TMEM64基因真核表达载体的建立与表达  

Construction and Expression of aTMEM64 Gene Eukaryotic Expression Vector

作　　者：武丽芳[1] 庄立琨[2] 邹清平[2] 贾培敏[2] 许桂平[3] 

机构地区：[1]重庆医科大学附属第二医院检验科,重庆市400010 [2]上海交通大学医学院附属瑞金医院上海血液学研究所,上海市200025 [3]重庆医科大学附属第二医院输血科,重庆市400010

出　　处：《医学分子生物学杂志》2013年第5期249-253,共5页Journal of Medical Molecular Biology

基　　金：国家自然科学基金（No.30670882）

摘　　要：目的构建带有GFP标签的TMEM64真核表达质粒，探求TMEM64在293T细胞中的表达特点。方法首先用RT—PCR的方法扩增白血病细胞株NB4中的TMEM64基因，并将其克隆到pEGFP—N2载体，然后用该质粒转染293T细胞，最后运用Western印迹检测TMEM64蛋白的表达情况，免疫荧光检测该蛋白亚细胞定位。结果在转染pEGFP—N2-TMEM64质粒的293T细胞中，能够检测到TMEM64一GFP重组蛋白的表达，而且该重组蛋白主要表达在细胞膜中。结论我们成功构建了带GFP标签的TMEM64真核表达质粒，为进一步研究TMEM64的功能奠定了分子基础。Objective To construct a TMEM64 gene eukaryotic expression vector with GFP tag and identify the expression features of TMEM64 in 293T cells. Methods TMEM64 gene in NB4 cells was amplified by RT-PCR and cloned into the pEGFP-N2 vector, and then transfected into 293T cells. The level of TMEM64-GFP recombinant protein was detected by Western blotting, and its intracellular localization was analyzed by immunofluorescence. Results The TMEM64-GFP re- combinant protein was expressed in 293T ceils transfected with pEGFP-N2-TMEM64 plasmid. Furthermore, this recombinant protein was mainly expressed on the cell membrane. Conclusions The TMEM64 gene eukaryotic expression vector with GFP tag was successfully constructed, providing a powerful molecular basis for subsequent investigation on the functions of TMEM64.

关 键 词：TMEM64基因 真核表达载体 急性早幼粒细胞白血病 

分 类 号：Q78[生物学—分子生物学]