人釉蛋白基因启动子在成釉细胞中的转录活性研究  

Study of Transcription Activity about Human Enamelin Promoter in Ameloblast

作　　者：刘晓影[1] 高志芹[1] 韩婷婷[2] 官秀梅[3] 高玉光[2] 

机构地区：[1]潍坊医学院细胞生物学教研室,山东潍坊261053 [2]潍坊医学院口腔研究所 [3]潍坊医学院生化教研室

出　　处：《潍坊医学院学报》2009年第6期405-408,F0003,共5页Acta Academiae Medicinae Weifang

基　　金：国家自然科学基金资助项目(课题编号:30672316);山东省自然科学基金资助项目(课题编号:Y2006C106)

摘　　要：目的构建人釉蛋白(Enamelin,ENAM)基因不同长度的上游启动子荧光素酶报告基因载体,通过分析不同的启动子片段的活性,确定启动子的功能区域,为进一步研究Enamelin基因的转录调控机制奠定基础。方法以PCR方法获取基因上游启动子片段,将其构建至报告基因载体pGL3-Basic中,瞬时转染成釉细胞及Hela细胞,通过检测荧光素酶活性来分析启动子区域的转录调控能力。结果成功地获得了不同长度的Enamelin基因启动子目的片段,酶切鉴定表明不同长度启动子荧光素酶报告基因载体构建成功,不同长度的启动子在成釉细胞中活性不同,-1216～-554与-312～-133区域为特异的转录调控作用区。结论成功构建了不同长度的Enamelin基因启动子的pGL3-Basic荧光素酶报告基因载体,初步确定Enamelin基因启动子的转录活性区,为进一步研究Enamelin基因转录调控特点奠定基础。Objective Constructing luciferase report gene vectors with different promoter segments of human Enamelin,and comparing the luciferase activity of different length segments of human Enamelin in Ameloblast and Hela cells, so as to provide a foundation for confirming transcription regulation district of promoter sequence in further study. Methods The different-length desired promoter segments were obtained by PCR method, and they were cloned into luciferase report gene vectors pGL3-Basic, then transiently transfected into ALC cells and Hela ceils. Transcriptional regulatory capacity of the promoter segments were analyzed through detecting luciferase activity. Results Different-length promoter segments of Enamelin gene have been obtained,and the different-length promoter recombinant luciferase reporter vectors were constructed successfully by cutting them with two different restrict enzymes. Different-length promoters have different activity in different-type ceils, and - 1216 - - 554 and - 312 ~ - 133 regions are specific transcriptional regulatory district. Conclusion Different-length recombinant pGL3-Basic lueiferase reporter gene vector were successfully constructed. The transcriptional activity regions of Enamelin gene promoter were initially determined, which will provide a foundation for further studying the transcriptional regulatory characteristics of Enamelin gene.

关 键 词：ENAMELIN pGL3-BaSic荧光素酶报告基因载体 成釉细胞 

分 类 号：R392.11[医药卫生—免疫学]