2C-Cas9: a versatile tool for clonal analysis of gene function
- Vincenzo Di Donato1,4,
- Flavia De Santis1,4,
- Thomas O. Auer1,5,
- Noé Testa1,
- Héctor Sánchez-Iranzo2,
- Nadia Mercader2,
- Jean-Paul Concordet3 and
- Filippo Del Bene1
- 1Institut Curie, PSL Research University, INSERM U 934, CNRS UMR3215, F-75005, Paris, France;
- 2Department of Cardiovascular Development and Repair, Atherothrombosis and Imaging, Centro Nacional de Investigaciones Cardiovasculares (CNIC), 28028 Madrid, Spain;
- 3Muséum National d'Histoire Naturelle, INSERM U1154, CNRS UMR 7196, Paris F-75231, France
- Corresponding authors: filippo.del-bene{at}curie.fr, jean-paul.concordet{at}mnhn.fr
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↵4 These authors contributed equally to this work.
Abstract
CRISPR/Cas9-mediated targeted mutagenesis allows efficient generation of loss-of-function alleles in zebrafish. To date, this technology has been primarily used to generate genetic knockout animals. Nevertheless, the study of the function of certain loci might require tight spatiotemporal control of gene inactivation. Here, we show that tissue-specific gene disruption can be achieved by driving Cas9 expression with the Gal4/UAS system. Furthermore, by combining the Gal4/UAS and Cre/loxP systems, we establish a versatile tool to genetically label mutant cell clones, enabling their phenotypic analysis. Our technique has the potential to be applied to diverse model organisms, enabling tissue-specific loss-of-function and phenotypic characterization of live and fixed tissues.
Footnotes
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[Supplemental material is available for this article.]
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Article published online before print. Article, supplemental material, and publication date are at http://www.genome.org/cgi/doi/10.1101/gr.196170.115.
- Received June 22, 2015.
- Accepted February 24, 2016.
This article is distributed exclusively by Cold Spring Harbor Laboratory Press for the first six months after the full-issue publication date (see http://genome.cshlp.org/site/misc/terms.xhtml). After six months, it is available under a Creative Commons License (Attribution-NonCommercial 4.0 International), as described at http://creativecommons.org/licenses/by-nc/4.0/.
