Cloning and prokaryotic expressing of human interleukin-23 receptor

Objective:To clone the encoding sequence of human interleukin-23 receptor(hIL-23R) from PBMC,to construct prokaryotic expression vector and to express hIL-23R in E.coli.Methods:Using the total RNA isolated from human PBMC as template,the 1890bp cDNA of hIL-23R was amplified by reverse transcription-polymerase chain reaction(RT-PCR).The PCR product was cloned into pMD-19 T vector,digested by restriction enzyme and sequenced,then subcloned into pGEX-4T-1 to form prokaryotic expression vector pGEX-4T-1-hIL-23R.After transforming with pGEX-4T-1-hIL-23R,the E.coli BL21(DE3) cells were induced by Isopropanol-β-D-galactoside(IPTG).Western blot was performed to identify the expression of hIL-23R fusion protein.Results:The full length of hIL-23R cDNA was cloned,constructed prokaryotic expression vector pGEX-4T-1-hIL-23R and expressed in E.coli.The recombinant protein mainly exist as inclusion bodies and the Mr of fusion protein was 97KDa.Conclusion:The prokaryotic expression vector pGEX-4T-1-hIL-23R is successfully constructed and the fusion protein is expressed in E.coli BL21(DE3) cells.