Dryopteris hikonensis is a terrestrial evergreen fern found in Korea, Japan, and eastern China. Although there are in vitro propagation techniques for several Dryopteris species, an optimized protocol for D. hikonensis has not yet been developed. In this study, we aimed to evaluate the effects of Murashige and Skoog (MS) medium on spore germination and gametophyte development to establish a species-specific propagation protocol. Mature spores of D. hikonensis were sown on full-strength (1×), half-strength (1/2×), and quarter-strength (1/4×) MS media and cultured under controlled light and temperature conditions. A Vittaria-type spore germination pattern was observed, which began five days after sowing. The highest final germination rate (94.66%) and fast early growth occurred under the 1/4× MS condition, suggesting favorable osmotic conditions for water absorption. Gametophytes developed via an Aspidium-type pathway to form cordate thalli and reproductive organs. Sporophytes were successfully formed in all treatment groups, most likely through apogamy. Full-strength MS medium maintained typical gametophyte morphology. Meanwhile, diluted media, particularly 1/4× MS medium, resulted in earlier and more rapid sporophyte development. These results highlight the importance of medium concentration in influencing gametophyte morphological characteristics and sporophyte emergence. 1/4× MS medium is recommended for efficient massive sporophyte induction, while 1× MS is more suitable for preserving normal gametophyte morphology. This study establishes an effective in vitro propagation protocol for D. hikonensis , supporting its conservation and potential horticultural applications.
Ku et al. (Wed,) studied this question.