Highly competent in vitro propagation of Thrixspermum japonicum (Miq.) Rchb.f., a rare epiphytic orchid

Abstract

An efficient protocol for asymbiotic seed germination and multiple shoot production of Thrixspermum japonicum has been established. Seeds were cultured on MSB (Murashige and Skoog, (MS) macro- and micro-nutrients, 0.1% (w/v) activated charcoal (AC), and 3.0% (w/v) sucrose) medium fortified with different concentrations of coconut water (CW), indole-3-acetic acid (IAA), or α-naphthaleneacetic acid (NAA), and gibberellic acid (GA3) to promote seed germination. The highest seed germination percentage (88.1%) was observed on MSB medium fortified with 0.2% (v/v) CW, 0.2 mg L−1 NAA and 0.5 mg L−1 GA3. Optimized shoot induction (94.3%) and maximum shoot numbers (8.3) were obtained on protocorm cultured on MS medium amended with 0.1% (w/v) AC, 3.0% (w/v) banana pulp, 2.0% (w/v) potato homogenate, 0.3 mg L−1 KN, 0.2 mg L−1 NAA, and 0.5 mg L−1 GA3. Rooting of the shoots was best achieved on MS medium supplemented with 0.1% (w/v) AC, 3.0% (w/v) banana pulp and 2.0% (w/v) potato homogenate, and 0.8 mg L−1 indole-3-butyric acid. Plantlets were acclimatized in the greenhouse with 88.1% survival rates. The protocol proposed in this study could be useful for conservation of S. japonicus, an important and rare epiphytic orchid.