甲状旁腺素1受体(PTH1R)调控成釉细胞MMP-20基因表达的分子机制研究  

Study of Parathyroid Hormone 1 Receptor on Regulating the Expression of Matrix Metalloproteinase-20

作　　者：曲政[1] 张世龙[1] 袁杰[1] 高玉光[2] 

机构地区：[1]潍坊医学院口腔医学研究所,山东潍坊261053 [2]滨州医学院口腔医学院

出　　处：《潍坊医学院学报》2013年第4期282-284,I0001,共4页Acta Academiae Medicinae Weifang

基　　金：国家自然科学基金(30973327);山东省自然科学基金(ZR2010HM076)

摘　　要：目的通过进行甲状旁腺素l受体(PTH1R)基因克隆以及真核表达载体的构建,分析PTH1R对成釉细胞MMP-20基因表达的影响.为进一步探讨PTH1R在牙釉质形成以及发育过程中的分子调控方面奠定基础。方法根据引物设计原则设计PTH1R基因的RT-PCR引物,用PCR法扩增得出含有EcoRⅠ和XhoL的PTH1R目的基因片段。通过T4连接酶连接到pcDNA3.1/myc-HisA真核表达载体上。并用双荧光素酶报告基因系统检测MMP-20启动子区域的转录活性。结果①经过PCR引物扩增得到1794BP的基因片段,将获得的重组质粒pcDNA3.1/myc-HisA-PTH1R双酶切分析鉴定,测序结果与GenBank登录基因完全一致。②经双荧光素酶报告基因系统检测得出PTH1R能够上调MMP-20基因的表达水平。结论成功实现了PTH1R基因克隆及真核表达载体的构建,初步证明了PTH1R可能调控MMP-20基因的表达。Objective To study the regulatory effects of PTH1R for matrix metalloproteinase-20 in enamel developing of mouse through the parathyroid hormone 1 receptor（PTH1R） gene cloning and construct eukaryotic expres-sion vector of PTH1R, and to study the significance of PTH1R in developing dental enamel .Methods According to the RT-PCR primer design principles, the cDNA coding sequence of PTH1R containing the digestion site of EcoRⅠand XhoL was amplified by PCR from ameloblasts and then was inserted into pcDNA 3.1/myc-HisA vector.Dual luciferase reporter assay was used to analysis the effects of PTH 1R for transcription activity of MMP-20 promoter.Results ①The amplified gene fragment was 1794bp.Restriction enzyme digestion and sequencing showed that pcDNA 3.1/myc-HisA-PTH1R was constructed correctly.And it was confirmed by sequencing which was consistent with samples in GenBank .②Double luciferase genetic testing report system analysis showed that the expression of Dlx 1 was up-regulated.Conclu-sion The PTH1R gene cloning and eukaryotic PTH1R are constructed successfully , PTH1R can up-regulate the expres-sion of the MMP-20.

关 键 词：甲状旁腺素1受体 基因克隆 载体构建 RT-PCR 

分 类 号：R329[医药卫生—人体解剖和组织胚胎学]