is a high-valued flatfish species because of the desirable taste of its flesh and its wide tolerance to salinity and temperature. It has become an economically important farmed marine fish in China in recent years. Insulin-like growth factor (IGF)- plays an important role in the regulation of growth and development in teleosts. Our objective was to evaluate the physiological functions of IGF-. The cDNA sequence encoding the mature peptide domain of the A pair of primers were designed based on the conserved sequences of IGF-from Pleuronectiformes and Perciformes species. A 210 bp fragment was obtained and verified as the mature peptide fragment. A homology comparison revealed that the B-, A-, and D-domains of the IGF- mature peptide were highly conserved among vertebrates, but with changes in the C-domain. The mature peptide fragment was reconstructed by RT-PCR with a pair of primers and the subcloned into the prokaryotic expression vector pET-28a with T4 DNA polymerase to successfully construct an recombinant plasmid. The recombinant IGF-II plasmid was highly expressed in being induced by IPTG with a special fusion polypeptide containing His6 at its N-terminus. The SDS-PAGE analysis showed that the polypeptide was expressed in the form of inclusion bodies with a molecular weight of 11.4 kD and accounted for . Western blotting analysis indicated the fusion polypeptide had antigenicity to His6 antibody with primary antibodies (mouse-anti-His6 monoclonal antibody) and secondary antibodies (goat-anti-mouse IgG labeled with horseradish peroxidases). The inclusion bodies were denaturalized using 6 mol/L guanidine HCl, purified using Ni-NTA affinity chromatography, and annealed by gradient dialysis (8 , 0) in urea, after which the purified protein with a molecular weight of 11.4 kD was obtained. The concentration of the recombinant IGF-II was measured using the BCA method after being subject to ultrafiltration. The proliferation experiment revealed that the recombinant protein significantly promoted the proliferation of human embryonic kidney cells (HEK293T) at a concentration of 1.8 μg/mL and inhibited proliferation at a concentration of 5.4 μg/mL, thus verifying its biological activity at a cellular level. In summary, we successfully constructed an prokaryotic expression system and optimized the expression conditions (induction temperature, time, and IPTG concentration). A biologically active fusion protein was obtained following separation, purification, and renaturation. Our results provide information to better understand the role of .