title: Efficient CRISPR/Cas9 plasmids for rapid and versatile genome editing in Drosophila.
abstract: The CRISPR-associated RNA-guided nuclease Cas9 has emerged as a powerful tool for genome engineering in a variety of organisms. To achieve efficient gene targeting rates in Drosophila, current approaches require either injection of in vitro transcribed RNAs or injection into transgenic Cas9-expressing embryos. We report a simple and versatile alternative method for CRISPR-mediated genome editing in Drosophila using bicistronic Cas9/sgRNA expression vectors. Gene targeting with this single-plasmid injection approach is as efficient as in transgenic nanos-Cas9 embryos and allows the isolation of targeted knock-out and knock-in alleles by molecular screening within 2 months. Our strategy is independent of genetic background and does not require prior establishment of transgenic flies.
authors.affiliation: Institute of Molecular Biotechnology of the Austrian Academy of Sciences, 1030 Vienna, Austria.
authors.forename: Joseph
authors.initials: J
authors.lastname: Gokcezade
authors.affiliation: Institute of Molecular Biotechnology of the Austrian Academy of Sciences, 1030 Vienna, Austria.
authors.forename: Grzegorz
authors.initials: G
authors.lastname: Sienski
authors.affiliation: Institute of Molecular Biotechnology of the Austrian Academy of Sciences, 1030 Vienna, Austria peter.duchek@imba.oeaw.ac.at.
authors.forename: Peter
authors.initials: P
authors.lastname: Duchek
mesh_terms: D000818:Animals; D064113:CRISPR-Cas Systems; D004330:Drosophila / Q000235:genetics*; D018390:Gene Targeting / Q000379:methods*; D049750:Genome, Insect; D010957:Plasmids / Q000235:genetics; D017393:RNA Editing
keywords: CRISPR; Cas9; Drosophila; HDR