电穿孔法介导抗猪瘟病毒质粒DNA转染仔猪成纤维细胞研究  

Study on Transfecting the Anti-Swine Fever Virus Plasmid DNA into the Piglet Fibroblasts by Electroporation

作　　者：张文平[1] 周婧[1] 李方正[1] 宋学雄[1] 

机构地区：[1]青岛农业大学动物科技学院,山东青岛266109

出　　处：《青岛农业大学学报（自然科学版）》2013年第2期91-95,共5页Journal of Qingdao Agricultural University(Natural Science)

基　　金：国家转基因重大专项重点课题(2009ZX08006-007B)

摘　　要：本研究旨在用电穿孔法介导抗猪瘟病毒基因质粒转染仔猪成纤维细胞,优化电穿孔法转染条件。采用本实验培养的P6代仔猪成纤维细胞,用电穿孔法介导转染抗猪瘟病毒质粒,在电压、脉冲时间、脉冲次数、质粒DNA用量、电击后温度、电击缓冲液、质粒DNA构型等方面进行了转染条件优化的探索。结果显示,当电压为220V、脉冲时间25ms、脉冲1次,环形DNA用量为12μg,电击缓冲液为限定电转液时,细胞转染率高达47.49%。这些结果表明,用电穿孔法介导抗猪瘟病毒基因质粒转染仔猪成纤维细胞,可获得比较理想的细胞转染率。The aim of this research is to transfect the anti--swine fever virus plasmid into the piglet fibro- blasts by electroporation and optimize the conditions of electroporation. The anti--swine fever virus plas- mid was transfected into the P6 piglet fibroblasts cultured in our lab by electroporation, and transfection factors which includes voltage, pulse time, the number of pulse, the amount of plasmid DNA, the tempera- ture after electric shock, electroporation buffer, the types of plasmid were explored and optimized. The re- sults showed that the transfection efficiency is the highest, reaching 47.49 o/6o, when transfecting the fibro- blasts with 220V voltage, 25ms pulse time, 1pulse, 12μg annular plasmid DNA, the limited electropora- tion fluid as the buffer of electric shock and the transfected cells kept in 4℃ for 10min. These results indi- cated when transfecting the anti--swine fever virus plasmid into the piglet fibroblasts by electroporation, an ideal transfection efficiency could be obtained.

关 键 词：仔猪成纤维细胞 抗猪瘟病毒质粒 电穿孔 转染 

分 类 号：S828[农业科学—畜牧学] Q78[农业科学—畜牧兽医]