In vitro asymbiotic seed germination and micropropagation of Dendrobium heyneanum Lindl. – an endemic orchid of Western Ghats, India

Abstract

Abstract Dendrobium heyneanum Lindl. or Heyne’s Dendrobium is an endemic epiphytic orchid of Western Ghats categorized as a threatened taxon. The present investigation was aimed for conservational strategy using in vitro regeneration methods. In this investigation, mature pods of the D. heyneanum were collected from the field, and were inoculated aseptically on various nutrient media. Half-strength macro-MS media was found to be an efficient asymbiotic seed germination with 86.70% in 12 days old culture. Different stages (I-VI) of morphogenic characters were observed during entire studies; however, 20.84% of seeds produced young seedlings with roots on half-strength macro-MS media. Micropropagation protocol of D. heyneanum was established by using the protocorms (Stage IV) from the asymbiotic germinated seeds. The metamorphosis was observed on half-strength macro-MS fortified with 0.1 mg/L to 2.0 mg/L of Thidiazuron (TDZ), Benzyl amino purine (BAP) and Kinetin (KN). Higher frequency of protocorms (90.20%) were found in 1.0 mg/L KN with 95.56% of proliferation, shoot number (6.56) and shoot length (1.13 cm). Different frequency of in vitro flower buds was observed in protocorm at 0.5 mg/L (30.2%) and 1.0 mg/L (22.37%); and callus induction at 2.0 mg/L (20.67%) of BAP. Synergistic effect of plantlets were assessed with different concentration of combination of KN (1.0 mg/L) and auxins - α- Naphthaleneacetic acid (NAA), Indole- 3- acetic acid (IAA), and Indole- 3- butyric acid (IBA) from 0.1 mg/L to 1.0 mg/L). Combination of KN + IAA (1.0 mg/L) induced elongation, 0.92 cm long pseudobulbs and 0.74 cm rooting. The plantlets were then subsequently acclimatized and hardened on pots containing cocopeat and brick pieces. The survival rate was 52.73%. The present study results finding introduced a protocol for in vitro propagation of Dendrobium heyneanum Lindl. and thus, can be implemented as an ex-situ conservation.