犬2型腺病毒E1转化细胞的建立

The Establishment of CAV-2 E1 Transformed Cell

为建立能表达犬 2型腺病毒 (CAV 2 )E1基因的辅助细胞 ,本研究用含CAV 2E1基因的重组质粒pc E1t对DK细胞进行转染 ,经G41 8的筛选 ,得到 2 6个细胞克隆。用CAV 2E1A特异引物对各细胞克隆进行PCR和RT PCR检测 ,PCR结果表明转染后的细胞克隆都能扩增出 5 3 6bp的特异性片段 ,但仅有 6株为RT PCR强阳性 ,能扩增出 6 5 2bp的特异带。再对RT PCR强阳性的细胞克隆进行Westernblot分析 ,最终挑选到一个既能转录E1A基因 ,又能表达E1B 1 9kD蛋白的克隆细胞株 (DK E1 )。

To establish the accessory cell which could express the CAV-2 E1gene,an expressing vector containing CAV-2 E1 fragment was used to transform the DK cells.During the screening of G418,several cell clones were obtained.A pair of CAV-2 E1A gene primers were used to test the cell clones by PCR and RT-PCR.Western blot was also used to test the cell clones.At last,only one cell clone which could transcript the E1A gene and express the E1B 19kD protein was screed.;