Efficient Regeneration and Genetic Transformation System for Cymbidium kanran ‘Zhushalan’

Background: Cymbidium kanran ‘Zhushalan’ is a famous traditional Chinese orchid with high ornamental and economic value. As its market expands, there is a need to improve its key horticultural traits and stress resistance. Unfortunately, these traits are difficult to breed using traditional methods, and an optimal regeneration and genetic transformation system for C. kanran has yet to be established. Methods: This study evaluated the factors affecting Agrobacterium-mediated genetic transformation and regeneration of C. kanran ‘Zhushalan’ using rhizomes obtained from seedlings as receptor material. Results: The highest regeneration frequency was achieved after pre-cultivating the rhizomes in the dark on ½ MS medium for 10 days. The genetic transformation system was optimized as follows: Agrobacterium strain, EHA105; optimal concentration of Agrobacterium solution, OD600 = 0.6; 100 mg·L−1 acetosyringone; an infection time of no more than 40 min; and co-culturing for one to three days. Positive strains were screened using meropenem (15 mg·L−1) and hygromycin (50 mg·L−1) and confirmed through PCR and qRT-PCR. A transformation rate of 11.67% was achieved. Conclusions: An efficient regeneration and genetic transformation system for C. kanran ‘Zhushalan’ has been established for developing transgenic technologies. Our findings will stimulate research on functional genes and molecular breeding related to C. kanran.