Molecular cloning of restin gene promoter and its transcriptional activities in HeLa cells

    WANG Ruiª²Hua, FU Haiª²Yan£¬YANG Guoª²Dong, LU Fan£¬ZHAO Zhongª²Liang

    Department of Biochemistry and Molecular Biology, State Key Laboratory of Cancer Biology, School of Basic Medicine, Fourth Military Medical University, Xi¬ðan 710033, China

    ¡¾Abstract¡¿ 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 allª²transª²retinoic 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 pG5ª²luc, replacing the five GALª²4 binding sites and adenovirus promoter to generate the plasmid Rpª²luc. Rpª²luc 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. ......