口蹄疫病毒P1+2A基因真核表达载体的构建

Construction of eukaryon expression vectors for P1+2A genes of foot-and-mouth disease virus

通过RT-PCR方法获得了口蹄疫病毒O型,编号为OF3毒的P1+2A区的目的基因,并将其克隆到克隆载体pMD18-T上,再根据表达载体pQE-Trisystem的特性及酶切位点设计合适的表达引物.由表达引物通过PCR技术从克隆载体上扩增出带有特定酶切位点的目的片段,再对载体和目的片段进行酶切处理,处理后由T4 DNA连接酶将二者连接.通过PCR及酶切鉴定,结果证明口蹄疫病毒目的基因片段已被正确克隆到表达载体pQE-Trisystem上.

The antigen DNA fragments of FMDV P1＋2A genes were obtained through RT-PCR method. Then the fragments were cloned into the cloning vectors,pMD18-T vectors. Due to the characteristic of the eu-expression vector,pQE-Trisystem,and its enzyme sites, the appropriate primers were designed. Using the expression primers cloned the aimed DNA fragements which contain the right enzyme sites were amplified. The expression vectors and the aimed fragements were digested respectively,and then,they were ligated by the T4 DNA ligatease. Through PCR,restriction analysis and DNA sequencing,the positive cloning plasmid pQE＋P12A was found.