Genome-Wide Assessment of Efficiency and Specificity in CRISPR/Cas9 Mediated Multiple Site Targeting in Arabidopsis
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Simultaneous multiplex mutation of large gene families using Cas9 has the potential to revolutionize agriculture and plant sciences. The targeting of multiple genomic sites at once raises concerns about the efficiency and specificity in targeting. The model Arabidopsis thaliana is widely used in basic plant research. Previous work has suggested that the Cas9 off-target rate in Arabidopsis is undetectable. Here we use deep sequencing on pooled plants simultaneously targeting 14 distinct genomic loci to demonstrate that multiplex targeting in Arabidopsis is highly specific to on-target sites with no detectable off-target events. In addition, chromosomal translocations are extremely rare. The high specificity of Cas9 in Arabidopsis makes this a reliable method for clean mutant generation with no need to enhance specificity or adopt alternate Cas9 variants.
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Next generation sequencing (NGS) is useful for sequencing large mutant populations at lower cost.
“The more advanced next generation sequencing (NGS) is useful for sequencing large mutant populations at lower cost (Peterson et al., 2016;Yang et al., 2022).”
From Genetic resources and precise gene editing for targeted improvement of barley abiotic stress tolerance (2023) · cites this paper as Peterson et al., 2016
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“The more advanced next generation sequencing (NGS) is useful for sequencing large mutant populations at lower cost (Peterson et al., 2016; Yang et al., 2022).”
- Authors
- Peterson, B. A., Haak, D. C., Nishimura, M. T., Teixeira, P. J. P. L., James, S. R., Dangl, J. L., & Nimchuk, Z. L.
- Affiliations
- University of North Carolina at Chapel Hill (US)
- Virginia Tech (US)
- Corresponding author
- Nimchuk, Zachary L.
- Venue
- PLOS ONE
- Publisher
- Public Library of Science (PLoS)
- Published
- 2016-09-13
- Volume
- 11
- Issue
- 9
- Pages
- e0162169
- ISSN
- 1932-6203 (ISSN-L)
- PMC
- PMC5021288
- OpenAlex
- W2519004356
- Cited by
- 172
- References
- 32
- Citations by year
- 2026: 3 · 2025: 10 · 2024: 10 · 2023: 9 · 2022: 16 · 2021: 19 · 2020: 37 · 2019: 21 · 2018: 26 · 2017: 20 · 2016: 1
- FWCI
- 11.78
- Type
- journal article
- Language
- en
- Open access
- gold
- doi.org
- Licence
- Locations
- PLoS ONE (publishedVersion, cc-by)
- PubMed Central (publishedVersion, cc-by)
- PubMed (publishedVersion)
- Carolina Digital Repository (University of North Carolina at Chapel Hill) (submittedVersion)
- DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
- Figshare (submittedVersion, cc-by)
- VTechWorks (Virginia Tech) (submittedVersion, cc-by)
- UNC Libraries
- Funders
- Directorate for Biological Sciences (IOS-1455607)
- Howard Hughes Medical Institute (GBMF3030)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › CRISPR and Genetic Engineering
- Life Sciences › Agricultural and Biological Sciences › Plant Science › Plant Genetic and Mutation Studies
- Life Sciences › Agricultural and Biological Sciences › Plant Science › Genetically Modified Organisms Research
- Keywords
- CRISPR/Cas9, multiplex genome editing, off-target effects, targeting specificity, Arabidopsis thaliana, chromosomal translocation
- MeSH terms
- Translocation, Genetic, Arabidopsis, Gene Targeting, Genome, Plant, Genome-Wide Association Study, Genetic Loci, CRISPR-Cas Systems
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