Direct somatic embryogenesis using leaf explants and short term storage of synseeds in Spathoglottis plicata Blume

Abstract

Spathoglottis plicata Blume is a vulnerable orchid species in various parts of the world, and the conventional propagation provides limited success to its cultivation and conservation. Therefore, present study deals with the direct induction of somatic embryos (SEs) from the leaf explants of S. plicata. Murashige and Skoog’s (MS) medium fortified with various types and concentrations of plant growth regulators were used to induce somatic embryogenesis and plantlet production. The highest percentage of somatic embryo formation (93.7 ± 0.56%) was achieved on MS medium supplemented with 1.0 mg L−1 2,4-dichlorophenoxyacetic acid (2,4-D), whereas, maximum proliferation and increase in fresh weight (FW) of SEs (149.5 ± 0.24 mg/ 50 mg initial FW) was achieved on MS medium fortified with 2.0 mg L−1 6-benzylaminopurine (BAP) within 5 weeks of incubation. The light microscopic observation revealed that SEs emerged directly from the leaf surface. The viable synthetic seeds (SS) from SEs were prepared by encapsulating with gel matrix of 3% (w/v) sodium alginate and 100 mM calcium chloride. The SS were successfully stored for 60 days at − 4 °C with 97.4% germination frequency and shoot proliferation using MS medium with 2.0 mg L−1 BAP and 0.25 mg L−1 indole-3-acetic acid. The synergistic rooting frequency was observed on half-strength MS medium with 1.0 mg L−1 indole-3-butyric acid. The rooted shoots were acclimatized in a greenhouse with a 90% survival rate using soilrite® and vermicompost. This is an efficient short-term storage and regeneration protocol for S. plicata, which could help in reducing pressure on its wild population and could also be extended to the cryopreservation of this orchid species. The somatic embryos were induced directly from the leaf explants of Spathoglottis plicata and an efficient short-term storage protocol was developed by encapsulating these somatic embryos. The light microscopic analysis confirmed the formation of somatic embryos. The protocol could be convenient for the commercial cultivation and conservation of S. plicata.