ZFIN ID: ZDB-PUB-080527-24
Heritable targeted gene disruption in zebrafish using designed zinc-finger nucleases
Doyon, Y., McCammon, J.M., Miller, J.C., Faraji, F., Ngo, C., Katibah, G.E., Amora, R., Hocking, T.D., Zhang, L., Rebar, E.J., Gregory, P.D., Urnov, F.D., and Amacher, S.L.
Date: 2008
Source: Nat. Biotechnol.   26(6): 702-708 (Journal)
Registered Authors: Amacher, Sharon, McCammon, Jasmine
Keywords: none
MeSH Terms:
  • Animals
  • Animals, Genetically Modified/physiology*
  • Deoxyribonucleases/genetics
  • Gene Targeting/methods*
  • Genetic Engineering/methods*
  • Mutagenesis, Site-Directed/methods*
  • Protein Engineering/methods
  • Zebrafish/genetics*
  • Zebrafish Proteins/genetics*
  • Zinc Fingers/genetics*
PubMed: 18500334 Full text @ Nat. Biotechnol.
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ABSTRACT
We describe the use of zinc-finger nucleases (ZFNs) for somatic and germline disruption of genes in zebrafish (Danio rerio), in which targeted mutagenesis was previously intractable. ZFNs induce a targeted double-strand break in the genome that is repaired to generate small insertions and deletions. We designed ZFNs targeting the zebrafish golden and no tail/Brachyury (ntl) genes and developed a budding yeast-based assay to identify the most active ZFNs for use in vivo. Injection of ZFN-encoding mRNA into one-cell embryos yielded a high percentage of animals carrying distinct mutations at the ZFN-specified position and exhibiting expected loss-of-function phenotypes. Over half the ZFN mRNA-injected founder animals transmitted disrupted ntl alleles at frequencies averaging 20%. The frequency and precision of gene-disruption events observed suggest that this approach should be applicable to any loci in zebrafish or in other organisms that allow mRNA delivery into the fertilized egg.
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