ArticleGenesis (New York, N.Y. : 2000)2018
Use of two gRNAs for CRISPR/Cas9 improves bi-allelic homologous recombination efficiency in mouse embryonic stem cells.
Article in Genesis (New York, N.Y. : 2000), 2018. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 18 papers.
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Who cites it
18 citing papers in PubMed.
- Heritable transgenic schistosomes as a living platform for SARS-CoV-2 neutralizing antibody secretion.Nature communications · 2026Article
- Serum coating enables feeder-free culture of naive human pluripotent stem cells preserving developmental potential.The EMBO journal · 2026Article
- CRISPR-MiX: A pooled single-stranded donor strategy to enhance HDR efficiency in human iPSCs.Molecular therapy. Nucleic acids · 2026Article
- Transferable approaches to CRISPR-Cas9 induced genome editing in non-model insects: a brief guide.Frontiers in zoology · 2025Review
- CRISPR-based functional genomics for schistosomes and related flatworms.Trends in parasitology · 2024Review
- Analysis of the genome-editing activity of microinjected CRISPR/Cas9 ribonucleoprotein complexes inmicroPublication biology · 2024Article
- PCR-Based Strategy for Introducing CRISPR/Cas9 Machinery into Hematopoietic Cell Lines.Cancers · 2023Article
- Targeted insertion and reporter transgene activity at a gene safe harbor of the human blood fluke,Cell reports methods · 2023Article
- The CRISPR/Cas System in Human Cancer.Advances in experimental medicine and biology · 2023Article
- Biallelic and gene-wide genomic substitution for endogenous intron and retroelement mutagenesis in human cells.Nature communications · 2022Article
- EditingSynthetic biology (Oxford, England) · 2022Article
- Article
- Investigating CRISPR/Cas9 gene drive for production of disease-preventing prion gene alleles.PloS one · 2022Article
- Efficient mutagenesis targeting theAmerican journal of translational research · 2021Article
- Systematic analysis of factors that improve homologous direct repair (HDR) efficiency in CRISPR/Cas9 technique.PloS one · 2021Article
- Bi-allelic Homology-Directed Repair with Helper-Dependent Adenoviruses.Molecular therapy. Methods & clinical development · 2019Article
- Imaging endogenous synaptic proteins in primary neurons at single-cell resolution using CRISPR/Cas9.Molecular biology of the cell · 2019Article
- Optimized CRISPR/Cas9-mediated in vivo genome engineering applicable to monitoring dynamics of endogenous proteins in the mouse neural tissues.Scientific reports · 2019Article
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4 authors.
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Abstract
Targeted genome editing in mouse embryonic stem cells (ESCs) is a powerful resource to functionally characterize genes and regulatory elements. The use of the CRISPR/Cas9 genome editing approach has remarkably improved the time and efficiency of targeted recombination. However, the efficiency of this protocol is still far from ideal when aiming for bi-allelic homologous recombination, requiring at least two independent targeting recombination events. Here we describe an improved protocol that uses two gRNAs flanking the selected targeted region, leading to highly efficient homologous recombination in mouse ESCs. The bi-allelic recombination targeting efficiency is over 90% when using two gRNAs together with the inhibition of non-homologous end-joint repair. Moreover, this technique is compatible with the generation of knocked-in mice and the use of ESC-derived differentiation protocols, therefore facilitating and accelerating the gene targeting in mice and ESCs.
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