猪源IRAV的克隆、表达及单克隆抗体的制备  

Cloning and Expression of Porcine Interferon-Regulated Antiviral Gene and Preparation of Monoclonal Antibodies

作　　者：王桦 焦亚娟 孔宁[1,2] 孙大格 董苏洁 陈晓勇 翟焕杰 童武 郑浩[1,2] 于海 虞凌雪[1,2] 童光志 单同领[1,2] WANG Hua;JIAO Yajuan;KONG Ning;SUN Dage;DONG Sujie;CHEN Xiaoyong;ZHAI Huanjie;TONG Wu;ZHENG Hao;YU Hai;YU Lingxue;TONG Guangzhi;SHAN Tongling(Shanghai Veterinary Research Institute,CAAS,Shanghai 200241,China;Jiangsu Co-Innovation Center for the Prevention and Control of Important Animal Infectious Disease and Zoonoses,Yangzhou University,Yangzhou 225009,China)

机构地区：[1]中国农业科学院上海兽医研究所,上海200241 [2]江苏高校动物重要疫病与人兽共患病防控协同创新中心,扬州225009

出　　处：《中国动物传染病学报》2022年第5期126-134,共9页Chinese Journal of Animal Infectious Diseases

基　　金：国家自然科学基金(31872478,32102665,31760741)。

摘　　要：干扰素调节的抗病毒基因(IRAV)是近年来被发现的一种具有抗病毒活性的新型干扰素刺激基因,其对登革热病毒的增殖具有良好的抑制作用,但目前鲜有猪源IRAV基因的研究。本研究,从PK-15细胞中克隆了IRAV全长的cDNA。猪源IRAV cDNA全长为1241 bp,其中包含858 bp的开放阅读框,编码一个大小为285个氨基酸的多肽,定位于细胞质。将猪源IRAV蛋白在BL21(DE3)中诱导表达,经纯化后免疫雌性BALB/c小鼠,最终获得5株分泌猪源IRAV单克隆抗体(mAb)的杂交瘤细胞,分别命名为2B10、2G12、2H1、5A8和2C5。将获得的分泌性抗IRAV单克隆抗体经Western blot和间接免疫荧光试验分析后,确定该抗体可以与PK-15细胞中过表达的猪源IRAV发生特异性反应。本研究成功制备针对猪源IRAV的单克隆抗体,为后续深入研究猪源IRAV的功能奠定了基础。IRAV(interferon-regulated antiviral gene)has been discovered as a novel interferon-stimulated gene with antiviral activity.However,little is known about porcine IRAV.In this study,we cloned the full-length IRAV complementary DNA(cDNA)from porcine kidney cells.The porcine IRAV cDNA was of 1241 bp with an open reading frame of 858 bp,encoding a polypeptide of 285 amino acids,which localized to the cytoplasm.The porcine IRAV protein was expressed in E.coli BL21(DE3),purifi ed and used to immunize BALB/c mice for preparation of monoclonal antibodies(mAbs).As a result,5 hybridomas(2B10,2G12,2H1,5A8 and 2C5)were found to secrete anti-IRAV mAbs.These mAbs specifi cally reacted with the overexpressed porcine IRAV protein in PK15 cells as demonstrated in Western blot and indirect immunofl uorescence assay.Taken together,the availability of mAbs against porcine IRAV provided a valuable tool to study the biological function of IRAV in the future.

关 键 词：猪源IRAV 系统发育分析 表达 定位 单克隆抗体 

分 类 号：S852.4[农业科学—基础兽医学]