非洲猪瘟病毒p72蛋白的原核表达及其单克隆抗体的制备  

African Swine Fever Virus p72 Protein Expression and Production of Monoclonal Antibodies

作　　者：靳家鑫 路闻龙 赵旭阳 张帅[1] 杜永坤 张改平[1] 孙爱军 庄国庆 JIN Jia-xin;LU Wen-long;ZHAO Xu-yang;ZHANG Shuai;DU Yong-kun;ZHANG Gai-ping;SUN Ai-jun;ZHUANG Guo-qing(International Joint Research Center of National Animal Immunology,College of Veterinary Medicine,Henan Agricultural University,Zhengzhou,Henan,450046,China)

机构地区：[1]河南农业大学动物医学院,国家动物免疫学国际联合研究中心,河南郑州450046

出　　处：《动物医学进展》2023年第3期38-42,共5页Progress In Veterinary Medicine

基　　金：国家自然科学基金非洲猪瘟重点专项(31941016和31941001);“十四五”国家重点研发计划项目(2021YFD1801404);2022年度河南省重点研发与推广专项(222102110373);河南农业大学高层次“拔尖人才”项目(30500690);河南省高等学校重点科研项目(21A230010)。

摘　　要：非洲猪瘟病毒(African swine fever virus, ASFV)B646L基因编码的结构蛋白p72是高度保守的抗原,常用作ASFV检测方法的建立。论文将截短的B646L基因连接在pET28b载体,重组质粒命名为pET28b-B646L。将重组质粒转化至大肠埃希氏菌BL21(DE3)感受态细胞,经异丙基-β-D-硫代半乳糖苷(IPTG)诱导表达,纯化p72蛋白并免疫Balb/c小鼠制备单克隆抗体。Western blot和IFA验证结果表明,成功获得1株IgG1亚型的ASFV p72特异性单克隆抗体,可为建立ASFV血清学检测方法提供参考。P72,a highly conserved structural protein encoded by African swine fever virus(ASFV)B646L gene,is usually used as antigen for ASFV detection method development.In this study,we constructed recombinant plasmid named as pET28b-B646L by ligating truncated B646L gene and the pET28b vector.Then,the pET28b-B646L recombinant plasmid was transformed into E.coli BL21(DE3)competent cells.The truncated p72 protein was purified and identified by SDS-PAGE and Western blot assays after induction by isopropyl-β-D-thiopental(IPTG),and was used to immunize Balb/c mice to produce monoclonal antibodies.The results showed that we successfully obtained a IgGl subtype of p72 specific monoclonal antibody,which is confirmed by Western blot and IFA assays.Thus,this study provides the basis for establishing an ASFV-associated serum detection method.

关 键 词：非洲猪瘟病毒 p72蛋白 原核表达 单克隆抗体 

分 类 号：S852.651[农业科学—基础兽医学]