重组猪干扰素β在CHO细胞中的表达及其抗口蹄疫病毒活性的研究  

Expression of recombinant porcine interferon β in CHO cells and study on its activity against foot-and-mouth disease virus

作　　者：王凌云 郝荣增 茹毅 常辉 杨洋[2] 李亚军 张丽伟 卢炳州 刘艳红 龚真莉 井波[1] 齐萌[1] 郑海学 WANG Ling-yun;HAO Rong-zeng;RU Yi;CHANG Hui;YANG Yang;LI Ya-jun;ZHANG Li-wei;LU Bing-zhou;LIU Yan-hong;GONG Zhen-li;JING Bo;QI Meng;ZHENG Hai-xue(College of Animal Science and Technology,Tarim University,Aral 843300,China;National Foot-and-Mouth Disease Reference Laboratory/State Key Laboratory of Veterinary Etiological Biology/Lanzhou Veterinary Research Institute,Chinese Academy of Agricultural Sciences,Lanzhou 730046,China)

机构地区：[1]塔里木大学动物科学与技术学院,新疆阿拉尔843300 [2]中国农业科学院、兰州兽医研究所、家畜疫病病原生物学国家重点实验室国家口蹄疫参考实验室,甘肃兰州730046

出　　处：《中国兽医科学》2023年第2期143-149,共7页Chinese Veterinary Science

基　　金：甘肃省科技重大专项(21ZD3NA001);中国农业科学院所级重点任务(CAAS-ASTIP-JBGS-20210401);中国农业科学院兰州兽医研究所所级基本科研业务费项目(1610312021013)。

摘　　要：构建猪IFN-β重组真核表达质粒pcDNA3.1-PoIFN-β,转染悬浮培养的CHO细胞,进行上清分泌表达;通过CHO细胞表达系统,成功分泌表达和制备纯化的rPoIFN-β。通过CCK-8试验,分析rPoIFN-β对细胞的毒性,结果显示纯化的rPoIFN-β对细胞无明显毒性;采用VSV/PK-15系统检测rPoIFN-β抗病毒活性效价,结果显示其抗病毒活性效价为4.54×10^(7)U/mg;制备的rPoIFN-β可显著抑制O型和A型FMDV在PK-15细胞的复制,表现出较强的抗病毒活性;通过分析rPoIFN-β对细胞ISGs的诱导作用,发现rPoIFN-β能够显著刺激12种ISGs的表达。研究结果表明,本试验制备纯化的rPoIFN-β具有较低的细胞毒性和较高的抗病毒活性效价,为口蹄疫防控制剂的开发和研究提供了基础数据。In this study,the porcine IFN-β recombinant eukaryotic expression plasmid pc DNA3.1-Po IFN-β was constructed,and the suspension cultured CHO cells were transfected for supernatant secretion expression,and the purified r Po IFN-β was successfully secreted and prepared by the CHO cell expression system.Through the CHO cell expression system,the purified r Po IFN-β was successfully secreted and prepared.The cytotoxicity of r Po IFN-β was analyzed by CCK-8 test,we found that the purified r Po IFN-β showed no apparent cytotoxicity to PK-15 cells.Antiviral activity assay for r Po IFN-β was performed and evaluated by standard procedures in VSV/PK-15(virus/cell) test system,the biological activity of r Po IFN-β was 4.54×10^(7)U/mg.The prepared r Po IFN-β significantly inhibited virus replication of the two serotype of FMDV and showed strong antiviral activity.By analyzing the induction effect of r Po IFN-β on cellular ISGs,and r Po IFN-β greatly induced the 12 ISGs production in PK-15 cells.The results showed that the purified r Po IFN-β prepared in this experiment has low cytotoxicity and high antiviral activity titer,which provides basic data for the development and research of FMD prevention and control agents.

关 键 词：猪IFN-β CHO细胞 蛋白表达 抗病毒活性 口蹄疫病毒 

分 类 号：S852.659.6[农业科学—基础兽医学]