嵌合OVA多肽的水貂肠炎病毒VP2病毒样颗粒的表达及鉴定  

Expression and identification of mink enteritis virus VLPs with chimeric OVA polypeptide

作　　者：吕甜甜 郭晓芹 曹海旭 朱言柱[1] 宋显晶[2] 张蕾[1] LÜTian-tian;GUO Xiao-qin;CAO Hai-xu;ZHU Yan-zhu;SONG Xian-jing;ZHANG Lei(Key Laboratory of Special Animal Epidemic Disease,Ministry of Agriculture and Rural Afars/Jilin Pror incial Biological Engineering Research Center for Special Economic Animals/Institute of Special Animal and Plant Sciences,Chinese Academy of Agricultural Sciences,Changchun 130122,China;The Second Hospital of Jilin University,Changchun 130041,China)

机构地区：[1]中国农业科学院特产研究所农业农村部经济动物疫病重点实验室吉林省特种经济动物用生物制品工程研究中心,吉林长春130122 [2]吉林大学第二医院,吉林长春130041

出　　处：《中国兽医科学》2022年第6期712-719,共8页Chinese Veterinary Science

基　　金：中国农业科学院科技创新工程项目(CAAS-ASTIP-2016-ISAPS)。

摘　　要：为探究水貂肠炎病毒(MEV)VP2病毒样颗粒(VLPs)作为嵌合载体携带外源抗原的能力,通过PCR技术将鸡卵清蛋白(OVA)第257~264位氨基酸短肽(SIINFEKL)插入到MEV VP2的N末端获得OVA_(257-264)-VP2,通过克隆重组和蓝白斑筛选分别获得重组质粒Bacmid-VP2、Bacmid-OVA_(257-264)-VP2,转染到昆虫细胞表达获得VP2、OVA_(257-264)-VP2蛋白的重组杆状病毒;通过间接免疫荧光和Western-blot分析蛋白反应原性,透射电镜观察VLPs形态,血凝试验检测其血凝特性。间接免疫荧光和Western-blot结果表明,重组蛋白VP2和OVA_(257-264)-VP2都具有良好的反应原性。透射电镜观察结果表明,VP2与OVA_(257-264)-VP2都可以形成病毒样粒子,颗粒直径均约为22 nm。血凝试验结果表明,重组蛋白VP2和OVA_(257-264)-VP2与天然病毒的凝集特性相同,均可达到1∶16 384。结论:在MEV VP2的N端插入OVA257-264,不影响VP2病毒样颗粒的组装,VP2和外源多肽都保持有良好的反应原性,MEV VP2可以作为疫苗载体递呈外源抗原。To explore the ability of mink enteritis virus(MEV)VP2 virus-Like particles(VLPs),as chimeric carriers,a short peptide of amino acids 257-264(SIINFEKL)of chicken ovalbumin(OVA)was connected to the N-terminus of MEV VP2 by PCR.The recombinant bacmid-VP2 and Bacmid-OVA_(257-264)-VP2 were cloned,genetic recombination and screened by the blue-white spot experiment,and then transfected into insect cells to obtain recombinant baculoviruses expressing VP2 and OVA_(257-264)-VP2 proteins.The protein reactogenicity was analyzed by indirect immunofluorescence assay and Western-blot,the morphology of VLPs was observed by transmission electron microscope,and the hemagglutination characteristics were detected by hemagglutination test.Indirect immunofluorescence assay and Western-blot showed the recombinant proteins had good reactogenicity.Transmission electron microscopy results showed that VP2 and OVA_(257-264)-VP2 can assemble VLPs with diameter 22 nm.The hemagglutination titer of VP2 and OVA_(257-264)-VP2 VLPs both reached 1∶16384.In conclusion,the OVA_(257-264)insertion into the N-terminus of MEV VP2 does not affect the assembling of the vector’s VLPs.VP2 and OVA peptides both have good reactogenicity,MEV VP2 can be used as a vaccine carrier to present foreign antigens.

关 键 词：病毒样颗粒 水貂肠炎病毒 VP2基因 OVA 

分 类 号：S852.659.2[农业科学—基础兽医学]