犬传染性肝炎病毒Hexon蛋白基因的克隆和原核表达质粒的构建

Cloning of Hexon Gene of Canine Infectious Hepatitis Virus and Construction of Recombined Prokaryotic Expression Plasmid

通过设计特异性引物,以犬传染性肝炎患犬的全血基因组DNA为模板,经PCR扩增,获得了犬传染性肝炎病毒Hexon蛋白基因,将其插入到pMD18-T载体进行测序、鉴定,构建原核表达质粒载体pGEX-6p-1-Hex-on。结果显示,犬传染性肝炎病毒Hexon基因已被成功地克隆,基因全长2 718 bp,包含从起始密码子ATG到终止密码子TAA的完整序列。与犬传染性肝炎病毒国际标准株Hexon蛋白基因的同源性为99.7%,通过实验鉴定表明pMD18-Hexon克隆载体和原核表达质粒载体pGEX-6p-1-Hexon已成功构建。

One pair of the special primers was designed by Oligo6.0 software according to the Hexon gene sequences of canine Infectious Hepatitis virus（CIHV） published in GenBank. The Hexon gene fragment in CIHV was amplified by PCR from the genomic DNA in the whole blood samples infected with CIHV and was cloned into a pMD18-T vector before sequencing and identification. A prokaryotic expression vector pGEX-6p-1-Hexon was then constructed. The result was shown that hexon protein gene sequence of the Infectious Canine Hepatitis Virus was successfully cloned. Sequence analysis was shown the total length of CIHV Hexon gene was 2 718 bp with integrated Sequence from start eodon ATG to stop codon TAA. It shared 99.7% nucleotide homology with that published in GenBank. The experimentat identification was shown that the pMD18-Hexon vector and the prokaryotic expression vector pGEX 6p-1-Hexon were successfully constructed.