表达及抗病毒活性比较-生物谷.docVIP

猪I型与II型干扰素的克隆、表达及抗病毒活性比较 陈雪梅, 薛清华, 祝荣格, 付显华, 杨利敏, 孙蕾, 刘文军 中国科学院病原微生物及免疫学重点实验室 中国科学院微生物研究所分子病毒中心, 北京 100101 摘 要: 干扰素(IFN)是由多种细胞受病毒感染或其他生物诱导剂刺激而产生的天然蛋白质, 主要功能为抗病毒增殖、调节免疫反应和激活免疫细胞等。本研究克隆并测序了猪干扰素(PoIFN)?、?、???及?基因。构建原核表达载体pET-His/PoIFN-?、pET-His/PoIFN-?、pET-His/PoIFN-???和pET-His/PoIFN-?, 转化大肠杆菌Rosetta(DE3)进行表达, 经纯化、复性得到具有生物学活性的蛋白。用细胞病变抑制法在Marc-145/PRRSV、Marc-145/VSV、PK-15/VSV、Vero/VSV、MDBK/VSV系统上进行抗病毒活性测定, 结果表明猪?和??融合干扰素有较为显著的抗病毒活性, 抗PRRSV活性高达108 U/mg; 猪?干扰素活性效价约为?干扰素的1/2到1/3; 猪?干扰素几乎未检测到抗病毒活性, 需进一步验证。本研究对干扰素在抗病毒、提高机体免疫方面的应用提供了理论依据。 关键词: 猪干扰素, 抗病毒活性 Comparison of antiviral activities of porcine interferon type I and type II Xuemei Chen, Qinghua Xue, Rongge Zhu, Xianhua Fu, Limin Yang, Lei Sun, and Wenjun Liu Chinese Academy of Sciences Key Laboratory of Pathogenic Microbiology and Immunology, Center for Molecular Virology, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China Abstract: Interferons (IFNs) are natural proteins produced by wide variety of cells in response to viral infection or other biological inducers, and they execute diversified functions as antiviral defense, immune activation and cell growth regulation. Four genes encoding porcine interferons (PoIFN), PoIFN-α, PoIFN-γ, PoIFN-αγ or PoIFN-ω, were cloned and sequenced. The four types of porcine interferon genes were subcloned into the pET-His vector, and expressed in Escherichia coli Rosetta (DE3). The recombinant products were purified and renaturalized from inclusion bodies to obtain a native state of well biological activity. Antiviral activity assays for porcine interferons were performed and evaluated by standard procedures in following cell/virus test systems: Marc-145/PRRSV, Marc-145/VSV, PK-15/VSV, Vero/VSV or MDBK/VSV. The data showed that both PoIFN-α and PoIFN-αγ demonstrated significant antiviral activities, and the titer of them against PRRSV was up to 108 U/mg. PoIFN-γ had approximately half or one-third