人酰基辅酶A:胆固醇酰基转移酶1基因P7启动子调控的报告基因载体的构建及鉴定  

Construction and Identification of Recombinant Firefly Luciferase Report Vector Containing Human Acyl Coenzyme A: Cholesterol Acyltransferase 1 Gene P7 Promoter

作　　者：葛晶[1] 成蓓[1] 何平[1] 文晖[1] 陆菡[1] 陈心[1] 曾永利[1] 

机构地区：[1]华中科技大学同济医学院附属协和医院老年病科,武汉430022

出　　处：《生物医学工程学杂志》2008年第6期1381-1384,共4页Journal of Biomedical Engineering

基　　金：国家自然科学基金资助项目(30471921)

摘　　要：应用PCR方法从人单核细胞系THP-1基因组DNA扩增人酰基辅酶A:胆固醇酰基转移酶1(acyl coen-zyme A:cholesterol acyltransferase1,ACAT1)基因P7启动子全长片段,进行TA克隆,经双酶切、测序鉴定阳性克隆。将该基因亚克隆至荧光素酶报告基因载体pGL3-Enhancer,用DEAE-葡聚糖(DEAE-dextran)法将重组质粒pGL3E-P7瞬时转染入单核细胞THP-1,检测其活性,证明pGL3E-P7能在体外表达荧光素酶。人ACAT1基因P7启动子调控的荧光素酶报告基因表达载体的成功构建,为研究动脉粥样硬化过程中ACAT1基因的转录调控机制提供新手段。The DNA segment of the human acyl coenzyme A： cholesterol acyltransferasel （ACAT1） gene P7 promoter was amplified by PCR from human monocytic leukemia cell line （THP-1） and cloned to TA vector, then the positive clone was confirmed by restriction enzymes and sequencing. The targeted segment was subcloned to Firefly luclferase report vector pOL3-Enhancer. The recombinant plasmid pGL3E-P7 was transfected transiently into THP-1, then the expression of luciferase could be detected in THP-1 by pGL3E-P7 transfection. We successfully constructed luciferase reporter vector containing P7 promoter of the human ACAT1 gene, and established a new means to study the transcriptional regulation mechanisms of ACAT1 during atherosclerosis.

关 键 词：动脉粥样硬化 酰基辅酶A:胆固醇酰基转移酶1 启动子 荧光素酶 

分 类 号：Q78[生物学—分子生物学]