ArticlePloS one2019
Generation of deletions and precise point mutations in Dictyostelium discoideum using the CRISPR nickase.
Article in PloS one, 2019. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 14 papers.
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Who cites it
14 citing papers in PubMed.
- Unlocking CRISPR-Cas9 editing for widely diverse Dictyostelid species.Molecular systems biology · 2026Article
- A nanobody-based degron system for targeted protein knockdown in Dictyostelium discoideum.BMC molecular and cell biology · 2026Article
- Genome editing across Dictyostelia species enables comparative functional genetics of social amoebas.Scientific reports · 2026Article
- The IQGAP-related RasGAP IqgC regulates cell-substratum adhesion in Dictyostelium discoideum.Cellular & molecular biology letters · 2025Article
- Efficient endogenous protein labelling in Dictyostelium using CRISPR/Cas9 knock-in and split fluorescent proteins.PloS one · 2025Article
- Use of paired Cas9-NG nickase and truncated sgRNAs for single-nucleotide microbial genome editing.Frontiers in genome editing · 2024Article
- CRISPR/Cas9-based genome-wide screening of Dictyostelium.Scientific reports · 2022Article
- Knock-in and precise nucleotide substitution using near-PAMless engineered Cas9 variants in Dictyostelium discoideum.Scientific reports · 2021Article
- Development of a rational strategy for integration of lactate dehydrogenase A suppression into therapeutic algorithms for head and neck cancer.British journal of cancer · 2021Article
- CRISPR Toolbox for Genome Editing inFrontiers in cell and developmental biology · 2021Article
- Novel Single-Cell and High-Throughput Microscopy Techniques to Monitor Dictyostelium discoideum-Mycobacterium marinum Infection Dynamics.Methods in molecular biology (Clifton, N.J.) · 2021Article
- Review
- Dictyostelium discoideum as a non-mammalian biomedical model.Microbial biotechnology · 2021Review
- Calmodulin and Calmodulin Binding Proteins inInternational journal of molecular sciences · 2020Review
Corrections and comments
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Authors and funding
3 authors.
Funding
No grant is acknowledged in the PubMed record.
Abstract
The CRISPR/Cas9 system enables targeted genome modifications across a range of eukaryotes. Although we have reported that transient introduction of all-in-one vectors that express both Cas9 and sgRNAs can efficiently induce multiple gene knockouts in Dictyostelium discoideum, concerns remain about off-target effects and false-positive amplification during mutation detection via PCR. To minimise these effects, we modified the system to permit gene deletions of greater than 1 kb via use of paired sgRNAs and Cas9 nickase. An all-in-one vector expressing the Cas9 nickase and sgRNAs was transiently introduced into D. discoideum, and the resulting mutants showed long deletions with a relatively high efficiency of 10-30%. By further improving the vector, a new dual sgRNA expression vector was also constructed to allow simultaneous insertion of two sgRNAs via one-step cloning. By applying this system, precise point mutations and genomic deletions were generated in the target locus via simultaneous introduction of the vector and a single-stranded oligonucleotide template without integrating a drug resistance cassette. These systems enable simple and straightforward genome editing that requires high specificity, and they can serve as an alternative to the conventional homologous recombination-based gene disruption method in D. discoideum.
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