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Cold Spring Harb Protoc. 2016 Feb 1;2016(2):pdb.prot090704. doi: 10.1101/pdb.prot090704.

Generation of Genetically Modified Mice Using the CRISPR-Cas9 Genome-Editing System.

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Institute for Immunology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104; and Division of Transplant Immunology, Department of Pathology and Laboratory Medicine, Children's Hospital of Philadelphia, University of Pennsylvania, Philadelphia, PA 19104.
The Jackson Laboratory for Genomic Medicine, Department of Genetics and Genome Sciences, University of Connecticut Health Center, Farmington, CT 06032.
Departments of Immunobiology, Yale University School of Medicine, New Haven, CT 06520.
Howard Hughes Medical Institute, Yale University School of Medicine, New Haven, CT 06520.
Contributed equally


Genetically modified mice are extremely valuable tools for studying gene function and human diseases. Although the generation of mice with specific genetic modifications through traditional methods using homologous recombination in embryonic stem cells has been invaluable in the last two decades, it is an extremely costly, time-consuming, and, in some cases, uncertain technology. The recently described CRISPR-Cas9 genome-editing technology significantly reduces the time and the cost that are required to generate genetically engineered mice, allowing scientists to test more precise and bold hypotheses in vivo. Using this revolutionary methodology we have generated more than 100 novel genetically engineered mouse strains. In the current protocol, we describe in detail the optimal conditions to generate mice carrying point mutations, chromosomal deletions, conditional alleles, fusion tags, or endogenous reporters.

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