脂质体介导抗猪瘟病毒基因转染小鼠胚胎成纤维细胞的研究

摘要

The study aimed to investigate the conditions of transfection of the anti-classical swine fever virus gene recombination plasmid Pgpug/GFP/Neo into the mouse embryonic fibroblasts mediated by liposome, in order to provide the technical platform for construction of animal disease model and transgenic pigs against classical swine fever virus. In this study, different methods of isolation and culture of mouse embryonic fibroblasts were compared, and transfected the anti-classical swine fever virus gene recombination plasmid Pgpug/GFP/Neo into the 3 to 5 generation of fetal fibroblast cells mediated by liposome (LipofectamineTM2000). The results showed that, in different concentrations of liposomes, recombinant plasmid and different combinations of transfection time used in this sdudy, it was obtained 19% transfection rate in 2 Μl liposome and 1. 5 μg recombinant plasmid, incubated 6 h of combination, it was significantly higher than the other combinations(P<0. 05). The results indicated that, the anti-classical swine fever virus gene recombination plasmid Pgpug/GFP/Neo could be effectively transfected into mouse embryonic fibroblasts mediated by optimal combinations of liposomes, recombinant plasmid and transfection time.%本研究旨在探索脂质体介导抗猪瘟病毒基因重组质粒pGPUG/GFP/Neo转染小鼠胚胎成纤维细胞的条件,为构建疾病模型动物及抗猪瘟病毒转基因猪提供技术平台.本研究比较了不同的小鼠胚胎成纤维细胞分离培养方法,将培养至3～5代的胚胎儿成纤维细胞,通过脂质体(LipofectamineTM 2000)介导转染抗猪瘟病毒基因重组质粒pGPUG/GFP/Neo.结果显示,在所选用的不同浓度脂质体和重组质粒,以及不同转染时间的组合中,2μL脂质体介导1.5 μg重组质粒,转染6h时可获得19％的转染效率,极显著高于其他剂量和时间组合(P＜0.01).这些结果表明,脂质体、重组质粒及转染时间的优化组合,能有效介导抗猪瘟病毒基因重组质粒pGPUG/GFP/Neo转染小鼠胚胎成纤维细胞.