Mutant rescue by BAC clone injection in zebrafish.
Genes essential for vertebrate body plan specification, organ development, and organ function are likely to be shared between mammals and zebrafish, but only in zebrafish have large-scale, genome-wide mutagenesis screens been conducted to isolate embryonic lethal mutations. Discovering the roles played by these disrupted genes requires their molecular characterization, which would be facilitated by assaying large cloned genomic DNAs for their potential to rescue mutant phenotypes. Here we demonstrate that bacterial artificial chromosomes can rescue the phenotype of floating head (flh) mutants. Homozygous flh embryos lack a differentiated notochord and have a reduced, discontinuous floor plate. Mutant embryos injected with genomic clones containing the flh+ gene often had stretches of several to many notochord cells overlaid by a row of floor-plate cells. In contrast, control mutant embryos injected with artificial chromosomes lacking the flh+ gene failed to form notochord. We conclude that the injection of large-insert genomic clones will speed the isolation of zebrafish genes disrupted by mutation and hence the identification of gene functions necessary for development of vertebrate embryos.
Yan
Y L
YL
Institute of Neuroscience, University of Oregon, Eugene 97403, USA.
Talbot
W S
WS
Egan
E S
ES
Postlethwait
J H
JH
D000818:Animals; D002876:Chromosomes, Bacterial / Q000235:genetics; D018507:Gene Expression Regulation, Developmental; D005801:Genes, Homeobox; D005816:Genetic Complementation Test; D018398:Homeodomain Proteins / Q000235:genetics*; D009154:Mutation; D009672:Notochord / Q000196:embryology; D014157:Transcription Factors / Q000235:genetics*; D015027:Zebrafish / Q000196:embryology* / Q000235:genetics*; D029961:Zebrafish Proteins
D018398:Homeodomain Proteins; D014157:Transcription Factors; D029961:Zebrafish Proteins; C096660:noto protein, zebrafish