ArticleScientific reports2020
Strategies to produce T-DNA free CRISPRed fruit trees via Agrobacterium tumefaciens stable gene transfer.
Article in Scientific reports, 2020. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 31 papers.
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Who cites it
31 citing papers in PubMed.
- Transgene-free genome editing in plants.aBIOTECH · 2026Review
- Article
- A Fruit-Pulp-Derived Callus-Level Agrobacterium-Mediated Transformation Platform forPlants (Basel, Switzerland) · 2026Article
- Reduced stomatal density improves water-use efficiency in grapevine under climate scenarios of decreased water availability.Plant cell reports · 2025Article
- CRISPR/Cas9-driven double modification of grapevine MLO6-7 imparts powdery mildew resistance, while editing of NPR3 augments powdery and downy mildew tolerance.The Plant journal : for cell and molecular biology · 2025Article
- Mini review: Apple improvement, traditional approaches, biotechnology options, and regulatory considerations.Frontiers in bioengineering and biotechnology · 2025Review
- Overview on Current Selectable Marker Systems and Novel Marker Free Approaches in Fruit Tree Genetic Engineering.International journal of molecular sciences · 2024Review
- Rare but diverse off-target and somatic mutations found in field and greenhouse grown trees expressing CRISPR/Cas9.Frontiers in bioengineering and biotechnology · 2024Article
- Citrus threat huanglongbing (HLB) - Could the rootstock provide the cure?Frontiers in plant science · 2024Article
- Application of CRISPR/Cas-based gene-editing for developing better banana.Frontiers in bioengineering and biotechnology · 2024Review
- CRISPR/Cas as a Genome-Editing Technique in Fruit Tree Breeding.International journal of molecular sciences · 2023Review
- Insights into the molecular mechanisms underlying responses of apple trees to abiotic stresses.Horticulture research · 2023Article
- Targeted mutagenesis with sequence-specific nucleases for accelerated improvement of polyploid crops: Progress, challenges, and prospects.The plant genome · 2023Review
- Review
- Applications of CRISPR/Cas genome editing in economically important fruit crops: recent advances and future directions.Molecular horticulture · 2023Review
- Efficient Agrobacterium-mediated transformation and genome editing ofFrontiers in plant science · 2023Article
- DNA-free genome editing in grapevine using CRISPR/Cas9 ribonucleoprotein complexes followed by protoplast regeneration.Horticulture research · 2023Article
- Lignin engineering in forest trees: From gene discovery to field trials.Plant communications · 2022Review
- Spontaneous, Artificial, and Genome Editing-Mediated Mutations inInternational journal of molecular sciences · 2022Review
- Powdery Mildew Resistance Genes in Vines: An Opportunity to Achieve a More Sustainable Viticulture.Pathogens (Basel, Switzerland) · 2022Review
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9 authors.
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No grant is acknowledged in the PubMed record.
Abstract
Genome editing via CRISPR/Cas9 is a powerful technology, which has been widely applied to improve traits in cereals, vegetables and even fruit trees. For the delivery of CRISPR/Cas9 components into dicotyledonous plants, Agrobacterium tumefaciens mediated gene transfer is still the prevalent method, although editing is often accompanied by the integration of the bacterial T-DNA into the host genome. We assessed two approaches in order to achieve T-DNA excision from the plant genome, minimizing the extent of foreign DNA left behind. The first is based on the Flp/FRT system and the second on Cas9 and synthetic cleavage target sites (CTS) close to T-DNA borders, which are recognized by the sgRNA. Several grapevine and apple lines, transformed with a panel of CRISPR/SpCas9 binary vectors, were regenerated and characterized for T-DNA copy number and for the rate of targeted editing. As detected by an optimized NGS-based sequencing method, trimming at T-DNA borders occurred in 100% of the lines, impairing in most cases the excision. Another observation was the leakage activity of Cas9 which produced pierced and therefore non-functional CTS. Deletions of genomic DNA and presence of filler DNA were also noticed at the junctions between T-DNA and genomic DNA. This study proved that many factors must be considered for designing efficient binary vectors capable of minimizing the presence of exogenous DNA in CRISPRed fruit trees.
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