ArticleGenome biology2018
Harnessing accurate non-homologous end joining for efficient precise deletion in CRISPR/Cas9-mediated genome editing.
Article in Genome biology, 2018. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 95 papers.
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Who cites it
95 citing papers in PubMed.
- Targeted gene editing ofMolecular therapy. Nucleic acids · 2026Article
- Cell-cycle-dependent regulation of DNA end resection by PLK1 and PLK3 without CtIP level modulation.iScience · 2026Article
- In vivo systematic detection of the outcomes of CRISPR-Cas9-mediated DNA repair in skeletal muscle stem cells.Molecular therapy : the journal of the American Society of Gene Therapy · 2026Article
- CRISPR/Cas9-Based Genome Editing: Understanding Differences in DNA Repair Pathways, Profiles, and Outcomes.International journal of molecular sciences · 2026Review
- 53BP1-independent Shieldin-BRCA1 antagonism at replication-coupled double-strand breaks.Nature communications · 2026Article
- CRISPR Gene Tagging for Illuminating Endogenous Protein Dynamics.International journal of molecular sciences · 2026Review
- Deep learning-driven prediction of on-target activity, off-target risk, and repair outcomes in CRISPR/Cas9: current landscape and multi-scale perspectives.Journal of translational medicine · 2026Review
- Safe and Efficient CRISPR Genome Editing of Primary Human T Cells Using a Droplet-Based Cell Mechanoporation Platform.Small (Weinheim an der Bergstrasse, Germany) · 2026Article
- Revealing the multiple faces of LRG1: gene expression, structure, function, and therapeutic potential.Journal of advanced research · 2026Review
- Conformational dynamics of CRISPR-Cas type I-F-HNH inform nickase engineering in a cascade scaffold.Nucleic acids research · 2026Article
- Targeting transthyretin by one Cas9 variant with superfidelity and broad compatibility.Science advances · 2026Article
- Macrophages promote aberrant DNA repair in multiple myeloma via the CXCL5/8-CXCR2 axis.Haematologica · 2026Article
- Programmable multistep CRISPR gene activation via control of RNA polymerase III termination.Science advances · 2025Article
- Efficient genome editing in Pseudomonas syringae pv. actinidiae using the CRISPR/FnCas12a system.Molecular horticulture · 2025Article
- Exonuclease-fused CRISPR-cas system enhances targeted genome editing for functional genomics in soybean.BMC plant biology · 2025Article
- Nanotechnology-Based Delivery of CRISPR/Cas9 for Cancer Treatment: A Comprehensive Review.Cells · 2025Review
- Late steps of allelic break-induced replication suppress tandem duplication associated with BRCA1 deficiency.Nucleic acids research · 2025Article
- Baculovirus Variant Detection from Transient CRISPR-Cas9-Mediated Disruption ofInternational journal of molecular sciences · 2025Article
- Single-guide RNA Cas9 and enhanced-deletion Cas9 rescue a recurrentMolecular therapy. Nucleic acids · 2025Article
- Robust prediction of synthetic gRNA activity and cryptic DNA repair by disentangling cellular CRISPR cleavage outcomes.Nature communications · 2025Article
35 more citing papers are in PubMed but not listed here.
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15 authors.
Funding
No grant is acknowledged in the PubMed record.
Abstract
backgroundMany applications of CRISPR/Cas9-mediated genome editing require Cas9-induced non-homologous end joining (NHEJ), which was thought to be error prone. However, with directly ligatable ends, Cas9-induced DNA double strand breaks may be repaired preferentially by accurate NHEJ.
resultsIn the repair of two adjacent double strand breaks induced by paired Cas9-gRNAs at 71 genome sites, accurate NHEJ accounts for about 50% of NHEJ events. This paired Cas9-gRNA approach underestimates the level of accurate NHEJ due to frequent + 1 templated insertions, which can be avoided by the predefined Watson/Crick orientation of protospacer adjacent motifs (PAMs). The paired Cas9-gRNA strategy also provides a flexible, reporter-less approach for analyzing both accurate and mutagenic NHEJ in cells and in vivo, and it has been validated in cells deficient for XRCC4 and in mouse liver. Due to high frequencies of precise deletions of defined "3n"-, "3n + 1"-, or "3n + 2"-bp length, accurate NHEJ is used to improve the efficiency and homogeneity of gene knockouts and targeted in-frame deletions. Compared to "3n + 1"-bp, "3n + 2"-bp can overcome + 1 templated insertions to increase the frequency of out-of-frame mutations. By applying paired Cas9-gRNAs to edit MDC1 and key 53BP1 domains, we are able to generate predicted, precise deletions for functional analysis. Lastly, a Plk3 inhibitor promotes NHEJ with bias towards accurate NHEJ, providing a chemical approach to improve genome editing requiring precise deletions.
conclusionsNHEJ is inherently accurate in repair of Cas9-induced DNA double strand breaks and can be harnessed to improve CRISPR/Cas9 genome editing requiring precise deletion of a defined length.
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