Identification of GFP gene traps in zebrafish

描述（由申请人提供）：斑马鱼胚胎的光学清晰度，加上外部发育和基因突变的可用性，为胚胎发生和器官发生的分析提供了理想的工具。充分利用斑马鱼需要单个细胞或组织的标记物，这些标记物可以在活胚胎中可视化。我们将利用在medaka tol2转座子中设计的基因/增强子陷阱的有效转座来识别在特定细胞、组织或器官中表达GFP的细胞系。

DESCRIPTION (provided by applicant): The optical clarity of the zebrafish embryo, together with external development and the availability of genetic mutations, provides an ideal tool for analysis of embryogenesis and organogenesis. Full utilization of the zebrafish requires the availability of markers for individual cells or tissues that can be visualized in the living embryo. We will take advantage of efficient transposition of gene/enhancer traps engineered in the medaka tol2 transposon to identify lines expressing GFP in specific cells, tissues or organs. (1) We will identify upwards of 500 gene traps or enhancer traps strongly expressing GFP in specific cells, tissues or organs, in embryonic, larval, or adult stages. These GFP lines will be provided to individual researchers as they are isolated, archived as sperm or live stocks in the PI's facility, and submitted to the Zebrafish International Resource Center. (2) We will identify integration sites of isolated gene- or enhancer-trap lines, identify tagged genes, and determine whether the integration results in a visible or lethal mutation. (3) We will develop techniques for more efficient identification of gene or enhancer traps. We will test whether expression patterns in chimeric F0 injected animals reliably predicts patterns in F1s, or whether injection into eggs prior to fertilization will result in integration prior to the first mitotic DMA synthesis, resulting in non-mosaic expression. This work will enhance the use of zebrafish in developmental biology. It will identify live fluorescent markers for many of the cells, tissues and organs of the zebrafish embryo and larvae, identify the expression patterns for up to 500 tagged genes or enhancers, and generate recessive-lethal mutations in approximately 250 genes. This will enhance analysis of existing mutations, identify new molecularly identified mutations, and identify new phenomena in development of the vertebrate organism.