María del Carmen Rocío Avila-Pérez, Laura White-Olascoaga, Amaury M. Arzate-Fernández
American Fern Journal 101 (1), 25-35, (1 January 2011) https://doi.org/10.1640/0002-8444-101.1.25
KEYWORDS: Leatherleaf fern, Rhumora, Sporophyte regeneration
Rumohra adiantiformis, also known as “leatherleaf fern”, is an ornamental species that, because of its long display life, is widely used in floral arrangements. In this study, a new protocol for in vitro regeneration of the leatherleaf fern was established. For spore germination, two culture media (MS and Knop) were assessed with presence or absence of 1g L−1 activated charcoal (AC) under different light and dark conditions. Frond, frond microcuttings, and prothallus explants were evaluated on Knop regeneration medium supplemented with 1g L−1 AC, 0.5% agar, pH 5.0 in combination with 2,4 dichlorophenoxyacetic acid (2,4-D: 0.0, 0.1, 0.5 and 1.0 mg L−1) and 6-benzylamino purine (BA: 0.0, 0.1, 0.5 and 1.0 mg L−1). For rooting, four levels of α-naphthaleneacetic acid (NAA: 0.0, 0.01, 0.1 and 0.2 mg L−1) were tested. After 18 days of culture, spore germination rate was 100% on Knop medium with AC and 8 h light/16 h dark. After 120 days of culture, sporophytes 1.7 ± 0.4 cm in length developed on Knop medium, while those spore-cultured on MS medium never produced sporophytes. From those germinated sporophytes, prothallus explants cultivated on Knop medium with AC and 0.5 mg L−1 BA showed the highest regeneration rate with 235.7 gametophytes. The best sporophyte rooting response was obtained with 0.01 mg L−1 NAA. Complete, regenerated sporophytes were obtained 183 days after culture initiation. By this procedure it would be possible to obtain up to 2 million sporophytes from one fertile frond. To determine the origin of the regenerated gametophytes, a histological analysis was performed with scanning electron microscopy (SEM). The analysis revealed that the gametophytes were regenerated from explant epidermal tissues on either the adaxial or abaxial surface.