贵州地区羊口疮病毒B2L基因克隆及原核表达载体构建

用H2L基因特异性引物对羊口疮临床疑似病料进行鉴定,采用PCR扩增出阳性样品羊口疮病毒结构蛋白B2L基因,回收纯化PCR产物,克隆至pMD18-T,测序结果表明插入的片段为目的基因,全长1137 bp。将目的基因定向克隆至pET-32a构建了B2L基因原核表达载体pET-32a-B2L,经PCR鉴定、酶切及进一步测序证明表达载体构建成功,为下一步的表达及基因工程疫苗的研究奠定了良好基础。

Cloning and Construction of Prokaryotic Expression Vector of B2L Gene of Orf Virus in Guizhou

To identify suspected goats orf case and construction of prokaryotic expression vector of B2L gene of orf virus (ORFV),clinical and suspect samples of orf could be amplified by a pair of specific primers,then the structural protein B2L gene of orf virus was amplified by PCR method using specific primers.The B2L gene was ligated with pMD18-T vector and the sequencing results showed that the full-length of the inserted gene fragment was 1137 kb. Cutting the target fragment,which was cloned into pET-32a vector and construction of prokaryotic expression vector pET-32a-B2L,the recombinant plasimid was identified by restriction enzyme and PCR,sequence analysis indicated that to be successful construction of pET-32a-B2L expression vector and it has laid a good foundation for the expression and genetic engineering vaccine research.