The African turquoise killifish Nothobranchius furzeri has recently gained interest as an emerging vertebrate model system for the study of aging, owing to its naturally short life span and generation time. Here, we provide a step-by-step guide for effective genome engineering using the CRISPR-Cas9 system to generate loss-of-function (i.e., knockout) alleles and for precise editing (i.e., knock-in) of short sequences into the genome. Using this approach, a new stable line can be created within several months. The killifish's tough chorion, rapid growth, and short life span are considered in this protocol and account for the key deviations from similar protocols in other fish models.
Bibliographical noteFunding Information:
We thank the Harel laboratory for stimulating discussion and feedback. Supported by the Zuckerman Program (I.H.); Israel Science Foundation (ISF) 2178/19 (I.H.); Israel Ministry of Science 3-17631 (I.H.), 3-16872 (I.H.), and 3-16591 (E.M); the Moore Foundation GBMF9341 (I.H.) and BSF-NSF 2020611 (I.H.); and the Israel Ministry of Agriculture 12-16-0010 (I.H).
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