Clone And Expression, Purification And Immunoactivity Assay Of Human Interleukin 21

Objective To get the human interleukin 21 encoded gene from human tonsil cells and construct an engineering bacteria which can express human interleukin 21 mature N terminus fusion protein. Do the purification and bioactivity assay about the recombinant protein.Methods Human interleukin 21 gene fragment was required from human tonsil cDNA by means of PCR. Primers including BamH I and Sal I digestion sites were designed.The encoded gene for the mature N-terminus of IL-21 was then inserted into prokaryotic expressing vector pGEX-4T-2 and transformed into E.coli DH5 a . After induced by 1PTG, the expressed product was purified by affinity chromatography using GST Fusion Protein Purification bead and its immunoactivity was analyzed by Western blotting test. Results PCR identification and digestion by BamHI and SalI of recombinant plasmid showed there was corresponding electrophoresis products same to expectation. After induced by IPTG, transformed E.coli DH5 a expressed the right molecular weight protein as well. The interest protein was about 10 percent in the whole bacteria protein. Western blotting test revealed that the recombinant protein had reaction with IL-2 monoclonal antibody.Conclusion Constructed the vector pGEX-4T-2/IL-21 and the transformed engineering bacteria E.coli DH5 a which can express human interleukin 21 mature N terminus fusion protein. Purification and Western blotting test showed the recombinant IL-21, with similarity to IL-2 in structure, may be a new drug in the clinic to regulate the immunological function.