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目的探讨Ets-1基因对人血清类黏蛋白1样蛋白3(ORMDL3)启动子的影响。方法 RT-PCR扩增Ets-1基因,将产物克隆至真核载体pcDNA3.1(+)内,构建含Ets-1基因的重组真核表达质粒pcDNA-Ets-1。重组质粒转染人胚肾HEK-293细胞,Western blot检测Ets-1蛋白表达;双荧光素酶报告基因检测Ets-1过表达后人ORMDL3启动子的活性。结果核酸序列分析及Western blot结果证实,构建含Ets-1基因的重组真核表达质粒pcDNA-Ets-1成功;过表达Ets-1基因能增强人ORMDL3启动子的活性。结论成功构建了含Ets-1基因的重组真核表达质粒。Ets-1基因可促进人ORMDL3启动子的转录活性。
Objective To investigate the effect of Ets-1 gene on the promoter of human serotype mucin-like protein 3 (ORMDL3). Methods Ets-1 gene was amplified by RT-PCR and cloned into eukaryotic vector pcDNA3.1 (+). The recombinant eukaryotic expression vector pcDNA-Ets-1 containing Ets-1 gene was constructed. The recombinant plasmid was transfected into human embryonic kidney HEK-293 cells and the expression of Ets-1 protein was detected by Western blot. The dual luciferase reporter gene was used to detect the activity of human ORMDL3 promoter after Ets-1 overexpression. Results Nucleic acid sequence analysis and Western blot confirmed that the recombinant eukaryotic expression plasmid pcDNA-Ets-1 containing Ets-1 gene was successfully constructed. The overexpression of Ets-1 gene could enhance the activity of human ORMDL3 promoter. Conclusion The recombinant eukaryotic expression plasmid containing Ets-1 gene was successfully constructed. The Ets-1 gene can promote the transcriptional activity of human ORMDL3 promoter.