关键词: [关键词] B7-H1, 启动子, 双荧光素酶报告基因检测, 活性测定

Abstract: Objective To construct luciferase reporter gene vectors containing B7-H1 promoter region，and detect promoter activity in HepG2 cells. Methods Firstly several fragments of the B7-H1 gene 5’-UTR promoters were amplified by PCR and cloned into pGL3 basic vector. After the identification by digestion and sequencing on the recombinant basic vector, they were named p88，p175, p203, p398, p723, and p960 respectively. Then one of the human HCC cell lines, HepG2 cells, was transfected with these constructed plasmids. The expression of luciferase before and after the addition of IFN-γ was detected by Dual-Luciferase(LUC) Reporter Assay System kit. Results Five luciferase reporter gene vectors containing B7-H1 promoter region were constructed successfully identified by restriction endonuclease analysis and sequences analysis. The recombinant constructs were transiently transfected into HepG2 cells. After the addition of IFN-γ, the results of LUC detection assay showed that the expression of LUC was significantly enhanced by P175, P203, P398, P723 or P960，but not influenced by P88. Conclusion Luciferase reporter gene vectors containing B7-H1 promoter region were constructed successfully，which may be valuable for further study on B7-H1 promoter activity and molecular mechanism.

Key words: [Key words] B7-H1, promoter, Dual-Luciferase Reporter Assay System, activity assay