Abstract: In order to develop a recombinant adenoviral vaccine, genes of a whole ORF coding genes of foot-and-mouth disease virus (FMDV) were amplified by RT-PCR and subsequently cloned into the vector pAdTrack-CMV to construct the recombinant adenoviral pAdTrack-CMV-ORF. PCR amplification and digestion with restriction endonucleases confirmed that the cloned ORF coding genes shared 99.8% similarity with that of original virulent strain type FMDV. Then the shuttle plasmid was linearized and transformed into competent cells of BJ5183 which containing the adenoviral backbone plasmid pAdeasy-2. The recombinant adenoviral plasmid (pAd-ORF) was identified by PCR amplification and digestion with PacⅠ. In this paper, we cloned the whole ORF coding genes of O type FMDV and successfully constructed the recombinant adenoviral shuttle vector.

摘要： 为研制O型口蹄疫病毒（FMDV）复制缺陷型腺病毒活载体疫苗毒株，通过RT-PCR方法获得了O型 FMDV的开放阅读框(ORF)基因，并定向克隆到腺病毒穿梭载体pAdTrack-CMV中，构建了重组腺病毒穿梭质粒pAdTrack-CMV-ORF，经PCR、酶切及测序鉴定，克隆得到的ORF编码基因序列与原始强毒株O型FMDV的ORF编码基因相似性达99.8%。将含有目的基因的穿梭质粒线性化后和腺病毒骨架载体pAdeasy-2共同电转化入大肠杆菌BJ5183感受态细胞中，得到重组腺病毒质粒pAd-ORF，经PCR和酶切鉴定正确。本研究成功获得了含有现代O型疫苗用FMDV毒株的全ORF编码基因的阳性克隆，并成功构建了含有完整编码基因表达盒的重组腺病毒穿梭质粒及其腺病毒骨架质粒。

关键词: 口蹄疫病毒, 开放阅读框, 腺病毒载体