Prokaryotic expression,purification,identification,and polyclonal antibody preparation of human SUMO-1 gene

Objective To construct a prokaryotic plasmid expressing the recombinant proteins of human SUMO-1 and prepare the polyclonal antibodies of rabbit anti-human SUMO-1.Methods The fragment of SUMO-1 gene encoding 97 amino acids in the N terminus was amplified from pcDNA-HA-SUMO-1-GG plasmid by PCR.The fragment was then inserted into the prokaryotic expression vector to construct the recombinant plasmid pET41a(+)-SUMO1,and expressed in E.coli.BL21(DE3)pLysS.After induction with IPTG,the fusion protein was obtained,and then purified by GST affinity chromatography.A polyclonal antibody,analyzed by Western blotting and ELISA staining,was developed by immunizing rabbit with the purified recombinant protein.Results The recombinant plasmid was constructed successfully;the fusion protein was successfully expressed and purified.Western blotting and ELISA staining demonstrated that the polyclonal antibody was obtained by immunizing rabbit with the purified recombinant protein.Conclusion The preparation of the polyclonal antibody against human SUMO-1 lays a foundation for the further study on the function of SUMO-1 protein.