Construction and ldentification of Cell Division Cycle 2 Promoter Reporter Gene Vector
10.3969/j.issn.0253-9896.2014.02.002
- VernacularTitle:细胞分裂周期蛋白2启动子报告基因载体的构建与鉴定
- Author:
Hui ZHOU
;
Wei WU
;
Ming ZHAO
- Publication Type:Journal Article
- Keywords:
cyclins;
genes,cdc;
cyclin-dependent kinases;
transcription initiation site;
cell cycle;
luciferases
- From:
Tianjin Medical Journal
2014;(2):101-104
- CountryChina
- Language:Chinese
-
Abstract:
Objective To construct the luciferase reporter gene vector of cell division cycle 2 (Cdc2) gene promoter and determine its transcriptional activity. Methods Primers were designed based on human Cdc2 promoter sequence from UCSC software. Then Cdc2 promoter from human genome DNA was replicated. After pGL3-Basic vector and Cdc2 promoter were digested with restriction enzymes SacⅠand XhoⅠseparately, Cdc2 promoter was inserted into pGL3-Basic vector. The recombinant plasmid named pGL3-Cdc2-promoter was transiently co-transfected into U2OS cells with control vector pRL-SV40, and then the activity of dual luciferase was detected. Results pGL3-Cdc2-promoter was constructed successfully. The restriction analysis and sequencing proved the entirely correct sequencing results. The luciferase activity was higher in pGL3-Cdc2-promoter/pRL-SV40 group than that of pGL3-Basic/pRL-SV40 group (1.591 5±0.199 8 vs 0.049 9±0.010 4). Conclusion pGL3-Cdc2-promoter can be transcribed and activated in U2OS cells. This study provided an important basis for screening and evaluation of anticancer drugs.
