猪圆环病毒2型ORF2基因片段的原核表达及间接ELISA诊断方法的初步建立  

Expression of fragment of porcine circovirus type 2 ORF2 in Escherichia coli and development of indirect ELISA with recombinant protein

作　　者：屈素洁[1] 何苹萍[1] 秦毅斌[1] 朱芸[1] 陈琼[1] 黄伟坚[1] 

机构地区：[1]广西大学动物科学技术学院,广西南宁530005

出　　处：《广西农业生物科学》2008年第3期201-205,共5页Journal of Guangxi Agricultural and Biological Science

基　　金：广西科学研究与技术开发项目(桂科攻0537008-3);广西科技攻关项目(0537022-1;0630006-6B)

摘　　要：根据猪圆环病毒2型(PCV2)LC株序列,设计合成3对引物,通过PCR扩增PCV2的3个不同ORF2基因片段,分别将其克隆到pET-32a载体中,将重组表达质粒转化大肠杆菌BL21,表达并纯化了重组蛋白。Western-blot分析表明,重组蛋白可以与PCV2阳性血清反应,表明该蛋白具有良好的抗原性。以纯化的重组蛋白初步建立了间接酶联免疫吸附实验(ELISA)方法,经方阵滴定确定最佳包被浓度为0.24μg/mL,血清最佳的稀释度为1∶40。According to the porcine circovirus type 2 （PCV2） LC strain, 3 pair of primers were designed and synthesized to amplify the ORF2b, ORF2c and ORF2d gene. The amplified products were cloned into expression vector pET-32a and expression in Escherichia coli BL21. The Westernblot analysis showed that the recombinant protein had a good antigenicity with positive PCV2 serum. An indirected enzymelinked immunosorbentassy （ELISA） was established with the purified recombinant protein, after the square matrix titrate determination best density was 0. 24 μg/mL, the blood serum best dilution ratio was 1 ： 40.

关 键 词：猪圆环病毒2型(PCV2) ORF2片段 原核表达 间接ELISA方法 

分 类 号：S854.43[农业科学—临床兽医学]