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猪流行性腹泻病毒COE基因的原核表达和免疫原性分析
杨榕,顾超逸,李郁,魏建忠,孙裴,杨勇
0
(安徽农业大学动物科技学院,合肥 230036;安徽安泰农业集团,合肥 230036)
摘要:
为研究猪流行性腹泻病毒中COE基因的原核表达情况及免疫原性,从感染猪流行性腹泻病毒的猪体内采集肠内容物、肠段及肠系膜淋巴结,参照GenBank中所收录的猪流行性腹泻病毒(PEDV)CV777株的COE基因序列设计1对特异性引物,通过RT-PCR扩增出COE基因,克隆到原核表达载体pET-32α上,并构建重组表达质粒pET-32α-COE,在大肠杆菌Rosetta中进行表达,并进行Western blot检测。结果显示,重组蛋白获得了高效的表达,并能与多抗血清发生特异的免疫印迹反应。COE基因是猪流行性腹泻病毒的中和抗原位点,具有良好的免疫学活性。
关键词:  猪流行性腹泻病毒  COE基因  原核表达  免疫印迹
DOI:
基金项目:现代农业产业技术体系建设专项资金(CARS-36-生猪)和安徽省生猪产业技术体系资金共同资助。
Prokaryotic expression and immunogenicity analysis of COE gene of porcine epidemic diarrhea virus
YANG Rong,GU Chao-yi,LI Yu,WEI Jian-zhong,SUN Pei,YANG Yong
(School of Animal Science and Technology, Anhui Agricultural University, Hefei 230036;Antai Agricultural Group of Anhui, Hefei 230036)
Abstract:
To study the prokaryotic expression and immunogenicity in the COE gene of porcine epidemic diarrhea virus, we collected intestinal contents, bowel and mesenteric lymph nodes in piglets from the pig which infected porcine epidemic diarrhea virus (PEDV). The specific primers were designed by the sequence of porcine epidemic diarrhea virus (PEDV) CV777 strain COE gene reported in GenBank. The purpose gene was amplified by RT-PCR, and then cloned into prokaryotic expression vector pET-32α, to construct a recombinant expression vector pET-32α-COE. The gene was expressed by transformation into in E.coli Rosetta, and was detected by Western blot. The result showed that the recombinant protein was highly expressed, and could react with polyclonal antibody. COE gene is neutralization antigenic sites of porcine epidemic diarrhea virus, and has a good immune activity.
Key words:  porcine epidemic diarrhea virus  COE gene  prokaryotic expression  Western blot

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