摘要
目的:构建人牙成釉细胞相关蛋白(odontogenic ameloblast-asssociated protein,ODAM)基因不同长度的上游启动子荧光素酶报告基因载体,比较不同的启动子片段在成釉细胞、Hela细胞中的活性,为进一步判定上游启动子的转录调控区奠定基础。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ODAM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不同的细胞中活性不同,-333～-195与-195～-96区域为特异的转录调控作用区。结论:初步确定了ODAM基因启动子的转录活性区,为进一步研究该基因转录调控特点奠定基础。
AIM：To construct luciferase report gene vectors with different promoter segments of human odontogenic ameloblast-asssociated protein （ODAM）,and to compare the luciferase activities of different length segments of human ODAM in ameloblast and Hela cells so as to provide information for determining transcription regulation district of promoter sequence in further study.METHODS：The different-length desired promoter segments were obtained by PCR method,and cloned into luciferase report gene vectors pGL3-Basic.The construction was transiently transfected into ameloblasts and Hela cells.Sussequently,the transcriptional regulatory capacity of the promoter segments was analysed by luciferase activity.RESULTS：The different-length promoter segments of ODAM gene was obtained.Different-length promoter recombinant luciferase reporter vectors were constructed successfully by cutting with two different restriction enzymes.Different-length promoters showed different activities in different cells.The specific transcriptional regulatory district were-333～-195 and-195～-96 regions.CONCLUSION：The transcriptional activity regions of ODAM gene promoter were determined,which will provide useful information for further studying the transcriptional regulatory characteristics of ODAM gene.
出处
《牙体牙髓牙周病学杂志》
CAS
北大核心
2010年第4期187-191,共5页
Chinese Journal of Conservative Dentistry
基金
国家自然科学基金(36072316)
山东省教育厅基金资助项目(J08LH65)
山东省自然科学基金资助项目(Y2006C106)
