Cloning and Prokaryotic Expression of Gene Encoding Cytoplasmic Domain of Human IL-23 Receptor

Objective To clone the gene encoding cytoplasmic domain of human IL-23 receptor(IL-23R)and express fusion protein in E.coli.Methods The gene encoding cytoplasmic domain of human IL-23R was amplified by PCR and cloned into vector pGEX-4T-1.The constructed recombinant plasmid pGEX-4T-1-hIL-23R(I)was transformed to E.coli BL21(DE3)for expression under induction of IPTG.The expressed product was identified by SDS-PAGE and Western blot.Results Agarose gel electrophoresis proved that the target gene fragment at a length of about 760 bp was amplified by PCR.Both restriction analysis and sequencing proved that recombinant plasmid pGEX-4T-1-hIL-23R(I)was constructed correctly.The fusion protein with a relative molecular mass of about 55 000 was expressed in a soluble form under induction of 0.5 mmol / L IPTG at 20℃.The expressed product contained about 16.1% of total soluble somatic protein and was recognized with rabbit anti-GST polyclonal antibody.Conclusion The gene encoding cytoplasmic domain of human IL-23R was successfully cloned, and soluble GST fusion protein was expressed in E.coli.