重组犬α6干扰素的酵母表达及其抗病毒活性评价

doi:10.16431/j.cnki.1671-7236.2019.07.032

Abstract

Abstract:

The aim of the experiment was to construct the expression system of canine interferon alpha 6 (CaIFN-α6) in Pichia pastoris,optimize and screen the expression system in order to obtain recombinant canine interferon alpha 6 with high activity.According to the sequence of CaIFN-α6 gene and the codon preference of Pichia pastoris,the whole gene sequence of CaIFN-α6 was optimized and synthesized.The recombinant expression plasmid of pPICZαA-CaIFN-α6 was constructed using Xho Ⅰ and Not Ⅰ and linked to the vector pPICZαA.The recombinant expression plasmid was transformed into E.coli DH5α competent cells.The plasmid pPICZαA-CaIFN-α6 was extracted and linearized.The recombinant strain was prepared by electroporation into yeast competent cell X33.The recombinant CaIFN-α6 was purified by methanol induction,supernatant collection and ultrafiltration concentration.The concentration of purified CaIFN-α6 protein was 1.5 mg/mL by BCA.Western blotting analysis showed that CaIFN-α6 protein had good reactivity.SDS-PAGE showed that the purity of CaIFN-α6 protein was above 95%.The titer of CaIFN-α6 protein was 2.37×10⁷ IU/mL by MDCK/VSV detection and the specific activity was 1.58×10⁷ IU/mg.The results showed that CaIFN-α6 was successfully expressed in Pichia pastoris pPICZαA expression vector system with high biological activity,which provided a good support for the clinical prevention and treatment of canine viral disease in the later stage.

Key words: canine interferon alpha 6(CaIFN-α6); eukaryotic expression; purification; antiviral activity

CLC Number:

S852.4

SONG Tianqi, HOU Shaohua, GUO Xiaoyu, XIN Ting, JIANG Yitong, YUAN Weifeng, ZHU Hongfei, JIA Hong. Yeast Expression of Recombinant Canine Alpha Interferon 6 and Evaluation of Their Antiviral Activities[J]. , 2019, 46(7): 2144-2150.

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Yeast Expression of Recombinant Canine Alpha Interferon 6 and Evaluation of Their Antiviral Activities

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