摘要
目的V构建全长及截短型Cx36基因重组表达载体,观察其在Hela细胞的表达。方法RTPCR获得全长及截短型Cx36编码区基因片段,与pCR2.1TOPO克隆载体连接酶切纯化鉴定,亚克隆到pcDNA3构建真核表达载体,脂质体介导法转染Hela细胞,G418抗性筛选阳性克隆,Western印迹和免疫荧光方法分析Hela细胞Cx36的表达。结果构建了全长及截短型重组真核表达载体Cx36-pcDNA3,转染的Hela细胞获得稳定表达Cx36的基因,Western印迹和免疫荧光显示转染的HeLa细胞表达Cx36蛋白,但转染截短型Cx36的Hela细胞表达量明显少于转染全长Cx36的细胞。结论全长及截短型Cx36基因可在Hela细胞中稳定表达,截短型Cx36基因表达较少。
Objective To construct full length and truncated Cx36 recombinant expression vector,and observe their expression in Hela cells.Methods Full length and truncated Cx36 were obtained by RT-PCR,PCR products were cloned into PCR2.1TOPO vector,Cx36 -pcDNA3 expression vector construction,cell culture,transient transfection using lipofectamine 2000,stable clone screening with G418. Western blotting and immunofluorescence were used to analyse the expression of Cx36 in Hela cells.Results Full length and truncated Cx3...
出处
《潍坊医学院学报》
2008年第6期-,共4页
Acta Academiae Medicinae Weifang
