Doherty Joanne R, Johnson Hamlet Michelle R, Kuliyev Emin, Mead Paul E
Department of Pathology, St. Jude Children's Research Hospital, Memphis, Tennessee 38105, USA.
Dev Dyn. 2007 Oct;236(10):2808-17. doi: 10.1002/dvdy.21321.
We have used the Sleeping Beauty (SB) transposable element to generate transgenic Xenopus laevis with expression of green fluorescent protein (GFP) in vascular endothelial cells using the frog flk-1 promoter. This is the first characterization of a SB-generated transgenic Xenopus that has tissue-restricted expression. We demonstrate that the transgene integrated into single genomic loci in two independent founder lines and is transmitted through the germline at the expected Mendelian frequencies. Transgene integration occurred through a noncanonical transposition process possibly reflecting Xenopus-specific interactions with the SB system. The transgenic animals express GFP in the same spatial and temporal pattern as the endogenous flk-1 gene throughout development and into adulthood. Overexpression of xVEGF122 in the transgenic animals disrupts vascular development that is visualized by fluorescent microscopy. These studies demonstrate the convenience of the SB system for generating transgenic animals and the utility of the xflk-1:GFP transgenic line for in vivo studies of vascular development.
我们利用睡美人(SB)转座元件,使用蛙flk-1启动子,在血管内皮细胞中表达绿色荧光蛋白(GFP),从而生成转基因非洲爪蟾。这是对具有组织限制性表达的SB生成的转基因非洲爪蟾的首次表征。我们证明,转基因整合到两个独立奠基系的单个基因组位点,并以预期的孟德尔频率通过种系传递。转基因整合通过非经典转座过程发生,这可能反映了非洲爪蟾与SB系统的特异性相互作用。在整个发育过程直至成年期,转基因动物中GFP的表达在空间和时间模式上与内源性flk-1基因相同。转基因动物中xVEGF122的过表达会破坏血管发育,这可通过荧光显微镜观察到。这些研究证明了SB系统在生成转基因动物方面的便利性,以及xflk-1:GFP转基因系在血管发育体内研究中的实用性。