人成釉蛋白基因在COS-7细胞中的表达  

Expression of human ameloblastin in mammalian COS-7 cells

作　　者：田宇[1] 鲁红[1] 倪龙兴[1] 肖明振[1] 余擎[1] 

机构地区：[1]第四军医大学口腔医学院,陕西西安710032

出　　处：《牙体牙髓牙周病学杂志》2006年第1期1-4,共4页Chinese Journal of Conservative Dentistry

基　　金：国家自然科学基金资助(30300386);全军"十五"计划基金重点课题(012089);第四军医大学学术新人基金资助

摘　　要：目的:通过构建真核表达载体,在COS-7细胞中表达人成釉蛋白基因。方法:分别将人成釉蛋白基因克隆入非融合和融合真核表达载体pcDNA 3.1和pEGFP-C3中,证实克隆正确后,用脂质体介导将载体转染哺乳动物细胞COS-7,通过免疫组织化学、W estern B lot等方法检测成釉蛋白在哺乳动物细胞中的表达。结果:表达的绿色荧光蛋白融合蛋白所发出的荧光位于细胞浆中,胞核中无荧光,与免疫组化结果一致,说明人成釉蛋白位于细胞浆中。W estern B lot检测显示在COS-7细胞中表达的成釉蛋白的分子量为65×103,分泌至培养基中的成釉蛋白的分子量为62×103。结论:成功在哺乳动物细胞中表达了人成釉蛋白,为研究成釉蛋白合成后的修饰加工和成釉蛋白的结构,为今后进一步研究成釉蛋白结构和功能的关系奠定了基础。AIM：To construct eukaryotic expression plasmids and express human ameloblastin in COS -7 cells. METHODS：We constructed eukaryotic expression plasmids by inserting ameloblastin cDNA into pcDNA 3.1 and pEGFP - C3 and transfected the recombinations into COS - 7 cells. The expressed ameloblastin was examined by fluorescence microscopy,immunohistochemistry and Western Blot. RESULTS：The green fluorescence emitted by the expressed fusion protein under fluorescence microscope located in the cell plasm, not in the nuclear. Immunohistochemistry showed the same results. Western blot revealed that the ameloblastin was synthesized as a 65 kDa protein and became 62 kDa when it was secreted, CONCLUSION： We successfully expressed human ameloblastin in mammalian, which would facilitate our understanding of the post - translation events and the structure of ameloblastin.

关 键 词：成釉蛋白 免疫组化 绿色荧光蛋白 蛋白基因 COS-7细胞 

分 类 号：R780.2[医药卫生—口腔医学]