In order to exogenously express the MAP3732c protein of Mycobacterium avium subspecies paratuberculosis,this study constructed the prokaryotic expression plasmid pET28a-MAP3732c,purified the recombinant protein of MAP3732c by His-tag affinity chromatography after the induced expression and identification,and then immunised Balb/c mice several times to prepare the polyclonal antibody,detected the antibody potency by indirect ELISA method.The results showed that the recombinant plasmid T1-MAP3732c was successfully constructed and the recombinant plasmid pET28a-MAP3732c was successfully induced at a final concentration of IPTG of 0.5 mmol·L-1 at 37℃;SDS-PAGE results showed that MAP3732c recombinant protein was about 28 ku in size and existed in the form of inclusion bodies;Western blot results showed that the recombinant protein had good immunoreactivity with His antibody and bovine paratuberculosis positive serum,while negative serum did not bind non-specifically;the ELISA results showed that the anti-MAP3732c polyclonal antibody had a potency of 1:204 800.This study provided a reference for the immunogenicity analysis of MAP3732c protein of Mycobacterium avium subspecies paratuberculosis and detection methodology of paratuberculosis.
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