Prokaryotic Expression Of GP5 And Nsp7 Of PRRSV And Establish An Indirect ELISA Based On Expression Produce

The outbreak of Porcine reproductive and respiratory syndrome(PRRS) bring huge economic losses of the global. It’s a hot topic about how to prevention and treatment PRRS effectively, the measures of RT-PCR; ELISA were more effect in the detection of PRRSV, ELISA test was used intensively because of rapidity and effectively. The prevention of PRRS in our country was mainly rely on the injection of inactivated vaccine, but single structural protein or single non-structural protein could not be used for the differentiation of post-infection and post-vaccination antibodies in animals vaccinated with inactivated vaccine, it made a trouble in the detection of PRRS, new ELISA test was urgent in need.The envelope glycoprotein(GP5) was the main target protein in the detection of PRRSV antigen, the non-structural protein 7(Nsp7) was coded by the relatively conserved region of the viral genome and have high similarity in different strains, could be used as an antigen to identify where did antibody come.The host could induce antibody to structural proteins when injected with inactivated vaccine and infected with PRRSV,therefore, it was not suitably to use structural proteins as antigens only. The goal of this work is to use non-structural protein as an antigen in an indirect ELISA test on the base of structural protein to identify PRRSV effectively. GP5 and Nsp7 were amplified from isolate strains of PRRSV by PCR, connected target gene with pET28 a, transformed pET28a-GP5, pET28a-Nsp7 into BL21(DE3)respectively, then identified them by sequence analysis. Induced expression with IPTG, the expression products were identified by SDS-PAGE. The result shows that GP5 and Nsp7 were expressed successfully, analyzed by western blot showed that the recombinant proteins have good sensitivity and specificity. The GP5-Nsp7-ELISA was established successfully, after terms and conditions were optimized.