【摘要】 目的:分析人β1整合素在HaCat细胞中核心启动片段。方法:以本室构建的含整合素β1全长启动子1745bp基因序列(-1745bp～+11bp)的重组载体pGL3-1756为模板,用含有酶切位点的特异性引物扩增出整合素β1启动子-845bp～-304bp间基因序列,克隆至荧光素酶表达载体pGL3basic,构建含正确目的基因重组载体pGL3-542。用pGL3-1756、和pGL3-542质粒转染人表皮细胞株HaCat进行活性分析。结果:酶切及序列测定表明,克隆后插入pGL3basic中的启动子片段与GenBank DNA序列数据库对比分析序列一致,且插入方向正确。在人永生化表皮细胞株(HaCat细胞)中构建的pGL3-542载体具有很强的启动活性。结论:成功构建了整合素β1远端启动子542bp片段载体,在HaCat细胞中具有非常强的启动活性。人整合素β1整合素启动子核心启动序列可能位于-845bp～-304bp。更多还原

【Abstract】 Objective:To analyze the core fragment of integrin beta1 in HaCat cells.Methods:pGL3-1756,a constructed vector containing the human integrin beta 1 promoter(1756 bp),was used as template for amplification of the vector,which was named pGL3-542.HaCat cells were transfected with pGL3-1756,promoter sequence at the distal part(-845 bp～-304 bp).Then the amplified se-quence was cloned into pGL3 basic pGL3-542,respectively,for the analysis of promoter activity,and pGL3-basic was used as negative controls.Results:The results of enzyme digestion and sequencing identified the correction of constructed vector.pGL3-542 exhibited very strong activity in HaCat cells.Conclusion:The reporter gene vector containing 542 base pairs at the distal part of integrin beta 1 pro-moter is constructed successfully,with strong activity in HaCat cells.The core sequences at human integrin β1 promoter may be located in-845 bp～-304 bp.更多还原