口蹄疫病毒3D基因在重组杆状病毒中的表达及检测  

Expression and detection of 3D gene of FMDV in recombinant baculovirus

作　　者：鲁会军[1] 金宁一[1] 韩松[1] 郑敏[1] 尹革芬[1] 张洪勇[1] 葛淑敏[1] 金扩世[1] 

机构地区：[1]解放军军需大学军事兽医研究所解放军基因工程重点实验室,吉林长春130062

出　　处：《中国预防兽医学报》2005年第3期175-178,共4页Chinese Journal of Preventive Veterinary Medicine

摘　　要：利用杆状病毒表达系统进行了口蹄疫病毒3D基因在Sf9细胞中的表达研究。首先克隆了3D基因片段,将pMD18_3D质粒及杆状病毒转座载体质粒pFastBacⅠ分别用EcoRⅠ及XbaⅠ酶切后,用T4DNA连接酶连接,构建了重组质粒pFastBac_3D;再将该重组质粒转化DH10Bac感受态细菌,在体内进行重组,并经三重抗性与蓝白斑筛选,得到杆状病毒重组质粒Bacmid_3D;将Bacmid_3D转染Sf9细胞,获得重组杆状病毒,并进行表达水平的检测。经SDS_PAGE和West ernblot检测,结果表明,3D蛋白在重组杆状病毒中获得表达。According to the 3D gene of FMDV,the primers of polymerase gene were designed,and the objective fragment was (1 410 bp) which was cloned into pMD18_T vector.Then the 3D gene was cloned into baculovirus expression vectors pFastBacⅠ,and the recombinant plasmid of pFastBac_3D was constructed.The plasmid were transformed to DH10 Bac competent cell,and was screened by three antibiotics and blue_white patch, the white colony was the recombinant Bacmid_3D.And then the bacmid_3D was transfected into the Sf9 insect cells,by three passaging the recombinant baculovirus was obtained.Finally,the expressed productions were detected by SDS_PAGE and Western_blotting,and the results showed that the polymerase(3D protein) was high_efficiently expressed in recombinant baculovirus.

关 键 词：口蹄疫病毒 3D基因 杆状病毒 表达 

分 类 号：S852.65[农业科学—基础兽医学] Q784[农业科学—兽医学]