An effective nutrient medium for asymbiotic seed germination and large-scale in vitro regeneration of Dendrobium hookerianum, a threatened orchid of northeast India

Background and aims Dendrobium hookerianum is a rare and threatened epiphytic orchid of northeast India. Prospects for conservation would be strengthened by developing an in vitro method for mass propagation. Seeds are minute and difficult to use directly in the field for this purpose, being non-endospermous with a low nutrient content and dependent on a specific fungus for germination and early seedling development. Although produced in large numbers (2–3 million per capsule), ,5 % germinate naturally in the wild. Our objective was to develop a rapid and successful method for in vitro propagation based on an initial in vitro asymbiotic seed germination step that achieved high percentages. Methodology Effects of four different media, i.e. (i) Murashige and Skoog (MS), (ii) Mitra et al., (iii) Knudson (KC) and (iv) Gamborg et al. (B5), were evaluated for large-scale multiplication by asymbiotic seed germination. Seedling leaf number, shoot number, shoot length, root number and root length were scored. After 7–8 months, large numbers of well-rooted plantlets were transferred to a glasshouse in thermocol pots containing compost. Six different composts based on broken brick and charcoal were compared for their ability to support further development over 90 days of hardening. Principal results The fastest and highest percentage seed germination was achieved using MS medium. Seeds on MS medium germinated in 3–4 weeks compared with 7–8 weeks on B5 medium. Seedling development was also superior on MS medium. The inclusion of plant growth regulators was unnecessary. Compost comprising broken brick and charcoal with an upper layer of moss was found to be the most suitable for the survival of transferred plantlets. Ninety per cent survival of plantlets was achieved 90 days after transfer to a glasshouse. Conclusions The use of MS culture medium is well suited for the mass multiplication of D. hookerianum plants intended for re-introducing this threatened orchid into the wild. * Corresponding author’s e-mail address: [email protected] Published by Oxford University Press. This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited. AoB PLANTS http://aobplants.oxfordjournals.org/ AoB PLANTS 2012: plr032; doi:10.1093/aobpla/plr032, available online at www.aobplants.oxfordjournals.org & The Authors 2012 1