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Patterns of CRISPR/Cas9 activity in plants,animals and microbes
Authors:Julia Zischewski  Lucia Perez  Ludovic Bassié  Riad Nadi  Giobbe Forni  Sarah Boyd Lade  Erika Soto  Xin Jin  Vicente Medina  Gemma Villorbina  Pilar Muñoz  Gemma Farré  Rainer Fischer  Richard M Twyman  Teresa Capell  Paul Christou  Stefan Schillberg
Institution:1. Institute for Molecular Biotechnology, RWTH Aachen University, Aachen, Germany;2. Department of Plant Production and Forestry Science, School of Agrifood and Forestry Science and Engineering (ETSEA), University of Lleida‐Agrotecnio Center, Lleida, Spain;3. Fraunhofer Institute for Molecular Biology and Applied Ecology IME, Aachen, Germany;4. TRM Ltd, York, UK;5. ICREA, Catalan Institute for Research and Advanced Studies, Barcelona, Spain
Abstract:The CRISPR/Cas9 system and related RNA‐guided endonucleases can introduce double‐strand breaks (DSBs) at specific sites in the genome, allowing the generation of targeted mutations in one or more genes as well as more complex genomic rearrangements. Modifications of the canonical CRISPR/Cas9 system from Streptococcus pyogenes and the introduction of related systems from other bacteria have increased the diversity of genomic sites that can be targeted, providing greater control over the resolution of DSBs, the targeting efficiency (frequency of on‐target mutations), the targeting accuracy (likelihood of off‐target mutations) and the type of mutations that are induced. Although much is now known about the principles of CRISPR/Cas9 genome editing, the likelihood of different outcomes is species‐dependent and there have been few comparative studies looking at the basis of such diversity. Here we critically analyse the activity of CRISPR/Cas9 and related systems in different plant species and compare the outcomes in animals and microbes to draw broad conclusions about the design principles required for effective genome editing in different organisms. These principles will be important for the commercial development of crops, farm animals, animal disease models and novel microbial strains using CRISPR/Cas9 and other genome‐editing tools.
Keywords:genome editing  mutational signature  off‐target mutations  on‐target mutations  sgRNA design  site‐directed mutagenesis  species‐dependent effects
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