Improved micropropagation protocol for leatherleaf fern (Rumohra adiantiformis) using rhizomes as donor explant

Abstract

A new micropropagation protocol for leatherleaf fern (Rumohra adiantiformis (G. Forst.) Ching) was successfully established using rhizomes as the donor explant, following appropriate sterilization. Murashige and Skoog (MS) medium containing 0.25mg/l 2,4-dichlorophenoxy acetic acid (2,4-D), 0.2mg/l α-naphthalene acetic acid (NAA), 1.0mg/l 6-benzyl adenine (BA), and 0.5mg/l thidiazuron (TDZ) with 30g/l sucrose (IM-4) was the most appropriate medium for culture initiation. When entire rhizomes harvested from mother stock plants were cultured on a simple paper bridge containing liquid IM-4 medium, culture initiation improved with 3.4 rhizomes regenerated per sub-culture. The average multiplication rate of newly regenerated rhizomes increased to 5.7/rhizome on half-strength MS medium supplemented with 0.05mg/l IAA, 0.25mg/l BA, 0.5mg/l Kin, 1g/l activated charcoal and 20g/l sucrose (MM-5). The level of multiplication peaked in the fifth subculture and retained high quality until the sixth subculture. From the seventh subculture onwards, the quality of regenerated fronds was reduced. The regenerated rhizomes rooted easily on MM-5 and could be acclimatized ex vitro with 97–100% survival.