摘要
为了研究猪流行性腹泻病毒（PEDV）S1蛋白中和表位区S1D的免疫原性,根据PEDV河南株（Gen Bank：YK649107）S1基因序列,设计1对特异性引物,PCR扩增S1D片段,克隆到p ET-28a（＋）中,构建原核表达载体,经优化的IPTG浓度诱导后进行SDS-PAGE分析,利用His标签的特性重组蛋白进行纯化及Western blot分析。结果显示：中和表位S1D片段在p ET-28a（＋）中高效表达;Western blot证明重组蛋白可以与PEDV阳性血清产生特异性反应;用纯化的蛋白免疫新西兰白兔,抗体水平升高,表明重组蛋白具有良好的免疫原性。
To investigate the immunogenicity of the neutralizing epitope region S1 D of S1 protein in porcine epidemic diarrhea virus（ PEDV）,a pair of primers was designed according to the nucleotide sequence of the S1 gene in the PEDV Henan strain（ Gen Bank：YK649107）. S1 D was amplified by PCR and cloned into the p ET-28a（ ＋） vector for constructing a prokaryotic expression vector. SDS-PAGE,purification of recombinant protein and Western blot were performed to analyze the expression of recombinant protein after induction of the optimal IPTG concentration. The results showed that the neutralizing epitope S1 D was efficiently expressed in the p ET-28a（ ＋） by SDS-PAGE analysis. The recombinant protein was purified by the characteristics of His tag and could produce specific positive binding reaction with PEDV positive serum,as confirmed by Western blot. The purified recombinant protein was used to immunize New Zealand white rabbits,and subsequently increases in antibody levels were observed,suggesting that the recombinant protein had good immunogenicity.
作者
张鸿鑫
韩昊莹
侯华琳
赵宇
郑兰兰
杨明凡
陈红英
ZHANG Hongxin;HAN Haoying;HOU Hualin;ZHAO Yu;ZHENG Lanlan;YANG Mingfan;CHEN Hongying(College of Animal Science and Veterinary Medicine,Henan Agricultural University,Zhengzhou 450002,China;Zhengzhou City Key Laboratory of Major Pig Disease Prevention and Control,Henan Agricultural University,Zhengzhou 450002,China)
出处
《畜牧与兽医》
北大核心
2018年第10期110-114,共5页
Animal Husbandry & Veterinary Medicine
基金
河南省产学研合作计划项目(152107000003)
河南省基础与前沿技术研究计划项目(142300410156)
