目的构建表达血清类黏蛋白1样蛋白3(ORMDL3)基因第一内含子中新启动子区的质粒,并研究其功能特征。方法以人基因组DNA为模板,PCR扩增ORMDL3基因第一内含子内新转录起始位点上下游776bp启动子区片段,酶切并连接此片段至pGL3-basic载体,构建含有ORMDL3基因第一内含子中新启动子区的基因重组体,并转染人胚肾293细胞。荧光素酶检测报告基因启动子的活性,计算相对活性单位。生物信息学方法分析转录因子结合位点。结果成功构建含有新启动子的重组质粒。与pGL3-basic空载体相比,新启动子活性明显增强,并定位于转录起始位点-363bp至-63bp之间,且该区域存在Sp1、GATA-1等转录因子的结合位点。结论 ORMDL3基因第一内含子中新转录起始位点上游具有启动子活性,在-363bp至-63bp区域存在转录调控元件,且Sp1、GATA-1等转录因子可能参与其转录调控。 Objective To construct a plasmid expressing a new promoter in the first intron of the serine mucin-like protein 3 (ORMDL3) gene and study its functional characteristics. METHODS: Human genomic DNA was used as a template to amplify the 776bp promoter region upstream and downstream of the new transcription initiation site in the first intron of ORMDL3 gene. The fragment was inserted into pGL3-basic vector to construct ORF gene fragment containing ORMDL3 gene A intron of the new promoter region of the recombinant gene and transfected human embryonic kidney 293 cells. Luciferase reporter assay gene promoter activity, calculate the relative activity unit. Bioinformatics Analysis of Transcription Factor Binding Sites. Results The recombinant plasmid containing the new promoter was successfully constructed. Compared with the pGL3-basic empty vector, the new promoter activity was significantly enhanced and located between -363bp and -63bp at the transcription start site, and the binding sites of transcription factors such as Sp1 and GATA-1 existed in this region. Conclusion The first intron of ORMDL3 gene possesses promoter activity upstream of transcriptional start site and transcriptional regulatory elements in -363bp to-63bp region. Transcription factors such as Sp1 and GATA-1 may be involved in transcriptional regulation.