ArticlePloS one2014
Optimization of genome engineering approaches with the CRISPR/Cas9 system.
Article in PloS one, 2014. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 89 papers.
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Who cites it
89 citing papers in PubMed.
- Double-stranded DNA donors and CRISPR-Cas9 for universal correction of mutations causing cystic fibrosis in human airway cells.Molecular therapy. Nucleic acids · 2026Article
- A bulk cell heterozygous knock-in strategy for targeted protein degradation.bioRxiv : the preprint server for biology · 2026Article
- Functional validation of a white pupae minimal gene construct in Ceratitis capitata (Diptera: Tephritidae).Insect science · 2026Article
- Precise CRISPR-Mediated Editing of theInternational journal of molecular sciences · 2026Article
- High efficiency CRISPR knock-in demonstrates that TCF1 is insufficient to reverse T cell exhaustion.Nature communications · 2026Article
- Gene modification: Exploring the potential in treating kidney diseases.Pharmacological research · 2026Review
- Compact bacterial recombination complexes drive efficient kilobase-scale knock-in in mammalian cells.Nucleic acids research · 2026Article
- Next-generation T cell immunotherapies engineered with CRISPR base and prime editing: challenges and opportunities.Nature reviews. Clinical oncology · 2025Review
- Optimization of gene knockout approaches and sgRNA selection in hPSCs with inducible Cas9 expression.Scientific reports · 2025Article
- ONE-STEP tagging: a versatile method for rapid site-specific integration by simultaneous reagent delivery.Nucleic acids research · 2025Article
- Genetic advancements in breast cancer treatment: a review.Discover oncology · 2025Review
- Development of efficient targeted insertion mediated by CRISPR-Cas12a and homology-directed repair in maize.Frontiers in genome editing · 2025Article
- The generation of stable transgenic lines in the human-infective nematode Strongyloides stercoralis.G3 (Bethesda, Md.) · 2024Article
- Current Strategies for Increasing Knock-In Efficiency in CRISPR/Cas9-Based Approaches.International journal of molecular sciences · 2024Review
- CRISPR-Cas9 immune-evasive hESCs are rejected following transplantation into immunocompetent mice.Frontiers in genome editing · 2024Article
- Article
- Enrichment strategies to enhance genome editing.Journal of biomedical science · 2023Review
- Gene Modulation with CRISPR-based Tools in Human iPSC-Cardiomyocytes.Stem cell reviews and reports · 2023Review
- Application of CRISPR/Cas9 Technology in Cancer Treatment: A Future Direction.Current oncology (Toronto, Ont.) · 2023Review
- Development of a Cellular Model Mimicking Specific HDAC Inhibitors.Methods in molecular biology (Clifton, N.J.) · 2023Article
29 more citing papers are in PubMed but not listed here.
Corrections and comments
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Authors and funding
5 authors.
Funding
Abstract
Designer nucleases such as TALENS and Cas9 have opened new opportunities to scarlessly edit the mammalian genome. Here we explored several parameters that influence Cas9-mediated scarless genome editing efficiency in murine embryonic stem cells. Optimization of transfection conditions and enriching for transfected cells are critical for efficiently recovering modified clones. Paired gRNAs and wild-type Cas9 efficiently create programmed deletions, which facilitate identification of targeted clones, while paired gRNAs and the Cas9D10A nickase generated smaller targeted indels with lower chance of off-target mutagenesis. Genome editing is also useful for programmed introduction of exogenous DNA sequences at a target locus. Increasing the length of the homology arms of the homology-directed repair template strongly enhanced targeting efficiency, while increasing the length of the DNA insert reduced it. Together our data provide guidance on optimal design of scarless gene knockout, modification, or knock-in experiments using Cas9 nuclease.
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Registered trials
Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.