High-efficiency cryopreservation of Araucaria angustifolia (Bertol.) Kuntze embryogenic cultures: ultrastructural characterization and morpho-physiological features

Fraga, Hugo Pacheco de Freitas; Vieira, Leila N.; Puttkammer, Catarina Corrêa; Silva, Jamily M. da; Anjos, Karina Goulart dos; Oliveira, Eliana M.; Guerra, Miguel Pedro

doi:10.1007/s11240-015-0895-z

Cited by 14 publications

(14 citation statements)

References 42 publications

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“…Cryopreservation has frequently been applied for the long-term maintenance of conifer embryogenic tissues [57][58][59], including Abies species such as A. nordmanniana [60], A. cephalonica [22,61], A. alba [29] and A. fraseri [21], or Abies hybrids [32,33].…”

Section: Discussionmentioning

confidence: 99%

“…In A. alba, Krajňáková et al [29] reported that four out of 12 cell lines recovered after cryostorage for six years. In our cryopreserved cell lines of A. alba, no significant differences in recovery frequencies among cell lines were recorded, although the cryotolerance in some conifer species was genotype dependent [56,58]. In control cell lines that were pretreated but not cryostored, the recovery frequencies reached 100%, indicating the cryoprotectant solutions had no harmful effect on recovery.…”

Section: Discussionmentioning

confidence: 99%

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Physiological and Structural Aspects of In Vitro Somatic Embryogenesis in Abies alba Mill

Salaj

Klubicová

Panis

et al. 2020

Forests

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Initiation of somatic embryogenesis from immature zygotic embryos, long-term maintenance of embryogenic tissue in vitro or by cryopreservation, as well as maturation, of somatic embryos of Abies alba Mill. are reported in this study. For the initiation of embryogenic tissues, a DCR medium containing different types of cytokinins (1 mg.L−1) were tested. During three consecutive years, 61 cell lines were initiated out of 1308 explants, with initiation frequencies ranging between 0.83 and 13.33%. The type of cytokinin had no profound effect on the initiation frequency within one given year. Microscopic observations revealed presence of bipolar somatic embryos in all initiated embryogenic tissues. Besides the typical bipolar somatic embryos, huge polyembryonal complexes, as well as “twin” embryos, were observed. Maturation of somatic embryos occurred on a DCR medium supplemented by abscisic acid (10 mg.L−1), polyethylene glycol (PEG-4000, 7.5%) and 3% maltose. The maturation capacity was cell-line dependent. All of the four tested cell lines produced cotyledonary somatic embryos, though at different quantities, of 16 to 252 per g of fresh weight. After germination, seedlings developed, but their further growth soon stopped after the formation of a resting bud. Altogether, seven cell lines were cryopreserved, using the slow-freezing technique. After rewarming, all tested cell lines showed regrowth rates between 81.8 and 100%.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Physiological and Structural Aspects of In Vitro Somatic Embryogenesis in Abies alba Mill

Salaj

Klubicová

Panis

et al. 2020

Forests

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“…In recent two decades optimized cryopreservation protocols were elaborated for many plant species, including conifers. Cryopreservation used for conifer embryogenic tissue is based on the slow-freezing method (Häggman et al, 1998; Fraga et al, 2016b; Nunes et al, 2017). Several pine species were cryopreserved for different periods of time with relatively high recovery frequencies reaching values 87% for P. pinea (Carneros et al, 2017), 100% for P. pinaster (Alvarez et al, 2012) and 100% for P. radiata (Lineros et al, 2018).…”

Section: Somatic Embryogenesis In Pinus Nigra Arnmentioning

confidence: 99%

Somatic Embryogenesis in Selected Conifer Trees Pinus nigra Arn. and Abies Hybrids

