Regulation on the Induction of Protocorm-like Bodies and Callus in Dendrobium officinale

As globally important ornamental and medicinal plants, orchids exhibit significant differences in the difficulty and pathways of in vitro regeneration. Most orchid species can directly form protocorm-like bodies (PLBs) through the differentiation of shoot tips or other explants, which then regenerate into new plantlets, while some species form callus through explant dedifferentiation followed by PLB differentiation from the callus. At present, the regenerative mechanisms underlying PLB and callus in orchids, as well as the key factors influencing their differentiation, remain poorly elucidated. In this study, seedlings of Dendrobium officinale obtained from aseptic seed germination were used to investigate the effects of explant type, 2,4-D concentration, temperature, light intensity and photoperiod on the induction of PLBs and callus. The results showed that there were no significant differences in callus induction among the tested explants in D. officinale, whereas stem nodal segments were more suitable for PLB induction. For both internodal and nodal segments, the incidence rate of callus formation was higher than that of PLBs. The concentration of 2,4-D influenced the induction direction of the explants; higher concentration promoted PLB induction, while lower concentration was sufficient for callus formation. Low temperature and low light intensity inhibited PLB induction while promoting callus formation in D. officinale. High temperature and intense light partially caused desiccation of explants. A temperature of 25/22 °C (day/night) and a photosynthetic photon flux density of 50 µmol m−2 s−1 were more suitable for callus or PLB induction in D. officinale. A shorter photoperiod favored callus induction, while a longer photoperiod was beneficial for PLB induction. This study reveals the differences in influencing factors for PLB and callus induction in D. officinale, providing important insights for the propagation of orchid seedlings and laying a significant foundation for elucidating the mechanisms of PLB and callus induction.