14 research outputs found
The influence of various sucrose concentrations, pH and hormone inhibitors on the efficiency of somatic embryogenesis in two fern species: Cyathea delgadii and Asplenium cuneifolium
Somatic embryogenesis of a tree fern Cyathea delgadii Sternb.: achievements and prospects
Genetic transformation of gentian Gentiana tibetica (King) leaf explants with Agrobacterium tumefaciens strain C58C1
Comparison of the morphogenic potential of five Gentiana species in leaf mesophyll protoplast culture and ploidy stability of regenerated calli and plants
CRISPR–Cas9-mediated genome editing in apple and grapevine
The CRISPR–Cas9 genome-editing tool and the availability of whole-genome sequences from plant species have
revolutionized our ability to introduce targeted mutations into important crop plants, both to explore genetic changes and
to introduce new functionalities. Here, we describe protocols adapting the CRISPR–Cas9 system to apple and grapevine
plants, using both plasmid-mediated genome editing and the direct delivery of CRISPR–Cas9 ribonucleoproteins (RNPs) to
achieve efficient DNA-free targeted mutations in apple and grapevine protoplasts. We provide a stepwise protocol for the
design and transfer of CRISPR–Cas9 components to apple and grapevine protoplasts, followed by verification of highly
efficient targeted mutagenesis, and regeneration of plants following the plasmid-mediated delivery of components. Our
plasmid-mediated procedure and the direct delivery of CRISPR–Cas9 RNPs can both be utilized to modulate traits of
interest with high accuracy and efficiency in apple and grapevine, and could be extended to other crop species. The
complete protocol employing the direct delivery of CRISPR–Cas9 RNPs takes as little as 2–3 weeks, whereas the plasmid-mediated
procedure takes >3 months to regenerate plants and study the mutation