安徽医科大学学报2017，Vol.52Issue(11)：1592-1596,5.DOI:10.19405/j.cnki.issn1000-1492.2017.11.003
人SAMHD1基因核心启动子区域的初步鉴定和分析
Identification and analysis of the core promoter region of human SAMHD1 gene
摘要
Abstract
Objective To identify the core promoter region of human antiviral SAMHD1 gene.Methods Genomic DNA was extracted from HepG2 cells,then 937,667,553 and 399 bp fragments before the translation start site of SAMHD1 gene were amplified by PCR from genomic DNA.Four fragments were inserted into the pMD19-T vector.After enzyme digestion and electrophoresis,DNA bands were separated and ligated into pGL3-Basic vector.The pGL3 plasmids containing four fragments were further identified by double enzyme digestion and DNA sequencing.The luciferase expression vector containing four fragments was co-transfected with pRL-TK into HepG2 cells and the promoter activities of four fragments were analyzed by luciferase assay.The transcription initiation site of SAMHD1 in HepG2 cells was identified by 5'RACE.Results Electrophoretic results showed that genomic DNA had been successfully extracted from HepG cells.The results of electrophoresis after PCR amplification showed that 937,667,553 and 399 bp fragments had been amplified.The results of electrophoresis showed that four fragments were successfully inserted into pMD19-T vector.Double enzyme digestion and DNA sequencing confirmed that luciferase expression vectors pGL3-937,pGL3-667,pGL3-553 and pGL3-399 were successfully constructed.Luciferase activity analysis showed that the SAMHD1 core promoter region was located in the 0 ～-399 region(the first base before ATG was set as-1).5'RACE results showed that the transcription initiation site of SAMHD1 in HepG2 cells was located at-101.Conclusion The core promoter of SAMHD1 is located at-101 ～-399 region,which lays a foundation for further study of SAMHD1 transcriptional regulation in hepatocytes.关键词
SAMHD1/核心启动子/转录起始位点/转录调控Key words
SAMHD1/the core promoter/transcription initiation site/transcriptional regulation分类
生物科学引用本文复制引用
张栋,李苗苗,董阳,刘星赟,应松成,方圣云,沈玉先..人SAMHD1基因核心启动子区域的初步鉴定和分析[J].安徽医科大学学报,2017,52(11):1592-1596,5.基金项目
国家自然科学基金(编号:81372576) （编号:81372576）
安徽省自然科学基金(编号:1508085MH158) （编号:1508085MH158）
安徽省博士后研究人员科研活动经费资助(编号:2015B066) （编号:2015B066）
