Dissemin is shutting down on January 1st, 2025

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Nature Research, Nature Communications, 1(12), 2021

DOI: 10.1038/s41467-021-21427-6

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Cre-Controlled CRISPR mutagenesis provides fast and easy conditional gene inactivation in zebrafish

This paper is made freely available by the publisher.
This paper is made freely available by the publisher.

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Preprint: archiving allowed
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Postprint: archiving forbidden
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Published version: archiving allowed
Data provided by SHERPA/RoMEO

Abstract

AbstractConditional gene inactivation is a powerful tool to determine gene function when constitutive mutations result in detrimental effects. The most commonly used technique to achieve conditional gene inactivation employs the Cre/loxP system and its ability to delete DNA sequences flanked by two loxP sites. However, targeting a gene with two loxP sites is time and labor consuming. Here, we show Cre-Controlled CRISPR (3C) mutagenesis to circumvent these issues. 3C relies on gRNA and Cre-dependent Cas9-GFP expression from the same transgene. Exogenous or transgenic supply of Cre results in Cas9-GFP expression and subsequent mutagenesis of the gene of interest. The recombined cells become fluorescently visible enabling their isolation and subjection to various omics techniques. Hence, 3C mutagenesis provides a valuable alternative to the production of loxP-flanked alleles. It might even enable the conditional inactivation of multiple genes simultaneously and should be applicable to other model organisms amenable to single integration transgenesis.