EphA3基因启动子区域定位表达及活性分析  

Promoter Regions and Transcriptional Activities of EphA3 in 293T and MEF Cell Lines

作　　者：曾莉[1] 卜碧涛[1] 

机构地区：[1]华中科技大学同济医学院附属同济医院神经科,武汉市430030

出　　处：《医学分子生物学杂志》2012年第2期88-92,共5页Journal of Medical Molecular Biology

基　　金：国家自然科学基金（No.30670735）

摘　　要：目的构建含有不同长度EphA3基因启动子片段的报告基因载体，研究其在293T细胞和MEF细胞中的转录活性。方法以Balb／C小鼠基因组DNA为模板，扩增不同长度的EphA3基因启动子片段，并克隆进入荧光素酶报告基因质粒pGL3-Basic真核表达载体内。酶切鉴定及基因测序无误后，将重组质粒和pRL—CMV内对照质粒共转染293T和MEF细胞，分析不同长度的OhA3基因启动子片段的转录活性。结果酶切和测序鉴定表明表达载体构建成功，EphA3基因的核心启动子区域位于-279bp～＋110bp之间，在293T细胞和MEF细胞中其转录活性相似。结论成功构建了荧光素报告基因重组质粒，并确定了BphA3基因的核心启动子区域。Objective To construct reporter vectors containing EphA3 promoter region in dif- ferent lengths and test their transcriptional activities in 293T and MEF cells. Methods Genomic DNA from Balb/C mice was used as a template to synthesize various PCR products in different lengths, and the PCR products were then cloned into pGL3-Basic vector to develop a series of re- porter constructs. These constructs were co-transfected with pRL-CMV vector into 293T or MEF cells, and luciferase reporter activities were measured. Results All constructs were verified by re- striction enzyme digestion and sequencing. The basic promoter region of EphA3 was from-279 bp to transcription start site （ ＋ 110 bp） . Promoter activities of EphA3 gene in 293T and MEF cells were measured similarly. Conclusion EphA3 promoter regions in different lengths are successfully cloned into reporter vector and the basic promoter region has been confirmed.

关 键 词：EphA3基因 启动子 荧光素酶报告基因载体 转录活性 

分 类 号：Q785[生物学—分子生物学]