Harel, Itamar ORCID: 0000-0001-9749-8279, Valenzano, Dario Riccardo ORCID: 0000-0002-8761-8289 and Brunet, Anne (2016). Efficient genome engineering approaches for the short-lived African turquoise killifish. Nat. Protoc., 11 (10). S. 2010 - 2029. LONDON: NATURE PUBLISHING GROUP. ISSN 1750-2799
Full text not available from this repository.Abstract
A central challenge in experimental aging research is the lack of short-lived vertebrate models for genetic studies. Here we present a comprehensive protocol for efficient genome engineering in the African turquoise killifish (Nothobranchius furzeri), which is the shortest-lived vertebrate in captivity with a median life span of 4-6 months. By taking advantage of the clustered regularly interspaced short palindromic repeats/CRISPR-associated protein-9 nuclease (CRISPR/Cas9) system and the turquoise killifish genome, this platform enables the generation of knockout alleles via nonhomologous end joining (NHEJ) and knock-in alleles via homology-directed repair (HDR). We include guidelines for guide RNA (gRNA) target design, embryo injection and hatching, germ-line transmission and for minimizing off-target effects. We also provide strategies for Tol2-based transgenesis and large-scale husbandry conditions that are critical for success. Because of the fast life cycle of the turquoise killifish, stable lines can be generated as rapidly as 2-3 months, which is much faster than other fish models. This protocol provides powerful genetic tools for studying vertebrate aging and aging-related diseases.
Item Type: | Journal Article | ||||||||||||||||
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URN: | urn:nbn:de:hbz:38-261727 | ||||||||||||||||
DOI: | 10.1038/nprot.2016.103 | ||||||||||||||||
Journal or Publication Title: | Nat. Protoc. | ||||||||||||||||
Volume: | 11 | ||||||||||||||||
Number: | 10 | ||||||||||||||||
Page Range: | S. 2010 - 2029 | ||||||||||||||||
Date: | 2016 | ||||||||||||||||
Publisher: | NATURE PUBLISHING GROUP | ||||||||||||||||
Place of Publication: | LONDON | ||||||||||||||||
ISSN: | 1750-2799 | ||||||||||||||||
Language: | English | ||||||||||||||||
Faculty: | Unspecified | ||||||||||||||||
Divisions: | Unspecified | ||||||||||||||||
Subjects: | no entry | ||||||||||||||||
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Refereed: | Yes | ||||||||||||||||
URI: | http://kups.ub.uni-koeln.de/id/eprint/26172 |
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