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Detection of on-target and off-target mutations generated by CRISPR/Cas9 and other sequence-specific nucleases
Open accessAbstract
The development of customizable sequence-specific nucleases such as TALENs, ZFNs and the powerful CRISPR/Cas9 system has revolutionized the field of genome editing. The CRISPR/Cas9 system is particularly versatile and has been applied in numerous species representing all branches of life. Regardless of the target organism, all researchers using sequence-specific nucleases face similar challenges: confirmation of the desired on-target mutation and the detection of off-target events. Here, we evaluate the most widely-used methods for the detection of on-target and off-target mutations in terms of workflow, sensitivity, strengths and weaknesses.
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“| Delivery methods improvement | Application in vivo [95–105] | LNP: Can be quickly degraded in vivo | Eliminate off-target effects |”
From Off-target effects in CRISPR/Cas9 gene editing (2023) · cites this paper as [103]
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“| Delivery methods improvement | Application in vivo [95–105] | AAV: Last for years in terminally differentiated cell types | May cause accumulation of off-target mutations |”
From Off-target effects in CRISPR/Cas9 gene editing (2023) · cites this paper as [103]
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“When the indels are deposited at the coding sequences of a given gene, NHEJ can cause frameshift mutations, resulting in non-sense-mediated mRNA decay and gene silencing (Wang et al., 2016a; Zischewski et al., 2017; Lindeboom et al., 2019).”
From Off-target effects in CRISPR/Cas9 gene editing (2023) · cites this paper as Zischewski et al., 2017
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“| Delivery methods improvement | Application in vivo [95–105] | LNP: Can be quickly degraded in vivo | Eliminate off-target effects |”
From Off-target effects in CRISPR/Cas9 gene editing (2023) · cites this paper as [103]
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“| Delivery methods improvement | Application in vivo [95–105] | AAV: Last for years in terminally differentiated cell types | May cause accumulation of off-target mutations |”
From Off-target effects in CRISPR/Cas9 gene editing (2023) · cites this paper as [103]
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CRISPR/Cas9 therapeutics: progress and prospects
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BackgroundMethodCRISPR-Cas9 genome editing induces megabase-scale chromosomal truncations
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Metadata
- Authors
- Zischewski, J., Fischer, R., & Bortesi, L.
- Affiliations
- RWTH Aachen University (DE)
- Fraunhofer Institute for Molecular Biology and Applied Ecology (DE)
- Corresponding author
- Bortesi, Luisa
- Venue
- Biotechnology Advances
- Publisher
- Elsevier BV
- Published
- 2017-01
- Volume
- 35
- Issue
- 1
- Pages
- 95-104
- ISSN
- 0734-9750 (ISSN-L)
- OpenAlex
- W2565555627
- Cited by
- 323
- References
- 78
- Citations by year
- 2026: 14 · 2025: 29 · 2024: 32 · 2023: 31 · 2022: 35 · 2021: 41 · 2020: 40 · 2019: 46 · 2018: 37 · 2017: 18
- FWCI
- 3.04
- Type
- journal article
- Language
- en
- Open access
- hybrid
- doi.org
- Licence
- Locations
- Biotechnology Advances (publishedVersion, cc-by-nc-nd)
- PubMed (publishedVersion)
- Research Publications (Maastricht University) (publishedVersion)
- RWTH Publications (RWTH Aachen) (publishedVersion, other-oa)
- Fraunhofer-Publica (Fraunhofer-Gesellschaft) (submittedVersion)
- Funders
- European Research Council (269110)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › CRISPR and Genetic Engineering
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA regulation and disease
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › DNA and Nucleic Acid Chemistry
- Keywords
- CRISPR/Cas9, off-target mutations, TALEN, zinc finger nucleases, sequence-specific nucleases, genome editing, sensitivity
- MeSH terms
- Gene Editing, Animals, Base Sequence, Cell Line, Deoxyribonucleases, Humans, Mutation, Mice, CRISPR-Cas Systems
About these checks
Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.
Statistics and arithmetic are recomputed, not judged.
Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.
What counts as a claim ›How we measure confidence ›How we check this ›
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