中国兽医科学2024，Vol.54Issue(11)：1473-1481,9.DOI:10.16656/j.issn.1673-4696.2024.0194
非洲猪瘟病毒I215L蛋白的原核表达及其多克隆抗体的制备
Prokaryotic expression and polyclonal antibody preparation of African swine fever virus I215L protein
摘要
Abstract
This study focused on the ubiquitin-conjugating enzyme(UBCv1)encoded by the African swine fever virus I215 L gene,expressed its prokaryotic system and prepared polyclonal antibodies.Ac-cording to the sequence of the ASFV-CN/GS/2018 strain isolated by the team,the I215L gene was intro-duced into the prokaryotic expression vector to form a recombinant plasmid(pET-28a-I215L),which was introduced into E.coli BL21(DE3)competent cells.After a large number of induction and expression,the recombinant protein was purified by gel cutting method.The purified I215L protein was mixed with adju-vant to immunize animals for many times to obtain mouse/rabbit polyclonal antibody against I215L pro-tein.The antibody titer was determined by enzyme-linked immunosorbent assay(ELISA).The mouse an-ti-I215 L polyclonal antibody titer reached more than 1∶64 000,and the rabbit anti-I215 L polyclonal antibody titer reached 1∶512 000.The results of indirect immunofluorescence(IFA)and Western-blot showed that the polyclonal antibody could specifically recognize the expression of I215L protein after PAMs infected with ASFV,and the effect was good.In this study,the African swine fever virus I215L pro-tein was expressed and purified in vitro,and the polyclonal antibody of mouse/rabbit anti-I215L pro-tein was successfully obtained,which laid a foundation for the structural analysis,functional study and virus-related pathogenesis of ASFV I215L protein.关键词
非洲猪瘟病毒/I215L蛋白/原核表达/多克隆抗体Key words
African swine fever virus/I215L protein/prokaryotic expression/polyclonal antibody分类
农业科技引用本文复制引用
杜煜,孙茂文,曹丽艳,孔祥雨,李想通,张宇,王琦,郑海学..非洲猪瘟病毒I215L蛋白的原核表达及其多克隆抗体的制备[J].中国兽医科学,2024,54(11):1473-1481,9.基金项目
国家重点研发计划项目(2021YFD1801300,2021YFD1800100) （2021YFD1801300,2021YFD1800100）
甘肃省科技重大专项(22ZD6NA001,22ZD6NA012) （22ZD6NA001,22ZD6NA012）
广东省"十四五"农业科技创新专项(2023SDZG02) （2023SDZG02）
四川省杰出青年基金项目(21JCQN0175) （21JCQN0175）
