大鼠GLUT1真核表达载体的构建及其在293细胞中的表达  

Construction of eukaryotic expression vector carrying rat GLUT1 gene and its expression in 293 cells

作　　者：李方成[1] 李军亮[1] 刘然义[2] 吴中华[1] 张培峰[1] 刘安民[1] 

机构地区：[1]中山大学附属第二医院神经外科 [2]中山大学肿瘤防治中心实验室,广东广州510120

出　　处：《中国病理生理杂志》2005年第10期1994-1998,共5页Chinese Journal of Pathophysiology

基　　金：国家自然科学基金资助项目(No.39900048);广东省自然科学基金资助项目(No.010721)

摘　　要：目的:构建大鼠葡萄糖转运体1(GLUT1)的真核表达载体。方法:以RT-PCR方法从大鼠脑组织中 获取GLUT1全长cDNA片段,将其克隆至真核表达质粒pcDNA3．1(+)中,构建重组真核表达质粒pcDNA3．1(+)- Glutl,随后用lipofectamineTM2000介导转染HEK293细胞,以RT-PCR法检测重组质粒在mRNA水平的表达,以免疫 组化的方法检测重组质粒在蛋白水平的表达。结果:以构建的重组真核表达质粒pcDNA3．1(+)-Glutl转染293细 胞后,在基因及蛋白表达水平均检测到了葡萄糖转运体1的表达。结论:成功构建了携带大鼠葡萄糖转运体1的真 核表达载体pcDNA3．1(+)-Glutl,且证实其可在293细胞中成功表达目的基因,为进一步研究外源性GLUT1表达对 缺血缺氧脑细胞的保护作用奠定了基础。AIM： To construct a eukaryotic expression vector carrying rat GLUTI gene. METHODS： GLUTI cDNA was obtained by RT - PCR technique from rat cerebral tissue and was inserted into eukaryotic expression vector pcDNA3.1 （ ＋ ） to construct the recombinant expression plasmid pcDNA3.1 （ ＋ ） - GLUTI. The recombinant plasmid was transfected into 293 cells by lipofectamineTM 2000. The transcription and the expression of GLUTI in 293 ceils were tested by RT- PCR and immunocytochemistry, respectively. RESULTS： The transcription of GLUTI gene and expression of GLUTI protein in transfected 293 cells were observed by RT- PCR and immunocytochemistry. CONCLUSION： The eukaryotic expression vector pcDNA3.1 （ ＋ ） - GLUTI is constructed successfully and it can express GLUTI protein in 293 cells.

关 键 词：葡萄糖转运体1 真核表达 脑缺血 293细胞 免疫组织化学 

分 类 号：R363[医药卫生—病理学]