Study On High Expression Of The Chemical Synthetic Human Interferon α-2b Gene In E.coli

The chemical synthetic human interferon alpha 2b (rhIFN α -2b) gene was amplified from pUC19/rhIFN α -2b by polymerase chain reaction and recombined into expression vector pBV220. The obtained pBV220/rhIFN α -2b was transformed into Escherichia coli DH5 α . After temperature induction, the total protein was analyzed by 15% SDS-PAGE and a 19KD specific protein band was observed. The target protein accounts for about 36% of the total protein. By western blotting analysis with IFN α antibody, the target protein was proved to be the recombinant human IFN α -2b. The expression level of chemical synthetic human IFN α -2b gene is 123.5 times more than that of wild human IFN α -2b gene.The expression level of target protein was compared at different induction temperatures and in different culture medium. Firstly the target protein accounts for about 35% of total protein, when the transformed E.coli DH5 α was cultured in LB at 35℃ until the OD600 reached to 0.4, then for 50min at a restrictive temperature 30℃, finally induced for 5h at an induction temperature 42℃. Secondly the expected protein accounts for about 36% of total protein at following condition: cultured at 30℃ for 4h, then induced at an induction temperature 42℃ for 3.5h in a specific culture medium. The ingredients of the specific culture medium are 1% tryptone, 0.6% glucose, 0.5% yeast extract, 0.6%Na2HPO4, 0.2%KH2PO4, 0.08%NH4Cl, 0.4%NaCl, 0.001% CaCl2, 0.02%MgSO4 (pH6.9).