ArticleFrontiers in genome editing2021
Efficiency, Specificity and Temperature Sensitivity of Cas9 and Cas12a RNPs for DNA-free Genome Editing in Plants.
Article in Frontiers in genome editing, 2021. 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, 29 citations in OpenAlex.
- CRISPR RNP-Mediated Transgene-Free Genome Editing in Plants: Advances, Challenges and Future Directions for Tree Species.Plant, cell & environment · 2026Review
- Cas9-Embedding Hyperactive TadA8e Confers Efficient and Highly Specific A-To-G Base Editing in Rice.Plant biotechnology journal · 2026Article
- Efficient CRISPR-Cas genome editing in brown algae.Cell reports methods · 2026Article
- Beyond GMOs: transgene-free gene-edited crops for global food security.Frontiers in plant science · 2026Review
- Highly efficient transgene-free ErCas12a RNP-protoplast genome editing and single-cell regeneration in Nicotiana benthamiana for glyco-engineering.Plant biotechnology journal · 2026Article
- Directed evolution expands CRISPR-Cas12a genome-editing capacity.Nucleic acids research · 2025Article
- RNAi and genome editing of sugarcane: Progress and prospects.The Plant journal : for cell and molecular biology · 2025Review
- Research Progress and Application of Miniature CRISPR-Cas12 System in Gene Editing.International journal of molecular sciences · 2024Review
- Unleashing the Potential of CRISPR/Cas9 Genome Editing for Yield-Related Traits in Rice.Plants (Basel, Switzerland) · 2024Review
- Generation of the transgene-free canker-resistant Citrus sinensis using Cas12a/crRNA ribonucleoprotein in the T0 generation.Nature communications · 2023Article
- Editing efficiencies with Cas9 orthologs, Cas12a endonucleases, and temperature in rice.Frontiers in genome editing · 2023Article
- Multiplex CRISPR/Cas9-mediated knockout of the phytoene desaturase gene in Coffea canephora.Scientific reports · 2022Article
- A Robust Expression and Purification Method for Production ofMethods and protocols · 2022Article
- Tissue Culture and Somatic Embryogenesis in Warm-Season Grasses-Current Status and Its Applications: A Review.Plants (Basel, Switzerland) · 2022Review
Corrections and comments
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Authors and funding
9 authors at 3 institutions in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Delivery of genome editing reagents using CRISPR-Cas ribonucleoproteins (RNPs) transfection offers several advantages over plasmid DNA-based delivery methods, including reduced off-target editing effects, mitigation of random integration of non-native DNA fragments, independence of vector constructions, and less regulatory restrictions. Compared to the use in animal systems, RNP-mediated genome editing is still at the early development stage in plants. In this study, we established an efficient and simplified protoplast-based genome editing platform for CRISPR-Cas RNP delivery, and then evaluated the efficiency, specificity, and temperature sensitivity of six Cas9 and Cas12a proteins. Our results demonstrated that Cas9 and Cas12a RNP delivery resulted in genome editing frequencies (8.7-41.2%) at various temperature conditions, 22°C, 26°C, and 37°C, with no significant temperature sensitivity. LbCas12a often exhibited the highest activities, while AsCas12a demonstrated higher sequence specificity. The high activities of CRISPR-Cas RNPs at 22° and 26°C, the temperature preferred by plant transformation and tissue culture, led to high mutagenesis efficiencies (34.0-85.2%) in the protoplast-regenerated calli and plants with the heritable mutants recovered in the next generation. This RNP delivery approach was further extended to pennycress (
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