ApoB RNA编辑高效蛋白检测体系的构建

Construction of an Efficient Protein Detection System to Monitor ApoB RNA Editing

[目的]建立一个载脂蛋白B(apoB)RNA编辑蛋白检测体系。[方法]在慢病毒表达质粒载体pCSII-CMV-IRES-Neor中插入apoB RNA编辑保守序列,并在其2端嵌入红色荧光蛋白(DsRed)、绿色荧光蛋白(GFP)表达序列,以此慢病载体转染大鼠肝癌细胞系CBRH-7919获得稳定表达。采用共聚焦显微镜测序方法检测apoB RNA的编辑。[结果]目的指示基因能够在CBRH-7919细胞中稳定表达,表现出指征RNA编辑的多色特征。[结论]试验成功构建了在细胞水平上以颜色变化指示apoB RNA编辑的检测体系,该体系为快速检测以apoB RNA编辑为靶的降胆固醇药物筛选提供了基础。

[ Objective] To develop a protein detection system monitor apoB RNA editing. [Method] We constructed a lentiviral vector pCSII- CMV-IRES-Neor, harboring the fluorescent markers discosoma sp. red fluorescent protein （DsRed） and green fluorescent protein （GFP） up- stream and downstream, respectively, of the conserved ApoB RNA editing sequence. A rat hepatoma （CBRH-7919） was then transfected, and its stable cell line expressing fluorescent markers was established. ApoB RNA editing was then detected under eonfocal microscopy and confirmed with sequencing. [ Result] The target gene that indicates apoB RNA editing was expressed stably in CBRH-7919 cells, exhibiting various fluorescent colors in cells. [ Conclusion] The system to monitor apoB RNA editing at the cellular level was constructed in this experi- ment, which provides a fast tool for screening the medicines that target the apoB RNA editing to reduce the cholesterol in plasma.