High-frequency protocorm-like bodies and shoot regeneration through a combination of thin cell layer and RITA® temporary immersion bioreactor in Cattleya forbesii Lindl.
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An efficient in vitro mass propagation through protocorm-like bodies (PLBs) was established in Cattleya forbesii Lindl., a commercially important orchid. Whole PLBs (W-PLB) and transverse thin cell layers of PLB (tTCL-PLB) explants were cultured in RITA® bioreactors based on temporary immersion system. Explants were transferred in semi-solid or RITA® bioreactor for protocorm production or shoot regeneration. The effect of different immersion frequencies, medium volumes and inoculum densities were studied and optimized. RITA® bioreactor cultures were found to be superior compared with semi-solid cultures regarding PLB production and shoot regeneration. tTCL-PLB explant types cultured in the RITA® bioreactor with immersion for 1 min/4 h, 250 mL of medium and 20 explants showed the highest number of PLBs per RITA® (2237 PLBs) and per explant (111.9 PLBs). The highest number of PLBs per explant was 21 times higher than those from semi-solid culture. The highest number of shoots per RITA® (3998 shoots) and per explant (199.9 shoots) were observed on tTCL-PLB cultured in RITA® bioreactor (1 min/4 h; 150 mL of medium and 20 explants). The highest number of shoots per explant was 95 times higher than those grown on semi-solid culture. Mass propagation of PLBs and shoots of C. forbesii Lindl. using combined thin cell layer and RITA® temporary immersion has been adapted in commercial practice.
KeywordsCattleya forbesii Lindl. Protocorm Thin cell layer In vitro Temporary immersion system Bioreactor Mass propagation
Authors are grateful to Republic of Turkey Ministry of Science, Industry, and Technology (Project No: 1217.STZ.2012-1), Ege University Scientific Research Projects Coordination Unit (Project No: 13-BIL-022) and Ön Danışmanlık Tourism Commitment and Trade Limited Company. In addition, we would like to thank Didem Ökmen, Biomedicine and Genome Center, Bioinformatics Department PhD student for her contributions to statistical analyzes and interpretations.
ME performed all experiments and wrote the manuscript. MB wrote the manuscript. ÖA calculated all statistical analyses and interpreted the data. AG supervised the research and edited and reviewed this manuscript. All authors designed research, read and approved the manuscript.
Compliance with ethical standards
Conflict of interest
The authors declare that they have no conflict of interest.
- Almeida V, Pacheco de Freitas Fraga H, Bachiega Navarro B, Guerra MP, Pescador R (2017) Dynamics in global DNA methylation and endogenous polyamine levels during protocorm-like bodies induction of Cattleya tigrina A. Richard. Acta Sci Biol Sci 39(4):497–505. https://doi.org/10.4025/actascibiolsci.v39i4.36656 CrossRefGoogle Scholar
- Bose B, Kumaria S, Choudhury H, Tandon P (2017) Insights into nuclear DNA content, hydrogen peroxide and antioxidative enzyme activities during transverse thin cell layer organogenesis and ex vitro acclimatization of Malaxiswallichii, a threatened medicinal orchid. Physiol Mol Biol Plants 23(4):955–968. https://doi.org/10.1007/s12298-017-0474-3 CrossRefGoogle Scholar
- De LC, Kumar R, Khan AM, Sangma R, Sailo N, Barman D (2014) Tropical and subtropical orchids. IJSAR 1(2):1–9Google Scholar
- Kamemoto H, Amore TD, Kuehnle AR (1999) Breeding Dendrobium orchids in Hawaii. University of Hawaii Press, HonoluluGoogle Scholar
- Knudson L (1946) A new nutrient solution for germination of orchid seed. Am Orchid Soc Bull 15:214–217Google Scholar
- Murthy HN, Paek KY, Park SY (2018) Micropropagation of orchids by using bioreactor technology. In: Lee YI, Yeung ET (eds) Orchid propagation: from laboratories to greenhouses—methods and protocols. Springer Protocols Handbooks, Humana Press, New York, pp 195–208. https://doi.org/10.1007/978-1-4939-7771-0_9 CrossRefGoogle Scholar
- Nayak NR, Sahoo S, Patnaik S, Rath SP (2002) Establishment of thin cross section (TCS) culture method for rapid micropropagation of Cymbidium aloifolium (L.) Sw. and Dendrobium nobile Lindl. (Orchidaceae). Scientia Horticult 94:107–116. https://doi.org/10.1016/S0304-4238(01)00372-7 CrossRefGoogle Scholar
- Pérez-Alonso N, Wilken D, Gerth A, Jähn A, Nitzsche HM, Kerns G, Capote-Perez A, Jiménez E (2009) Cardiotonic glycosides from biomass of Digitalis purpurea L. cultured in temporary immersion systems. Plant Cell Tissue Organ Cult 99(2):151–156. https://doi.org/10.1007/s11240-009-9587-x CrossRefGoogle Scholar
- Ross S, Castillo A (2009) Mass propagation of Vaccinium corymbosum in bioreactors. Agrociencia 13(2):1–8Google Scholar
- Roy AR, Patel RS, Patel VV, Sajeev S, Deka BC (2011) Asymbiotic seed germination, mass propagation and seedling development of Vanda coerulea Griff ex. Lindl. (Blue Vanda): an in vitro protocol for an endangered orchid. Sci Hortic 128:325–331. https://doi.org/10.1016/j.scienta.2011.01.023 CrossRefGoogle Scholar
- Teisson C, Alvard D (1995) A new concept of plant in vitro cultivation liquid medium: temporary immersion. In: Terzi M, Cella R, Falavigna A (eds) Current issues in plant molecular and cellular biology. Springer, Dordrecht, pp 105–110. https://doi.org/10.1007/978-94-011-0307-7_12 CrossRefGoogle Scholar
- Teixeira da Silva JA, Winarto B (2016) Somatic embryogenesis in two orchid genera (Cymbidium, Dendrobium). In: Germanà MA, Lambardi M (eds) in vitro embryogenesis in higher plants. Methods in molecular biology, vol 1359. Springer, New York, pp 371–386. https://doi.org/10.1007/978-1-4939-3061-6_18 CrossRefGoogle Scholar
- Teixeira da Silva JA, Singh N, Tanaka M (2006) Priming biotic factors for optimal protocorm-like body and callus induction in hybrid Cymbidium (Orchidaceae), and assessment of cytogenetic stability in regenerated plantlets. Plant Cell Tissue Organ Cult 84:135–144. https://doi.org/10.1007/s11240-005-9003-0 CrossRefGoogle Scholar