Somatic embryogenesis in ferns: a new experimental system

Abstract

Key messageSomatic embryogenesis has never been reported in ferns. The study showed that it is much easier to evoke the acquisition and expression of embryogenic competence in ferns than in spermatophytes.AbstractWe discovered that the tree fern Cyathea delgadii offers an effective model for the reproducible and rapid formation of somatic embryos on hormone-free medium. Our study provides cyto-morphological evidence for the single cell origin and development of somatic embryos. Somatic embryogenesis (SE) in both primary and secondary explants was induced on half-strength micro- and macro-nutrients Murashige and Skoog medium without the application of exogenous plant growth regulators, in darkness. The early stage of SE was characterized by sequential perpendicular cell divisions of an individual epidermal cell of etiolated stipe explant. These resulted in the formation of a linear pro-embryo. Later their development resembled that of the zygotic embryo. We defined three morphogenetic stages of fern somatic embryo development: linear, early and late embryonic leaf stage. The transition from somatic embryo to juvenile sporophyte was quick and proceeded without interruption caused by dormancy. Following 9 weeks of culture the efficiency of somatic embryogenesis reached 12–13 embryos per responding explant. Spontaneous formation of somatic embryos and callus production, which improved the effectiveness of the process sevenfold in 10-month-long culture, occurred without subculturing. The tendency for C. delgadii to propagate by SE in vitro makes this species an excellent model for studies relating to asexual embryogenesis and the endogenous hormonal regulation of that process and opens new avenues of experimentation.

The content of endogenous hormones and sugars in the process of early somatic embryogenesis in the tree fern Cyathea delgadii Sternb.

“…As a result of the light deficit, plants show a dramatic elongation of the stem (Suzuki and Kerbauy 2006). The plant response is often linked to reduced leaf development, enhanced apical dominance, and a re-allocation of resources from storagefavouring shoot elongation to reproductive growth (Suzuki and Kerbauy 2006;Mikuła et al 2015b). Dark treatment of donor plantlets was reported to favour SE in Helianthus annuus L. (Fiore et al 1997), Dendranthema grandiflora Tzvelev.…”

“…Explants excised from sporophytes that developed under 16/8 photoperiod conditions are incapable of SE. In contrast, in etiolated explants, the first cell divisions typically starts at 10th day of culture, and during the next 4-6 days, many somatic embryos at linear developmental stage are observed (Mikuła et al 2015b). The present study was undertaken to find out if the etiolation, which is a key factor in the capacity of stipe explants to form somatic embryos, has an effect on the content and balance of hormones and sugars.…”

“…on PGR-free half-strength Murashige and Skoog's (1/2MS) medium, in darkness, we established a unique model system for the study of SE (Mikuła et al 2015b). In this system, somatic embryos regenerate directly from the single epidermal cells of stipe explants.…”

Effect of TIBA, fluridone and salicylic acid on somatic embryogenesis and endogenous hormone and sugar contents in the tree fern Cyathea delgadii Sternb.

“…is the first cryptogamic plant to be used as a model system for studying SE (Mikuła et al 2015b;Domżalska et al 2017;Grzyb et al 2017). It is also one of the few plant species in which direct SE can be induced on plant growth regulator-free medium (Mikuła et al 2015a).…”

Integration of tissue culture and cryopreservation methods for propagation and conservation of the fern Osmunda regalis L.

“…In vitro methods have been successfully exploited for many years for the micropropagation of plants, including ferns (Fernández and Revilla 2003;Bharati et al 2013b). Tissue culture provides optimal nutritional and environmental conditions for large-scale fern production, often in the absence of exogenous plant growth regulators (PGRs), even for inducing somatic embryogenesis (Mikuła et al 2015b). During the in vitro culture of ferns, many factors affecting the efficiency of multiplication of both gametophyte (Fernández et al 1997;Sheffield et al 1997;Goller and Rybczyński 2007;Wu et al 2010;Moura et al 2012) and sporophyte (Hirsch 1975;Materi and Cumming 1991;Teng 1997;Ambrósio and de Melo 2004;Kuriyama et al 2004b;Moura et al 2012;Mikuła et al 2015a) have been investigated, besides PGRs.…”