【摘要】 通过RT-PCR法获得阿克苏(Akesu/O/58)FMDV结构蛋白基因vp1,将该基因克隆到pGEM-T Easy载体进行核苷酸序列测定。将vp1基因、Kozak序列等插入中间表达质粒pBin438,构建重组中间表达载体pBinVP1。采用三亲融合法构建植物双元表达载体pBinFMDV-VP1。结果表明:vp1基因为639bp。重组中间表达载体pBinVP1经BamHⅠ/SalⅠ双酶切、PCR扩增和序列测定,表明vp1基因、Kozak序列等插入pBinVP1中。在含50mg/L卡那霉素、25mg/L链霉素和50mg/L利福平的YEB培养基上筛选及对融合农杆菌质粒进行vp1基因的PCR检测,证明植物双元表达载体pBinFMDV-VP1构建正确。更多还原

【Abstract】 The structural gene vp1 was obtained by reverse transcriptase-polymerase chain reaction(RT-PCR) from RNA of foot-and-mouth disease virus strain Akesu/58 Serotype O.The vp1 gene was cloned into the pGEM-T Easy vector and sequenced.The results showed that the vp1 gene contains 639 bp nucleotides.The recombinant mini-expression vector pBinVP1 was constructed by cloning the vp1 gene and Kozak sequence into plasmid pBin438 and confirmed by PCR,restriction enzyme analysis and nucleotide sequencing.Binary expression vector pBinFMDV-VP1 was constructed by triparental mating followed by selective screening of agrobacterium tumefaciens on YEB medium and PCR confirmation.更多还原