斑马鱼ID1启动子体内外活性分析

摘要

目的:分析斑马鱼分化抑制因子1(ID1)启动子在体内、外的活性情况,为研究ID1基因表达调控奠定基础.方法:通过构建斑马鱼ID1启动子不同长度萤光素酶报告质粒,分别以鲤鱼上皮瘤细胞(EPC)和斑马鱼胚胎为材料进行体外转染和体内注射,并采用双萤光素酶报告系统对不同长度ID1启动子活性进行评估.结果:在体内、外实验中,不同长度ID1启动子活性均表现出随启动子长度变化而变化的情况;启动子活性变化趋势在体内和体外并不一致,呈现出较大差异性.结论:ID1启动子活性随长度变化而表现不同,说明启动子上存在许多顺式调控元件,且体内、外ID1活性变化可能受多种因素的影响,提示不同细胞中的调控机制有所不同.%Objective:To explore the biological activity of zebrafish ID1(inhibitor of differentiation/DNA binding protein 1) promoter through in vitro and in vivo study,and to provide basis for expression regulation study for ID1 gene.Methods:A set of zebrafish ID1 promoter with different length were amplified from genomic DNA and inserted to luciferase vector pGL3-basic,and subsequently were transfected into endothelial progenitor cells(EPC) and microinected into zebrafish embyo at 1～2 cell stage.The promoter activity was determined with the dual-luciferase reporter system assay.Results:The ID1 promoter activity varied as the length change,and the in vivo and in vtro studies presented great different paterns.Conclusion:The different promoter activity between in vitro and in vivo study indicate that there are a lot of cis-acting element located at the ID1 promoter,and the regulatory mechanism of ID1 promoter are different in different types of cells.