Asymbiotic seed germination and in vitro development of orchid Papilionanthe Miss Joaquim

Abstract

Orchids with their sheer variety of species are amazing, a major sharer in global floriculture trade. Papilionanthe Miss Joaquim, well known hybrid orchid, also recognized as official National flower of Singapore is a resilient, sun loving orchid, which blossoms throughout the year. Though a seed pod derived from a single flower contains millions of dusty seeds, due to lack of endosperm, seeds cannot germinate without the help of symbiont fungus. Commercial production of seedlings thus remains challenging. Therefore, in this study, we report a novel asymbiotic seed germination protocol standardized for ‘Papilionanthe Miss Joaquim’ (a hybrid of Papilionanthe hookeriana x Papilionanthe teres) which could be adapted for mass cultivation in a commercial setup. Seed of Papilionanthe Miss Joaquim were treated with 0.1% H2O2 and 0.1% KNO3 to increase the germination rates. Seed pods were also subjected to various regimes of surface sterilization methods to reduce contaminations. Employing Carbendazim (1% w v-1), Tricyclazole (1% w v-1), Sodium hypochlorite (0.5% v v-1) and Ethanol (80% v v-1) in surface sterilization process, resulted in highest percentage of aseptic cultures. Seeds were inoculated on to Murashige and Skoog (MS) medium supplemented with BAP (6-Benzylaminopurine), NAA (1-Naphthaleneacetic acid) and 2,4-Dichlorophenoxyacetic acid (2,4-D) combination of 3 mg L-1 BAP (6-Benzylaminopurine) + 0.5 mg L-1 NAA (1-Naphthaleneacetic acid) found to be most effective to induce germination (84.67 ± 3.2%). Subsequently, germinated seedlings were subjected to different levels of BAP and NAA levels to achieve the highest number of plantlets. thus, multiplied plantlets were later subcultured onto MS medium containing 0.5 mg L-1 BAP and 3 mg L-1 NAA to induce rooting. Consequently, developed plantlets were acclimatized on a substratum containing coconut husk and charcoal pieces.