Unbiased detection of off-target cleavage by CRISPR-Cas9 and TALENs using integrase-defective lentiviral vectors
Abstract
The utility of CRISPR-Cas9 and TALENs for genome editing may be compromised by their off-target activity. We show that integrase-defective lentiviral vectors (IDLVs) can detect such off-target cleavage with a frequency as low as 1%. In the case of Cas9, we find frequent off-target sites with a one-base bulge or up to 13 mismatches between the single guide RNA (sgRNA) and its genomic target, which refines sgRNA design.
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IDLV displays a propensity to integrate into the vicinity of DSBs.
“One such labeling tool is called the integrase-defective lentiviral vectors (IDLV), which displays the propensity to integrate into the vicinity of DSBs (Wang et al., 2015; Kim et al., 2019).”
From Off-target effects in CRISPR/Cas9 gene editing (2023) · cites this paper as Wang et al., 2015
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“[29] found off-target sites harboring one-base bulge or up to 13 mismatches between the gRNA and its genomic target.”
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- Authors
- Wang, X., Wang, Y., Wu, X., Wang, J., Wang, Y., Qiu, Z., Chang, T., Huang, H., Lin, R.-J., & Yee, J.-K.
- Affiliations
- City Of Hope National Medical Center (US)
- First Affiliated Hospital Zhejiang University (CN)
- Corresponding author
- Yee, Jiing-Kuan
- Venue
- Nature Biotechnology
- Published
- 2015-01-19
- Volume
- 33
- Issue
- 2
- Pages
- 175-178
- ISSN
- 0733-222X (ISSN-L)
- 1087-0156
- 1546-1696
- OpenAlex
- W2051255304
- Cited by
- 442
- References
- 20
- Citations by year
- 2026: 5 · 2025: 17 · 2024: 18 · 2023: 25 · 2022: 31 · 2021: 38 · 2020: 47 · 2019: 36 · 2018: 46 · 2017: 57 · 2016: 70 · 2015: 52
- FWCI
- 32.55
- Type
- journal article
- Language
- en
- Open access
- closed
- Licence
- www.springer.com/tdm (tdm)
- Locations
- Nature Biotechnology (publishedVersion)
- PubMed (publishedVersion)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › CRISPR and Genetic Engineering
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Virus-based gene therapy research
- Health Sciences › Medicine › Oncology › CAR-T cell therapy research
- Keywords
- off-target cleavage, CRISPR/Cas9, TALEN, integrase-defective lentiviral vectors, genome editing, single guide RNA design, mismatch tolerance
- MeSH terms
- Genetic Vectors, Humans, Genome, Human, Lentivirus, RNA Editing, Integrases, HEK293 Cells, High-Throughput Nucleotide Sequencing, CRISPR-Cas Systems
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