Expression，Purification of Recombinant Luxi Yellow Cattle IFN-α Fusion Protein and Its Antiviral Activities

Abstract:

Interferon α gene was cloned from genomic DNA of Chinese Luxi yellow cattle by PCR, and the PCR product was inserted into vector pET32a(+) to make a recombinant plasmid pET32a(+)/BoIFN-α. The expression of BoIFN-α in Escherichia coli was induced by addition of IPTG. Sequence analysis showed that the Chinese Luxi yellow cattle IFN-α gene is composed of 498 nucleotides, encoding a mature polypeptide of 166 amino acids. Compared with other BoIFN-α subtypes, it shares the highest identity of 97.6% to the C-subtype. SDS-PAGE results showed that recombinant proteins were expressed in inclusion bodies in Escherichia coli with molecular weight of 40kD and the recombinant proteins accounted for 26.7% of the whole proteins.The expressed product was purified by affinity chromatography with immobilized nickel chelating NTA (Ni-NTA) and its antiviral activities were tested on MDBK/VSV cell system. Its antiviral activities were 5×10⁵u/mg on MDBK/VSV cell system. The results showed that the expression plasmid was successfully constructed and BoIFN-α C2 protein was expressed in Escherichia coli. Moreover the purification had good effects on antiviral activities.

Key words:yellow cattle， interferon-α gene， expression， antiviral activities