Construction of a luciferase reporter vector directed by DNA polβ promoter

Aim:To construct the luciferase reporter gene expression vector pGL3polβ/promoter containing wild-type DNA polβ promoter.Methods:The core fragment of DNA polβ gene promoter was amplified from human genomic DNA by polymerase chain reaction(PCR).The amplified fragment was subsequently cloned into pGL3-Neo-enhancer plasmid which contain neomycin-resistant gene.Then the recombinant was confirmed by restriction enzyme digestion,PCR analysis and DNA sequencing.Plasmid pGL3polβ/promoter was transfected into EC-1 cells by lipofectamine.The active of luciferase was detected,and the effect of polβ promoter was studied.Results:The result of DNA sequencing showed that the sequence of cloned polβ promoter was identical to that GenBank had reported and the fragment was inserted in right direction.The activity of luciferase was significantly increased in the pGL3polβ/promote transfected cells over 400 times than that in the pGL3-Neo-enhancer.Conclusion:The luciferase expression vector,pGL3polβ/promoter containing polβ promoter is constructed successfully.It will be foundation for studying its tissue-specificity and target gene therapy of tumor.