摘要
为探讨载脂蛋白CⅡ基因启动子区 - 190bp处的碱基突变 (T→A)对载脂蛋白CⅡ基因转录的影响 ,及其与血浆载脂蛋白CⅡ浓度降低的关系。通过定点诱变技术构建带有 - 190bp处T→A突变的载脂蛋白CⅡ启动子的荧光素酶表达载体pGL3;应用脂质体介导的基因转染技术 ,将带有正常的载脂蛋白CⅡ启动子的表达质粒和构建的突变质粒分别与内参照质粒pRL TK共转染HepG2细胞 ,瞬时表达 ;利用双荧光素酶测试系统检测荧光素酶活性。结果发现 ,带有启动子区 - 190T→A突变的载脂蛋白CⅡ启动子的荧光素酶表达活性比正常的载脂蛋白CⅡ启动子的表达活性低 18.5 6 %。结果提示 ,载脂蛋白CⅡ基因启动子区 - 190bp处的碱基由T突变为A降低了载脂蛋白CⅡ启动子的转录活性。
Aim To study the effect of apolipoprotein CⅡ promoter T→A mutation at position -190 on the transcription of the apolipoprotein CⅡ gene and its relationship to the decrease of apolipoprotein CⅡ concentration in plasma. Methods Firstly, the expression vector pGL3 with -190 T→A mutant promoter was constructed using site-directed mutagenesis; Secondly, the expression vector pGL3 with mutant promoter and normal promoter respectively was co-transfected with pRL-TK as an internal control into HepG2 cells by liposome-mediated methods, and expressed transiently. Thirdly, Both firefly luciferase activity drived by normal/mutant apolipoprotein CⅡ promoter from pGL3 and renilla luciferase activity drived by TK promoter from pRL-TK were measured by dual luciferase reporter assay system. Results The results showed that the T→A mutation at position -190 in apolipoprotein CⅡ promoter caused a decrease of 18.96% in luciferase activity compared with the normal apolipoprotein CⅡ promoter. Conclusions The T→A mutation of -190 base in apolipoprotein CⅡ promoter region could debase transcription activity of apolipoprotein CⅡ promoter.
出处
《中国动脉硬化杂志》
CAS
CSCD
2003年第6期509-511,共3页
Chinese Journal of Arteriosclerosis
基金
国家自然科学基金 ( 3 0 0 70 3 5 4)资助
