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Integrated computational guide design, execution, and analysis of arrayed and pooled CRISPR genome editing experiments

Matthew C. Canver, Maximilian Haeussler, Daniel E. Bauer, Stuart H. Orkin, Neville E. Sanjana, Ophir Shalem, Guo-Cheng Yuan, Feng Zhang, Jean-Paul Concordet, Luca Pinello
doi: https://doi.org/10.1101/125245
Matthew C. Canver
1Division of Hematology/Oncology, Boston Children’s Hospital, Department of Pediatric Oncology, Dana-Farber Cancer Institute, Harvard Stem Cell Institute, Department of Pediatrics, Harvard Medical School, Boston, Massachusetts, USA
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Maximilian Haeussler
2Santa Cruz Genomics Institute, MS CBSE, University of California, Santa Cruz, California, USA
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Daniel E. Bauer
1Division of Hematology/Oncology, Boston Children’s Hospital, Department of Pediatric Oncology, Dana-Farber Cancer Institute, Harvard Stem Cell Institute, Department of Pediatrics, Harvard Medical School, Boston, Massachusetts, USA
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Stuart H. Orkin
1Division of Hematology/Oncology, Boston Children’s Hospital, Department of Pediatric Oncology, Dana-Farber Cancer Institute, Harvard Stem Cell Institute, Department of Pediatrics, Harvard Medical School, Boston, Massachusetts, USA
3Howard Hughes Medical Institute, Boston, Massachusetts, USA
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Neville E. Sanjana
4New York Genome Center and Department of Biology, New York University, New York City, New York, USA
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Ophir Shalem
5Raymond G. Perelman Center for Cellular and Molecular Therapeutics, Children’s Hospital of Philadelphia, Department of Genetics, University of Pennsylvania, Philadelphia, Pennsylvania, USA
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Guo-Cheng Yuan
6Department of Biostatistics and Computational Biology, Dana-Farber Cancer Institute and Harvard T.H. Chan School of Public Health, Boston, Massachusetts, USA
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Feng Zhang
7The Broad Institute, Cambridge, Massachusetts, USA.
8McGovern Institute for Brain Research, Department of Brain and Cognitive Sciences, Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA.
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Jean-Paul Concordet
9INSERM U1154, CNRS UMR 7196, Muséum National d’Histoire Naturelle, Paris, France.
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Luca Pinello
10Department of Molecular Pathology, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts, USA
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  • For correspondence: lpinello@mgh.harvard.edu
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ABSTRACT

CRISPR genome editing experiments offer enormous potential for evaluation of genomic loci using arrayed or pooled lentiviral libraries of single guide RNAs (sgRNAs). Numerous computational tools are available to help design sgRNAs with optimal on-target efficiency and minimal off-target effects. In addition, a few computational tools have been developed to analyze data resulting from genome editing experiments. However, these tools are typically developed in isolation and oftentimes are not readily translatable into laboratory-based experiments. Here we present a protocol that describes in detail both the computational and the benchtop implementation of an arrayed and/or pooled CRISPR genome editing experiment. This protocol provides instructions for sgRNA design with CRISPOR, experimental implementation, and analysis of the resulting high-throughput sequencing data with CRISPResso.

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  • ↵* Co-first authors

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The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC-ND 4.0 International license.
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Posted April 07, 2017.
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Integrated computational guide design, execution, and analysis of arrayed and pooled CRISPR genome editing experiments
Matthew C. Canver, Maximilian Haeussler, Daniel E. Bauer, Stuart H. Orkin, Neville E. Sanjana, Ophir Shalem, Guo-Cheng Yuan, Feng Zhang, Jean-Paul Concordet, Luca Pinello
bioRxiv 125245; doi: https://doi.org/10.1101/125245
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Integrated computational guide design, execution, and analysis of arrayed and pooled CRISPR genome editing experiments
Matthew C. Canver, Maximilian Haeussler, Daniel E. Bauer, Stuart H. Orkin, Neville E. Sanjana, Ophir Shalem, Guo-Cheng Yuan, Feng Zhang, Jean-Paul Concordet, Luca Pinello
bioRxiv 125245; doi: https://doi.org/10.1101/125245

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