ArticleNature communications2016
Real-time observation of DNA recognition and rejection by the RNA-guided endonuclease Cas9.
Article in Nature communications, 2016. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 154 papers.
What it found
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The abstract states no effect estimate the extractor could read, or names no intervention and outcome on the map, so this paper lights no cell and moves no belief. It is still indexed, cited and linked below.
The trial behind it
Trials whose registry record cites this paper, or whose number appears in the abstract. A trial that started after this paper was published is citing it as background, not reporting it.
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Who cites it
154 citing papers in PubMed.
- Exchange dynamics and kinetic control of gene regulation complexes.Nature reviews. Molecular cell biology · 2026Review
- Single-cell and in vivo profiling reveal heterogeneous and organ-specific CRISPR-Cas9 off-target and translocation outcomes.Nature communications · 2026Article
- Biophysical characterization ofSensors & diagnostics · 2026Article
- Article
- Operating CRISPR/Cas12a in a complex nucleic acid sequence background.Nucleic acids research · 2026Article
- A kinetic approach for mapping seed regions of CRISPR ribonucleoprotein and improving specificity.Nucleic acids research · 2026Article
- CRISPR-mediated targeting of theMolecular therapy. Advances · 2026Article
- Exploring CRISPR-Cas: The transformative impact of gene editing in molecular biology.Molecular therapy. Nucleic acids · 2025Review
- And… cut! - how conformational regulation of CRISPR-Cas effectors directs nuclease activity.The Biochemical journal · 2025Review
- Off-target interactions in the CRISPR-Cas9 Machinery: mechanisms and outcomes.Biochemistry and biophysics reports · 2025Review
- Small RNA Toxin-Assisted Evolution of GC-Preferred ErCas12a for Enhanced Genome Targeting Range.Advanced science (Weinheim, Baden-Wurttemberg, Germany) · 2025Article
- Rapid two-step target capture ensures efficient CRISPR-Cas9-guided genome editing.Molecular cell · 2025Article
- Mechanism of Cas9 inhibition by AcrIIA11.Nucleic acids research · 2025Article
- Dynamic basis of supercoiling-dependent DNA interrogation by Cas12a via R-loop intermediates.Nature communications · 2025Article
- Dynamic sampling of a surveillance state enables DNA proofreading by Cas9.Cell chemical biology · 2025Article
- Exploring CRISPR-Cas9 HNH-Domain-Catalyzed DNA Cleavage Using Accelerated Quantum Mechanical Molecular Mechanical Free Energy Simulation.Biochemistry · 2025Article
- Structural basis of Cas9 DNA interrogation with a 5' truncated sgRNA.Nucleic acids research · 2025Article
- Single-molecule perspectives of CRISPR/Cas systems: target search, recognition, and cleavage.BMB reports · 2025Review
- Analysis of metal-dependent DNA nicking activities by Cas endonucleases.Methods in enzymology · 2025Article
- Current status of recombinant duck enteritis virus vector vaccine research.Frontiers in veterinary science · 2025Review
94 more citing papers are in PubMed but not listed here.
Corrections and comments
PubMed lists nothing against this paper. Absence here is not a guarantee, only a check that was made.
Authors and funding
5 authors.
Funding
Abstract
Binding specificity of Cas9-guide RNA complexes to DNA is important for genome-engineering applications; however, how mismatches influence target recognition/rejection kinetics is not well understood. Here we used single-molecule FRET to probe real-time interactions between Cas9-RNA and DNA targets. The bimolecular association rate is only weakly dependent on sequence; however, the dissociation rate greatly increases from <0.006 s(-1) to >2 s(-1) upon introduction of mismatches proximal to protospacer-adjacent motif (PAM), demonstrating that mismatches encountered early during heteroduplex formation induce rapid rejection of off-target DNA. In contrast, PAM-distal mismatches up to 11 base pairs in length, which prevent DNA cleavage, still allow formation of a stable complex (dissociation rate <0.006 s(-1)), suggesting that extremely slow rejection could sequester Cas9-RNA, increasing the Cas9 expression level necessary for genome-editing, thereby aggravating off-target effects. We also observed at least two different bound FRET states that may represent distinct steps in target search and proofreading.
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Registered trials
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