Anming Meng

In this study, DNA constructs containing the putative zebrafish promoter sequences of GATA-1, an erythroid-specific transcription factor, and the green fluorescent protein reporter gene, were microinjected into single-cell zebrafish embryos. Erythroid-specific activity of the GATA-1 promoter was observed in living embryos during early development. Fluorescent circulating blood cells were detected in microinjected embryos 24 hours after fertilization and were still present in 2-month-old fish. Germline transgenic fish obtained from the injected founders continued to express green fluorescent protein in erythroid cells in the F1 and F2 generations. The green fluorescent protein expression patterns in transgenic fish were consistent with the pattern of GATA-1 mRNA expression detected by RNA in situ hybridization. These transgenic fish have allowed us to isolate, by fluorescence-activated cell sorting, the earliest erythroid progenitor cells from developing embryos for in vitro studies. By generating transgenic fish using constructs containing other zebrafish promoters and green fluorescent protein reporter gene, it should be possible to visualize the origin and migration of any lineage-specific progenitor cells in a living embryo.

The zebrafish (Danio rerio) has emerged as a powerful model to study vertebrate development and disease. Its short generation time makes it amenable to genetic manipulation and analysis, and its small size and high fecundity make it especially well suited for large-scale forward genetic and chemical screens. Fast-developing zebrafish embryos are transparent, facilitating live imaging of a variety of developmental processes in wild-type and mutant animals. ...\nThis brief document provides an updated set of guidelines regarding morpholino use in zebrafish that we anticipate will be of value for experimentalists as well as journal and grant reviewers, and decision makers.