Genetic dissection of neural circuits by Tol2 transposon-mediated Gal4 gene and enhancer trapping in zebrafish.

Targeted gene expression is a powerful approach to study the function of genes and cells in vivo. In Drosophila, the P element-mediated Gal4-UAS method has been successfully used for this purpose. However, similar methods have not been established in vertebrates.

Insertional mutagenesis by the Tol2 transposon-mediated enhancer trap approach generated mutations in two developmental genes: tcf7 and synembryn-like.

Gene trap and enhancer trap methods using transposon or retrovirus have been recently described in zebrafish. However, insertional mutants using these methods have not been reported. We report here development of an enhancer trap method by using the Tol2 transposable element and identification and characterization of insertional mutants.

Localization of ammonia transporter Rhcg1 in mitochondrion-rich cells of yolk sac, gill, and kidney of zebrafish and its ionic strength-dependent expression.

Members of the Rh glycoprotein family have been shown to be involved in ammonia transport in a variety of species. Here we show that zebrafish Rhcg1, a member of the Rh glycoprotein family, is highly expressed in the yolk sac, gill, and renal tubules. Molecular cloning and characterization indicate that zebrafish Rhcg1 shares 82% sequence identity with the pufferfish ortholog fRhcg1.

Genomic organization, alternative splicing, and multiple regulatory regions of the zebrafish fgf8 gene.

Fgf8 is among the members of the fibroblast growth factor (FGF) family that play pivotal roles in vertebrate development. In the present study, the genomic DNA of the zebrafish fgf8 gene was cloned to elucidate the regulatory mechanism behind the temporally and spatially restricted expression of the gene in vertebrate embryos. Structural analysis revealed that the exon-intron organization of fgf8 is highly conserved during vertebrate evolution, from teleosts to mammals.

Transposon-mediated gene trapping in zebrafish.

The Tol2 transposon system can create chromosomal insertions in the zebrafish germ lineage very efficiently. We constructed a Tol2-based gene trap vector, T2KSAG, which contains a splice accepter, the GFP gene and the polyA signal. In the pilot screen for gene trapping using T2KSAG, we identified 38 fish lines expressing GFP in specific organs and tissues.