Vol.:(0123456789) 1 3 Plant Cell, Tissue and Organ Culture (PCTOC) https://doi.org/10.1007/s11240-020-01899-y ORIGINAL ARTICLE Short‑term cold storage of encapsulated somatic embryos and retrieval of plantlets in grey orchid (Vanda tessellata (Roxb.) Hook. ex G.Don) M. Manokari 1  · R. Latha 2  · S. Priyadharshini 2  · Phanikanth Jogam 3  · Mahipal S. Shekhawat 2 Received: 1 May 2020 / Accepted: 28 July 2020 © Springer Nature B.V. 2020 Abstract Vanda tessellata (Roxb.) Hook. ex G.Don (grey orchid) is a horticultural and medicinally potent, and conservation prior- itized orchid species. The present study investigates asymbiotic seed germination, callus induction, somatic embryogenesis, encapsulation of somatic embryos (SEs), and retrieval of plantlets of V. tessellata from the artifcial seeds after short-term cold storage at − 4 °C for 12 months. Seeds cultured on full strength Murashige and Skoog’s (MS) medium containing 2.0 mg L −1 6-benzylaminopurine (BAP) had the highest frequency of asymbiotic seed germination (94%) as compared with half- and one-fourth strength MS media supplemented with various concentrations and combinations of 6-furfurylami- nopurine (kinetin) and BAP. The protocorm like bodies (PLBs) gained dediferentiation and proliferated into embryogenic calli on MS medium containing 2.0 mg L −1 BAP and 0.5 mg L −1 indole-3 acetic acid (IAA). Somatic embryos (SEs) were diferentiated from the callus when cultured on MS medium in combination with 1.0 mg L −1 BAP and 0.5 mg L −1 IAA. The morpho-anatomical elucidations authenticated somatic embryogenesis and various developmental stages of SEs from embryogenic calli. Short-term cold storage of SEs was carried out by encapsulating with 2% sodium alginate and 100 mM calcium chloride and stored under dark at − 4 °C for 12 months to conserve this valuable germplasm. The stored artifcial seeds were germinated (91%) on MS medium incorporated with additives and 0.5 mg L −1 each of BAP, kinetin, and IAA. Complete retrieval of plantlets was achieved in 8 week with formation of shoots (4.2 cm average length) and roots (2.5 cm). The well developed plantlets were acclimatized in the greenhouse with 90% survival rate. Key message Vanda tessellata (grey orchid) is a horticultural and medicinally potent, and conservation prioritized orchid species. A repro- ducible protocol has been successfully established for asymbiotic seed germination, callus induction, somatic embryogenesis, encapsulation of somatic embryos, and retrieval of plantlets of V. tessellata from the artifcial seeds after short-term cold storage at − 4 °C for 12 months. Keywords Artifcial seeds · Asymbiotic seed germination · Cold storage · Grey orchid · Somatic embryos Introduction The members of the family Orchidaceae are among the most diverse of the fowering plants, with about 800 genera and 25,000 species distributed worldwide (Chugh et al. 2009). In India, this family is represented by 1289 species, fve subspecies, and 28 varieties under 186 genera which are distributed from the alpine to coastal regions and the islands, but their maximum diversity occur in the Eastern Himalayan and Peninsular regions (Misra 2007). Primarily, the orchids are valued for fowers due to their exotic beauty and long Communicated by Qiao-Chun Wang. * Mahipal S. Shekhawat smahipal3@gmail.com 1 Siddha Clinical Research Unit, Central Council for Research in Siddha, Tirunelvelli, Tamil Nadu, India 2 Biotechnology Unit, Kanchi Mamunivar Government Institute for Postgraduate Studies and Research, Lawspet, Puducherry, India 3 Department of Biotechnology, Kakatiya University, Warangal, Telangana 506009, India