人BRD8 启动子系列报告基因的构建和鉴定.pdfVIP

684 J Bengbu Med Coll,July2011,Vol.36,No.7 [文章编号] 1000鄄2200(2011)07鄄0684鄄03 ·基础医学· 人 BRD8启动子系列报告基因的构建和鉴定 王传付,马摇 华 [摘要]目的:构建具有相同3忆末端而5忆末端逐渐截短的人 BRD8启动子系列报告基因,并进行鉴定。 方法:聚合酶链反应 (PCR)从人基因组中扩增BRD8 启动子片段并插入荧光素酶报告基因载体pGL3鄄Basic;以已构建的BRD8报告基因为模板, 分别用PCR扩增具有相同3忆末端而5忆末端逐渐截短的BRD8启动子系列片段,并分别插入pGL3鄄Basic载体。 对BRD8启动 子系列报告基因进行酶切鉴定、测序。 结果:通过酶切鉴定、测序,证明构建的BRD8启动子系列报告基因序列和方向正确。 结论:成功构建了人BRD8启动子系列报告基因,为进一步研究调节BRD8基因表达的特异性转录因子和(或)沉默子奠定了 基础。 [关键词]基因,BRD8;启动子;荧光素酶;报告基因 [中国图书资料分类法分类号] Q343.1摇 摇 摇 [文献标识码]A Construction and identification of serial human BRD8 reporter genes WANG Chuan鄄fu,MA Hua (Department of Immunology,BengbuMedical CollegeKey Laboratory of InfectionandImmunity inAnhui,BengbuAnhui233030,China) [Abstract] Objective:To construct and identify serial human BRD8 luciferase reporter genes with the identical 3忆鄄end but gradually truncated5忆鄄end.Methods:The BRD8 promotor fragment from human genomewas amplified by polymerase chain reaction(PCR) and inserted into the luciferase reporter gene vector pGL3鄄Basic. A series of BRD8 promotor fragments with the identical 3忆鄄end but gradually truncated5忆鄄endwereproducedby PCRusingthe above constructed BRD8reporter genevector astemplate and inserted into pGL3鄄Basic vector,respectively.The series of BRD8 reporter genes were identified by restriction enzyme digestion and sequencing. Results:The restriction enzyme digestion and DNA sequencing results showed that the sequences and orientation of the serial BRD8 reporter geneswere correct.Conclusions:The serial human BRD8 reporter genes had been successfully constructed.These provide an experimental base for further research on the regulation of BRD8 gene transcription by some specific transcription factors and/ or silencers. [Key words] gene,BRD8