Effective targeted gene 'knockdown' in zebrafish

Citation
A. Nasevicius et Sc. Ekker, Effective targeted gene 'knockdown' in zebrafish, NAT GENET, 26(2), 2000, pp. 216-220
Citations number
30
Categorie Soggetti
Molecular Biology & Genetics
Journal title
NATURE GENETICS
ISSN journal
10614036 → ACNP
Volume
26
Issue
2
Year of publication
2000
Pages
216 - 220
Database
ISI
SICI code
1061-4036(200010)26:2<216:ETG'IZ>2.0.ZU;2-J
Abstract
The sequencing of the zebrafish genome should be completed by the end of 20 02. Direct assignment of function on the basis of this information would be facilitated by the development of a rapid, targeted 'knockdown' technology in this model vertebrate. We show here that antisense, morpholino-modified oligonucleotides(1) (morpholinos) are effective and specific translational inhibitors in zebrafish. We generated phenocopies of mutations of the gene s no tail (ref. 2), chordin (ref. 3), one-eyed-pinhead (ref. 4), nacre (ref . 5) and sparse (ref. 6), removing gene function from maternal through post -segmentation and organogenesis developmental stages. We blocked expression from a ubiquitous green fluorescent protein (GFP) transgene, showing that, unlike tissue-restricted limitations found with RNA-based interference in the nematode(7), all zebrafish cells readily respond to this technique. We also developed also morpholino-based zebrafish models of human disease. Mor pholinos targeted to the uroporphyrinogen decarboxylase gene(8) result in e mbryos with hepatoerythropoietic porphyria. We also used morpholinos for th e determination of new gene functions. We showed that embryos with reduced sonic hedgehog (ref. 9) signalling and reduced tiggy-winkle hedgehog(ref. 1 0) function exhibit partial cyclopia and other specific midline abnormaliti es, providing a zebrafish genetic model for the common human disorder holop rosencephaly. Conserved vertebrate processes and diseases are now amenable to a systematic, in vivo, reverse-genetic paradigm using zebrafish embryos.