Licht Alexander H, Raab Sabine, Hofmann Ursula, Breier Georg
Department of Molecular Cell Biology, Max Planck Institute for Physiological and Clinical Research, Bad Nauheim, Germany.
Dev Dyn. 2004 Feb;229(2):312-8. doi: 10.1002/dvdy.10416.
The use of the Cre-loxP recombination system allows the conditional inactivation of genes in mice. The availability of transgenic mice in which the Cre recombinase expression is highly cell type specific is a prerequisite to successfully use this system. We previously have characterized regulatory regions of the mouse flk-1 gene sufficient for endothelial cell-specific expression of the LacZ reporter gene in transgenic mice. These regions were fused to the Cre recombinase gene, and transgenic mouse lines were generated. In the resulting flk-1-Cre transgenic mice, specificity of Cre activity was determined by cross-breeding with the reporter mouse lines Rosa26R or CAG-CAT-LacZ. We examined double-transgenic mice at different stages of embryonic development (E9.5-E16.5) and organs of adult animals by LacZ staining. Strong endothelium-specific staining of most vascular beds was observed in embryos older than E11.5 in one or E13.5 in a second line. In addition, the neovasculature of experimental BFS-1 tumors expressed the transgene. These lines will be valuable for the conditional inactivation of floxed target genes in endothelial cells of the embryonic vascular system.
Cre-loxP重组系统的应用使得小鼠体内基因能够实现条件性失活。成功使用该系统的一个前提条件是要有Cre重组酶表达具有高度细胞类型特异性的转基因小鼠。我们之前已对小鼠flk-1基因的调控区域进行了表征,这些区域足以使LacZ报告基因在转基因小鼠中实现内皮细胞特异性表达。将这些区域与Cre重组酶基因融合,并构建了转基因小鼠品系。在所得的flk-1-Cre转基因小鼠中,通过与报告基因小鼠品系Rosa26R或CAG-CAT-LacZ杂交来确定Cre活性的特异性。我们通过LacZ染色检查了处于胚胎发育不同阶段(E9.5-E16.5)的双转基因小鼠以及成年动物的器官。在一条品系中,E11.5以上的胚胎以及另一条品系E13.5以上的胚胎中,大多数血管床均观察到强烈的内皮细胞特异性染色。此外,实验性BFS-1肿瘤的新生血管表达了转基因。这些品系对于胚胎血管系统内皮细胞中floxed靶基因的条件性失活将具有重要价值。