ULBP1启动子报告质粒的构建及NS3/4A蛋白对ULBP1基因转录的作用

The construction of reporter plasmid of ULBP1 and preliminary studying on the influence of NS3/4A on transcription of ULBP1

目的:构建含有ULBP1基因启动子的报告质粒,并初步研究NS3/4A对ULBP1转录的影响。方法:将ULBP1启动子核心区序列连接在pGL3-enhance载体上构建ULBP1报告质粒pGL3-ULBP1质粒;将HCV的蛋白酶NS3/4A基因亚克隆到pcDNA3-Flag载体上(pcDNA3-Flag-NS3/4A),Western blot检测其表达。用荧光分度计检测NS3/4A对ULBP1转录水平的影响。结果:成功构建了含有ULBP1启动子的报告质粒pGL3-ULBP1和FLAG-NS3/4A真核表达质粒,并检测到NS3/4A可以抑制ULBP1的转录。结论:HCV的蛋白酶NS3/4A可以抑制ULBP1的转录。

AIM：To construct a report vector of ULBP1 promoter gene and preliminary study on the influence of NS3/4A on transcription of ULBP1. METHODS：ULBP1 core promoter sequence was amplified by PCR,and inserted into pGL3-enhance vector,constructing ULBP1 reporter plasmid pGL3-ULBP1; The HCV protease NS3/4A gene was subcloned into pcDNA3-Flag vector （pcDNA3-Flag-NS3/4A）,and the expression of this plasmid was demonstrated by Western blot. The influence of inhibition by NS3/4A on the level of ULBP1 transcription was tested by assaying the Luciferase activity in cells transfected with pGL3-ULBP1 with Luminometer. RESULTS：The reporter plasmid of ULBP1 promoter gene and the eukaryotic expression plasmid Flag-NS3/4A have been constructed and expressed successfully; and the protease NS3/4A inhibit the level of ULPB transcription. CONCLUSION：The protease NS3/4A of HCV down-regulates ULBP1 expression by inhibiting the transcription of ULBP1.