title: Gene editing in mouse zygotes using the CRISPR/Cas9 system.
abstract: The generation of targeted mouse mutants is a key technology for biomedical research. Using the CRISPR/Cas9 system for induction of targeted double-strand breaks, gene editing can be performed in a single step directly in mouse zygotes. This article covers the design of knockout and knockin alleles, preparation of reagents, microinjection or electroporation of zygotes and the genotyping of pups derived from gene editing projects. In addition we include a section for the control of experimental settings by targeting the Rosa26 locus and PCR based genotyping of blastocysts.
authors.affiliation: German Center for Neurodegenerative Diseases (DZNE), Feodor-Lynen Str. 17, 81377 Munich, Germany; Helmholtz Zentrum München, German Research Center for Environmental Health, Institute of Developmental Genetics, Ingolstädter Landstr. 1, 85764 Neuherberg, Germany. Electronic address: benedikt.wefers@dzne.de.
authors.forename: Benedikt
authors.initials: B
authors.lastname: Wefers
authors.affiliation: Max-Delbrück-Centrum für Molekulare Medizin, Robert-Rössle Str. 10, 13125 Berlin, Germany; Berlin Institute of Health, Kapelle-Ufer 2, 10117 Berlin, Germany. Electronic address: sanum.bashir@mdc-berlin.de.
authors.forename: Sanum
authors.initials: S
authors.lastname: Bashir
authors.affiliation: Max-Delbrück-Centrum für Molekulare Medizin, Robert-Rössle Str. 10, 13125 Berlin, Germany. Electronic address: jana.rossius@mdc-berlin.de.
authors.forename: Jana
authors.initials: J
authors.lastname: Rossius
authors.affiliation: German Center for Neurodegenerative Diseases (DZNE), Feodor-Lynen Str. 17, 81377 Munich, Germany; Helmholtz Zentrum München, German Research Center for Environmental Health, Institute of Developmental Genetics, Ingolstädter Landstr. 1, 85764 Neuherberg, Germany; Technische Universität München-Weihenstephan, Chair of Developmental Genetics, c/o Helmholtz Zentrum München, Ingolstädter Landstr. 1, 85764 Neuherberg, Germany; Munich Cluster for Systems Neurology (SyNergy), Feodor-Lynen-Str. 17, 81377 Munich, Germany. Electronic address: wurst@helmholtz-muenchen.de.
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Wurst
authors.affiliation: Max-Delbrück-Centrum für Molekulare Medizin, Robert-Rössle Str. 10, 13125 Berlin, Germany; Berlin Institute of Health, Kapelle-Ufer 2, 10117 Berlin, Germany. Electronic address: ralf.kuehn@mdc-berlin.de.
authors.forename: Ralf
authors.initials: R
authors.lastname: Kühn
mesh_terms: D000818:Animals; D000831:Animals, Newborn; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D000076987:CRISPR-Associated Protein 9; D064113:CRISPR-Cas Systems; D064112:Clustered Regularly Interspaced Short Palindromic Repeats; D004247:DNA / Q000235:genetics / Q000378:metabolism; D053903:DNA Breaks, Double-Stranded; D059766:DNA End-Joining Repair; D018274:Electroporation / Q000379:methods; D004720:Endonucleases / Q000235:genetics* / Q000378:metabolism; D000072669:Gene Editing / Q000379:methods*; D055879:Gene Knock-In Techniques; D055786:Gene Knockout Techniques; D018390:Gene Targeting / Q000379:methods; D018014:Gene Transfer Techniques; D016678:Genome; D051379:Mice; D008822:Mice, Transgenic; D008845:Microinjections; D000094704:RNA, Guide, CRISPR-Cas Systems / Q000235:genetics* / Q000378:metabolism; D022661:RNA, Untranslated / Q000235:genetics / Q000378:metabolism; D059767:Recombinational DNA Repair; D015053:Zygote / Q000166:cytology / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; C110853:Gt(ROSA)26Sor non-coding RNA, mouse; D000094704:RNA, Guide, CRISPR-Cas Systems; D022661:RNA, Untranslated; D004247:DNA; D000076987:CRISPR-Associated Protein 9; C000606107:Cas9 endonuclease Streptococcus pyogenes; D004720:Endonucleases
keywords: CRISPR; Cas9; Gene editing; Mouse; Zygotes