Genome editing in almond using hairy root transformation system

Woody plant species are often challenging to transform, which complicates gene function studies using molecular biology techniques. To develop an effective gene editing tool for almond (Prunus dulcis, syn. Prunus amygdalus), we established a new method for inducing hairy roots in almond seedlings based on the injection of agrobacterial culture. We generated composite plants comprising wild-type shoots and transgenic hairy roots. This approach was also successful in almond x peach hybrids. The hairy root transformation system was used alongside Clustered Regularly Interspaced Short Palindromic Repeats/CRISPR-associated protein 9 gene editing to target two transcription factor genes (ETHYLENE RESPONSE FACTOR 74 and GIBBERELLIC ACID INSENSITIVE). We successfully generated hairy roots with knockouts of the target genes and evaluated the efficiency of guide RNAs for gene editing in almond. Our transgenic root method could be a valuable tool for routine gene function studies in almond.