Research abstract
Letter abstract
Nature Biotechnology 26, 702 - 708 (2008)
Published online: 25 May 2008 | doi:10.1038/nbt1409
Heritable targeted gene disruption in zebrafish using designed zinc-finger nucleases
Yannick Doyon1,3, Jasmine M McCammon2,3, Jeffrey C Miller1, Farhoud Faraji1, Catherine Ngo1, George E Katibah1, Rainier Amora1, Toby D Hocking1, Lei Zhang1, Edward J Rebar1, Philip D Gregory1, Fyodor D Urnov1,2 & Sharon L Amacher2
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.
- Sangamo BioSciences, Inc., Pt. Richmond Tech Center, 501 Canal Blvd., Suite A100, Richmond, California 94804, USA.
- Department of Molecular and Cell Biology and Center for Integrative Genomics, University of California, Berkeley, California 94720-3200, USA.
- These authors contributed equally to this work.
Correspondence to: Sharon L Amacher2 e-mail: amacher@berkeley.edu
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