Abstract: Objective To construct a dual luciferase reporter expression vector of sterol regulatory element-binding protein 1 (SREBP1) gene and to screen natural active inhibitors for SREBP1 targets. Methods The bioinformatics approaches were applied to predict and analyze the promoter region of SREBP1 from 5′ upstream 5 000 bp regulatory region. The dual luciferase reporter expression vector was constructed by Gibson Assembly method. The recombinant plasmid pGL3- SREBP1-pro and control plasmid pRL-SV40 were co-transfected into HepG2 cells. The inhibitory effects of the transcriptional activity of SREBP1 with or without natural active inhibitors were detected. Results There were promoter sequences and transcription factor binding sites in the 5′ upstream 2 000 bp region of SREBP1. The recombinant plasmid was confirmed as a positive clone by enzyme digestion and sequencing. The plasmid was transiently transfected in HepG2 cells to detect promoter activity of SREBP1 by dual luciferase reporter system, and transcriptional activity of SREBP1 was further verified by natural active inhibitors. Conclusion The pGL3-SREBP1-pro dual luciferase reporter expression vector is successfully constructed and its activity is confirmed, which can be further used for the screening of natural active inhibitors for SREBP1 targets.

Key words: sterol regulatory element binding protein 1, transcription initiation site, genes, reporter, luciferases, emodin, anthraquinones, bioinformatics