The ability to create targeted mutations in specific genes, and therefore a loss-of-function condition, provides essential information about their endogenous functions during development and homeostasis. The discovery that CRISPR-Cas9 can target specific sequences according to base-pair complementarity and readily create knockouts in a desired gene has elevated the implementation of genetic analysis in numerous organisms. As CRISPR-Cas9 has become a powerful tool in a number of species, multiple methods for designing, creating, and screening editing efficiencies have been published, each of which has unique benefits. This chapter presents a cost-efficient, accessible protocol for creating knockout mutants in zebrafish using insertions/deletions (INDELS), from target site selection to mutant propagation, using basic laboratory supplies. The presented approach can be adapted to other systems, including any vertebrate species.
CITATION STYLE
Moravec, C. E., & Pelegri, F. J. (2019). An accessible protocol for the generation of CRISPR-Cas9 knockouts using INDELs in Zebrafish. In Methods in Molecular Biology (Vol. 1920, pp. 377–392). Humana Press Inc. https://doi.org/10.1007/978-1-4939-9009-2_23
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