Prokaryotic expression of porcine interleukin-18 gene and purification of the expressed IL-18 protein

The mature protein gene of porcine interleukin-18 was amplified from recombinant plasmid pcDNA3.1-pIL-18 by PCR.The prokaryotic expression plasmid pET32c-pIL-18 was constructed through KpnⅠ+ SacⅠ digestion and ligation,and the recombination was identified by enzyme digestion,PCR amplification and DNA sequencing.The recombinant plasmid was transformed into Escherichia coli BL21(DE3) competent cells,and expression of the protein was induced with IPTG at 37℃ overnight.SDS-PAGE(analysis) showed that the recombinant plasmid was highly expressed in E.coli, and molecular weight of the expressed protein was approximately 33ku.After denaturation with 8 mol/L urea,the recombinant IL-18 protein which was purified with Ni~(2+)-NTA His Bind Resin was refolded by dialysis against PBS and water.Western-blotting analysis confirmed that the purified protein had immunogenicity.These results provide the foundation for the practical application of the recombinant IL-18.