In order to acquire vaccine antigen of triple outer membrane protein (omp) which can simultaneously prevent three common pathogens in eel,three genes of ompU,ompA and omp porin П were selected from Vibrio vulnificus, Edwardsiella tarda and Aeromonas hydrophila,respectively.Three gene fragments which encoded the outer regions of the three omps and showed rich epitopes were amplified and combined by two times of “two-steps” fusion PCR.According to the restriction sites of the expression vector (pGEX-2T-his plasmid),two restriction sites of BamHⅠand EcoRⅠwere added to the 5′ points of two primers which are used to amplify fragments of ompU and omp porin П,respectively.The fusion fragment was connected with pGEX-2T-his plasmid and the recombinant expression vector which expressed the fusion omp was constructed.After the vector was transformed into E.coli BL21,the fusion omp with a molecular weight of 85 ku was highly expressed by using 0.25 mmol/L IPTG to induce the tansformant (E.coli BL21) overnight at 16 ℃ in a suitable bacteria concentration (D600 nm=0.8).The expressed protein was eluted and purified by nickel column,and then renaturated by dialysis with gradient of urea.