Restin gene promoter Cloning and transcriptional activity of HeLa cells.doc

 PAGE \* MERGEFORMAT 13 Restin gene promoter Cloning and transcriptional activity of HeLa cells Author: Rui-Hua Wang, Fu Haiyan, Yang Guodong, Lu Fan, Zhong-Liang Zhao [Keywords:] all-trans-retinoic acid Molecular cloning of restin gene promoter and its transcriptional activities in HeLa cells [Abstract] AIM: To clone the promoter and construct its luciferase report vector of our newly found gene restin and to analyze its response to the stimulation with alltransretinoic acid (ATRA). METHODS: Approximate 2000 bp up from the ATG of gene restin was bioinformatics analyzed with computer. The fragment was generated by polymerase chain reaction and cloned the upstream of luciferase report gene into plasmid pG5luc, replacing the five GAL4 binding sites and adenovirus promoter to generate the plasmid Rpluc. Rpluc was transfected into HeLa cells and luciferase activities were measured with or without the treatment of ATRA at indicated time. RESULTS: We successfully cloned and identified the promoter of the gene restin and constructed its luciferase report plasmid. Obvious response to ATRA was found when transfected into HeLa cells and a high expression of luciferase was detected when induced with ATRA. CONCLUSION: The luciferase report system of restin gene promoter we have constructed can be applied for further study of the function and transcriptional regulation of restin. [Keywords] restin; transcription regulation; promoter; alltransretinoic acid [Abstract] Objective: expanded cells resting factor (restin) gene promoter and construct the promoter of the fluorescence reporting system to explore its all-trans retinoic acid (alltrans retinoic acid, ATRA) to stimulate responses. Methods: ① right restin gene is about 2000 bp upstream genomic sequence analysis of biological information by computer. ② restin gene was amplified by PCR using the upstream sequence and cloned into the pG5luc, insert the luciferase reporter gene upstream, to replace the five GAL4 b