Expression of overlapping fragments of IPNV VP3 gene and analysis of antigenic epitope domains

Four overlapping fragments L1,L2,L3 and L4 of VP3 gene of infectious pancreatic necrosis virus(IPNV) were amplified by PCR.PCR products were cloned into the expression vector pGEX-6P1 and pET32a.The recombinant plasmids pGEX-6P1-VP3(L1),pET32a-VP3(L2),pGEX-6P1-VP3(L3) and pGEX-6P1-VP3(L4) were identified by digestion,PCR and sequencing,and transformed into E.coli Rosetta and BL21 cells.The target proteins of 32 kD,26 kD,30 kD and 31 kD were obtained by induction using IPTG at 1.0 mmol/L.The VP3(L1) and VP3(L4) protein reacted with IPNV positive serum in Western-blotting,indicating that the immunodominant region of the VP3 protein was located in 1~70 and 143~206 amino acid sequence of VP3 protein.The VP3 recombinant fusion protein can be used as the specific diagnosis antigen for ELISA assay.This study will serve as the basis for further precise identification of VP3 protein epitope.