et al. 2019

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Somatic embryogenesis was achieved in the conifers Pinus nigra Arn. and in the hybrids Abies alba ×A. cephalonica and Abies alba ×A. numidica. For initiation of embryogenic tissue in P. nigra, immature zygotic embryos enclosed in megagametophytes were used. The initiated embryogenic cultures were maintained and proliferated on solid culture medium DCR supplemented with 9 μM 2,4-D and 2.2 μM BA. Microscopic investigations revealed the presence of bipolar early somatic embryos in proliferating tissue. Suspension cultures have also been established by resuspending the embryogenic tissue in liquid culture medium. Experimentation with abscisic acid concentration resulted in successful somatic embryo maturation. Besides abscisic acid, the carbohydrate content or higher concentration of gelling agent in maturation medium were also important requirements for somatic embryo maturation. Germination of cotyledonary somatic embryos occurred on hormone-free medium and terminated in somatic seedlings regeneration. The regenerated somatic seedlings were transferred to soil and were capable of successful development. For initiation of embryogenic tissue in Abies hybrids juvenile explants as immature or mature zygotic embryos as well as cotyledons were used and 4.4 μM BA as sole plant growth regulator was sufficient. Medium of the same composition was also suitable for their long-term maintenance. Maturation of somatic embryos was achieved on solid DCR medium supplemented with 38 μM abscisic acid, polyethylene glycol (0, 5, 7.5, and 10% PEG-4000) and different carbohydrates such as maltose, sucrose and glucose (each 3%). PEG-4000 stimulated somatic embryo development depending on the carbohydrate source used. Cotyledonary somatic embryos germinated after desiccation treatment and the regenerated somatic seedlings were transferred to soil. Cryopreservation of embryogenic tissue could be an alternative method for long-term maintenance. For cryopreservation the slow-freezing method was used with success. Tissue regeneration in the post thaw period was relatively high and the regenerated tissue produced mature somatic embryos and subsequent plantlets. The embryogenic tissue was also used in experiments focused on genetic transformation either by biolistic (P. nigra) or Agrobacterium-mediated (Abies hybrids) methods. A proteomic study was performed to gain a deeper insight into the early stages of P. nigra somatic embryogenesis.

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“…For conifer embryogenic tissues successful cryopreservation has been reported in Abies nordmanniana (Norgaard et al, 1993a), A. cephalonica (Aronen et al, 1999;Misson et al, 2006;Krajňáková et al, 2011), A. alba (Krajnaková et al, 2013, A. fraseri (Pullman et al, 2016), hybrids Abies alba x A. cephalonica and Abies alba x A. numidica (Salaj et al, 2010;Salaj et al 2016), Araucaria angustifolia (Fraga et al, 2016), Picea abies (Gupta et al, 1987;Norgaard et al, 1993b;Vondráková et al, 2010), P. glauca engelmanni (Cyr et al, 1994), P. mariana (Klimaszewska, 1995), P. sitchensis (Find et al, 1993;Gale et al, 2007), Pinus caribaea (Laine et al, 1992), P. nigra (Salaj et al, 2007), P. patula (Ford et al, 2000), P. radiata (Hargreaves et al, 2002), P. sylvestris (Häggman et al, 1998), P. roxburghii (Mathur et al, 2003), P. pinaster (Marum et al, 2004;Lelu-Walter et al, 2006;Alvarez et al, 2012), Pinus elliottii x P. caribaea (Nunez et al, 2017), Torreya taxifolia (Ma et al, 2012), Taxus x media, T. floridana (Skrlep et al, 2008), Tsuga canadensis, T. caroliniana (Merkle et al, 2014), Chamaecyparis thyoides (Ahn et al, 2017).…”

Section: Introduction Introduction Introduction Introductionmentioning

confidence: 99%

Cryopreservation of Abies alba embryogenic tissues by slow-freezing method

Salaj

Panis

Klubicová

et al. 2022

Not Bot Horti Agrobo

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Embryogenic tissues of Abies alba Mill. were cryopreserved using the slow-freezing approach. Four cell lines were incubated for 24 h on a medium with 0.5 M sorbitol and pre-treated with 5% DMSO. Subsequently, the tissues were frozen at a cooling rate of 1 °C min-1 to -40 °C and transferred to liquid nitrogen for 72 hours. After thawing in a water bath at 40 °C, the tissues were cultivated on a proliferation medium. All tested lines recovered, but variations in regrowth frequencies across cell lines were noticed (91.66 to 100%). The recovered tissues showed similar features to the control 2 (non-pre-treated and non-cryopreserved tissues). In the accumulation of fresh and dry mass, no statistically significant differences were observed between cryopreserved cultures and control 2. The cryopreserved tissues produced cotyledonary somatic embryos capable of germination. Microscopic observations revealed considerable structural changes as a consequence of the cryopreservation procedure. The long vacuolated suspensor cells were disrupted, and mostly the meristematic cells of the embryonal region survived. The typical bipolar structure of early somatic embryos has been regained during the post-thaw period. Differences in cryotolerance across cell lines were also observed.

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High-efficiency cryopreservation of Araucaria angustifolia (Bertol.) Kuntze embryogenic cultures: ultrastructural characterization and morpho-physiological features

Cited by 14 publications

References 42 publications

Physiological and Structural Aspects of In Vitro Somatic Embryogenesis in Abies alba Mill

Physiological and Structural Aspects of In Vitro Somatic Embryogenesis in Abies alba Mill

Somatic Embryogenesis in Selected Conifer Trees Pinus nigra Arn. and Abies Hybrids

Cryopreservation of Abies alba embryogenic tissues by slow-freezing method

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