鸡β干扰素在毕赤酵母中的分泌表达及其抗病毒活性测定

doi:10.16431/j.cnki.1671-7236.2017.07.043

摘要/Abstract

摘要：

为研究鸡β干扰素（chicken interferon-β，ChIFNβ）蛋白在毕赤酵母中的表达及其抗病毒活性，将克隆的ChIFNβ基因亚克隆至酵母表达载体pPICZα-A中，构建重组质粒pPIC-ChIFNβ。将鉴定正确的质粒pPIC-ChIFNβ线性化，通过电转化整合到毕赤酵母菌株X-33基因组中，得到阳性菌株。经1.0%甲醇连续诱导4 d，表达产物利用SDS-PAGE电泳、细胞病变抑制法等进行分析。ChIFNβ在酵母中获得了分泌型表达，表达产物分子质量约为17 ku。表达产物以细胞病变抑制法检测其活性，效价为10^7.5 U/mL；在鸡胚中能有效抑制禽流感病毒H9N2的增殖。动物攻毒保护试验结果显示，表达的蛋白可提供70%的免疫保护。表达的ChIFNβ蛋白具有较好的抗病毒活性，本研究可为ChIFNβ的应用提供重要参考。

关键词: 鸡β 干扰素; 分泌表达; 抗病毒活性

Abstract:

In order to study the expression of chicken interferon-β (ChIFNβ) in Pichia pastoris and its antiviral effect,ChIFNβ gene was subcloned into pPICZα-A vector and constructed pPIC-ChIFNβ recombinant plasmid. After linearized by Sac Ⅰ, the plasmid was transferred into Pichia pastoris X-33 strain. The recombinant strain was induced by 1.0% methanol for 4 d,the product of recombinant ChIFNβ was detected by SDS-PAGE,the antiviral activity was determined by classical cell pathological effect inhibition assay. The protein weight was approximate 17 ku and the recombinant protein was secreted effectively in Pichia pastoris. Potency of interferon was 10^7.5 U/mL. The expressed protein could also inhibit H9N2 avian influenza virus proliferation in chick embryo. Animal experiment suggested that recombinant protein could provide 70% immune protection for aviadenovirus. The results showed that the recombinant ChIFNβ had good antivirus activity,it would be reference for clinical application of ChIFNβ.

Key words: chicken interferon-β; secreted expression; antiviral activity

中图分类号:

TQ929⁺.1

刘新文, 王秀丽, 郭东春, 郭伟伟, 邹敏, 范根成. 鸡β干扰素在毕赤酵母中的分泌表达及其抗病毒活性测定[J]. 《中国畜牧兽医》, 2017, 44(7): 2203-2208.

LIU Xin-wen, WANG Xiu-li, GUO Dong-chun, GUO Wei-wei, ZOU Min, FAN Gen-cheng. Expression of Chicken Interferon-β in Pichia pastoris and Its Antiviral Effect[J]. , 2017, 44(7): 2203-2208.

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