of diagnosing Y.pestis.Methods Primers were designed according to the YPO1996 gene sequence of Y.pestis strain CO92.The target DNA fragments of YPO1996 were amplified by polymerase chain reaction(PCR),and the products were inserted directionally into a pET32a(+) vector.The recombinant plasmid was transformed into E.coli BL21(DE3) and inducted by IPTG.The fusion protein was effectively expressed and purified.The expressed protein was analyzed by SDS-PAGE and Western blot. Results The target gene YPO1996 had been inserted successfully into pET32a(+),and the results of SDS-PAGE showed that the expression product was about 53.11 ku as predicted.The fusion protein rYPO1996 performed well in terms of solubility and can be recognized by antibodies against the Y.pestis strain(EV strain) in Western blot.Conclusion The new pET32a-ypo1996 plasmid is successfully constructed in a prokaryotic expression system,and the protein rYPO1996 showed satisfactory solubility and antigenicity.It may be an acceptable candidate antigen for the diagnosis of Y.pestis infection.