Effects of Type of Explant and Dark Preconditioning on Bud Formation in Habenaria radiata (Thunb.) in Vitro

Abstract

Habenaria radiata is a terrestrial orchid with beautiful bird-shaped petals. The wild H. radiata population has been severely affected by environmental disruption and overexploitation. In micropropagation of H. radiata, although aseptic germination has been studied, tissue culture methods have not yet been established. Shoot apexes and leaf explants from vegetative plants and flower stalks, stolons, and floret explants from reproductive plants were chosen for this study. Explants were cultured on half-strength inorganic salts and full-strength vitamins of Murashige and Skoog (1/2 MS) medium containing 30 g·L−1 sucrose, 8 g·L−1 agar (pH 5.6) supplemented with 4.44 μM N6-benzyladenine, and 0.54 μM α-naphthaleneacetic acid. After 8 weeks of culture, the highest survival rate was obtained with floret explants excised from plants at the reproductive phase. In floret culture, the number of adventitious bud formation per explant was 5.4 per upper floret and 4.0 per lower floret. Dark preconditioning, which inhibited browning and contamination, of explants before shoot apex culture increased survival rates of explants (53%) and bud formation (83%). Consequently, a tissue culture method using florets and shoot apexes as explant material was established for H. radiata.