摘要
为了获得具有活性的鸡传染性贫血病毒(CIAV)的衣壳蛋白,给疫苗的研制和诊断试剂的开发提供新的思路。根据已发表的CIAV VP 1编码基因的序列并结合大肠杆菌偏爱密码子分析,合成了VP 1(去掉N′端80个氨基酸)的编码序列,并利用大肠杆菌进行表达,SDS-PAGE结果表明,表达产物主要为不可溶的包涵体蛋白。进一步利用伴侣蛋白使VP1蛋白的可溶性表达产量得到提高,并确定了诱导重组表达VP1蛋白的最佳条件:16℃、0.08 mmol/L IPTG诱导。利用Ni 2+树脂对重组VP1蛋白进行纯化,然后利用间接ELISA方法进行检测,发现该重组蛋白可与CIAV阳性血清发生反应。本研究成功利用伴侣蛋白促进VP1蛋白的可溶性表达,而且表达的VP1蛋白有良好的免疫反应,具有开发为新型亚单位疫苗和诊断抗原的潜力。
According to published CIAV capsid protein(VP1)coding gene sequence,which is used for codon preference analysis in E.coli,we synthesized gene fragment for the truncated N-terminal 80aa of VP1 protein,and then cloned it into vector and expressed the products in E.coli as insoluble proteins.The chaperone protein was further used to increase the soluble expression of VP1 protein in E.coli.The results showed that the chaperone protein significantly increased the expression level of VP1 protein,andoptimal induction conditions were 16℃,0.08 mmol/L of IPTG.The results of ELISA showed that the obtained soluble VP1 protein could be identified by standard positive serum of CIAV,which confirmed that the fusion protein had good aresponsiveness.Therefore,recombinant soluble VP1 protein has the potential to be developed as a new subunit vaccine and diagnostic antigen.
作者
黄英
付磊
靳泽华
易辰阳
王晓萍
张安定
HUANG Ying;FU Lei;JIN Ze-hua;YI Chen-yang;WANG Xiao-ping;ZHANG An-ding(State Key Laboratory of Agricultural Microbiology,Huazhong Agricultural University,Wuhan 430070,China;Key Laboratory of Preventive Veterinary Medicine in Hubei Province,Cooperative Innovation Center for Sustainable Pig Production,Wuhan 430070,China;Key Laboratory of Development of Veterinary Diagnostic Products,Ministry of Agriculture,International Research Center for Animal Disease,Ministry of Science and Technology,Wuhan 430070,China)
出处
《中国兽医杂志》
CAS
北大核心
2020年第8期16-19,24,共5页
Chinese Journal of Veterinary Medicine
基金
国家重点研发计划项目(2016YFD0500801)。
