PRRSV GP2a胞外区的原核表达及免疫原性分析  

Prokaryotic expression and immunogenicity analysis of ectodomain of GP2a of PRRSV

作　　者：扣莉云 张改平 史西保[3] 张小转[3] 车志芬 周景明 祁艳华 王爱萍 KOU Li-yun;ZHANG Gai-ping;SHI Xi-bao;ZHANG Xiao-zhuan;CHE Zhi-fen;ZHOU Jing-ming;QI Yan-hua;WANG Ai-ping(Department of Bioengi neering,Zhengzhou Universi-ty,Zhengzhou 450001,China;College of Veterinary Medicine and Animal Science,Henan Ag-ricultural University,Zhengzhou 450002,China;College of Life Sciences,Henan Normal Uni-versity,Xinx iang,Henan 453007,China)

机构地区：[1]郑州大学生命科学学院,河南郑州450001 [2]河南农业大学牧医工程学院,河南郑州450002 [3]河南师范大学生命科学学院,河南新乡453007

出　　处：《中国兽医学报》2020年第2期231-236,共6页Chinese Journal of Veterinary Science

基　　金：国家自然科学基金资助项目(31472177);河南省高等学校重点科研基因资助项目(17A180006)。

摘　　要：为获得有活性的猪繁殖与呼吸综合征病毒(porcine reproductive and respiratory syndrome virus,PRRSV)BJ-4株GP2a胞外区蛋白,以真核质粒pcDNA3.1-ORF2为模板,设计针对GP2a胞外区(41-208aa)的特异性引物,通过PCR等方法,获得重组表达质粒pET-32a-eORF2,转化宿主菌Rosetta(DE3),并用1 mmol/L异丙基硫代半乳糖苷(IPTG)进行诱导。SDS-PAGE结果表明,重组的GP2a胞外区蛋白以包涵体形式表达,相对分子质量约36000;将包涵体纯化、复性后,以50μg/只的剂量免疫BALB/c小鼠,三免后,抗体效价达1∶102400;这表明GP2a胞外区成功表达,且具有良好的免疫原性。In order to obtain the activated glycoprotein 2 a(GP2 a) of porcine reproductive and respiratory syndrome virus(PRRSV),a pair of primers were designed and the DNA sequences of the ectodomain of GP2 a were amplified from the eukaryotic plasmid pcDNA3.1-ORF2 and were restructured in the prokaryotic expression vector pET-32 a(pET-32 a-eORF2).Then the plasmid was transferred into the E.coli Rosetta(DE3).The results of SDS-PAGE showed that the recombinant protein about 36 000 was successfully expressed in form of inclusion body by the induction of the 1 mmol/L IPTG.At the same time,through the purification,denaturation and renaturation of the recombinant protein.The each BALB/c mouse was injected subcutaneously for 50 μg recombinant protein,after three immunization,the antibody titers were up to 1∶102 400.In conclusion,the ectodomain of GP2 a protein was expressed successfully and had well immunogenicity.

关 键 词：PRRSV GP2a胞外区 原核表达 蛋白纯化 

分 类 号：S852.65[农业科学—基础兽医学]