Construction of red fluorescent protein reporter gene vector driven by mouse HMGB1 promoter and expression in mammalian cells
10.3760/cma.j.issn.1671-8925.2008.08.003
- VernacularTitle:小鼠HMGB1启动子红色荧光蛋白报告基因载体的构建及在真核细胞中的表达
- Author:
Jia XU
1
;
Li YANG
;
Zheng LIU
;
Juan WANG
;
Shao HUANG
;
Peng DENG
;
Yong JIANG
Author Information
1. 南方医科大学
- Keywords:
High mobility group box 1;
Promoter;
Red fluorescent protein;
Reporter gene
- From:
Chinese Journal of Neuromedicine
2008;7(8):767-770
- CountryChina
- Language:Chinese
-
Abstract:
Objective To construct the vector of mouse HMGB1 promoter-driven red fluorescent protein reporter gene so as to supply a tool for the study on the expression regulation of HMGB1 gene in mammalian cells and related signal transduction mechanism. Methods The mouse HMGB1 promoter sequence was subcloned into a red fluorescent protein reporter gene vector, pDsRed1-1. The recombinant vector pDsRed1-1-HMGB1p was then transfected into NIH3T3 cells by liposome, and the intracellular activity of HMGB1 promoter was observed under a fluorescence microscope in normal condition or after tumor necrosis factor-α (TNF-α) stimulation. Results The recombinant plasmid was confirmed by enzyme digestion and DNA sequence analysis. The vector was expressed with red fluorescence at a low level in the rest NIH3T3 cells, but the expression was highly increased by the stimulation with TNF-α. Conclusion A red fluorescent protein reporter gene vector driven by mouse HMGBI promoter is constructed successfully, which can be expressed in mammalian cells with a physiological response to TNF-α stimulation, thus providing an important and convenient tool for the study on the regulatory mechanisms of HMGB 1 gene expression.
