¹Wuhu Overseas Students Pioneer Park, Wuhu, Anhui Province, 241000, CHINA
²Department of Microbiology, Anhui Medical University, Hefei, Anhui Province, 230032, CHINA
³Anhui JiuChuan Biotech Co., Ltd, Wuhu, Anhui Province, 241007, CHINA
⁴Department of Pathology & Cell Biology, Columbia University, New York, 10032, USA DOI : 10.9775/kvfd.2017.17372 Herein, we reported an Escherichia coli-based expression and purification method of recombinant porcine interferon alpha (rPoIFN-α). PoIFN-α coding sequence was cloned into pMD18-T vector and then subcloned into pET-32a (+) vector using standard recombinant DNA techniques and the resulting plasmid was transformed into BL21(DE3) competent cells. After induction with isopropyl-β-D-1-thiogalactopyranoside (IPTG), rPoIFN-α was purified from the supernatant of the bacteria lysate using a simple two-step chromatography process consisting of a Ni2+ affinity chromatography and a DEAE anion exchange chromatography. rPoIFN-α was purified to >95% homogeneity with a yield of 48 mg/L of culture. It has isoelectic point of 6.09 and bacterial endotoxin was less than 1 EU/mg. N-terminal amino acid sequence and the peptide map digested by trypsin provided additional evidence for the authenticity of rPoIFN-α. The biological activity of rPoIFN-α was 1.1×106 IU/ mL in HEp-2/ Vesicular Stomatitis Virus (VSV) titration system and its specific activity reached to 1.0×106 IU/mg. In conclusion, we obtained high-level expression of a soluble form of bioactive rPoIFN-α by using pET-32a (+) prokaryotic expression system. Keywords : Soluble expression, Protein purification, Quality control, Porcine interferon-α, Vesicular Stomatitis Virus (VSV)