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The introgression of rol-genes of rhizogenic Agrobacterium into the plant genome induces changes in plant phenotype and physiology. However, only limited experience with this technique is available for woody ornamentals. To induce new variation within the Escallonia genus, several species were co-cultivated with rhizogenic Agrobacterium strains. Co-cultivation of three rhizogenic Agrobacterium strains (Arqua1, LMG 63 and MAFF210266) with four Escallonia species (E. illinita, E. myrtoidea, E. rosea, and E. rubra), resulted in hairy roots production with a varying efficiency. Co-cultivation of E. rubra with MAFF210266, and E. myrtoidea with LMG63 did not yield hairy roots, while co-cultivation of E. rubra leaves with LMG63 was most successful for hairy root production (up to 80.6%). In addition, the efficiency of hairy root induction depended on the explant type (leaves or nodal sections). The presence of inserted rol-genes and aux-genes in hairy roots was molecularly confirmed using qPCR. Few shoots regenerated from hairy roots, but regeneration needs to be optimized for efficient implementation of rol-genes introgression in Escallonia breeding.