Abstract: LrrG (leucine-rich repeat protein from GBS) and Sip (surface immunogenic protein), which are two kinds of surface antigen proteins from Streptococcus agalactiae in tilapia, have good immunogenicity. To obtain the LrrG-Sip fusion protein via coalescing surface antigen protein LrrG and Sip of S.agalctiae in tilapia, we cloned Sip and LrrG genesinto vector pColdⅡone by one using double enzyme method of gene splicing technology, and constructed a prokaryotic expression vector pColdⅡ-LrrG-Sip. The recombinant plasmid was transformed into E.coli BL21(DE3), and the result indicated that 9 h, 15 ℃, 0.5 mmolL-1 IPTG were the optimum inducing conditions under which fusion protein was most soluble and abundant. Western blotting test showed that the LrrG-Sip fusion protein was about 160 kDa, consistent with the prediction (162 kDa), which suggested the prokaryotic expression vector pColdⅡ-LrrG-Sip was constructed successfully and laid the foundation for developing subunit vaccines forS.agalctiae in tilapia.