A cryopreservation protocol for immature zygotic embryos of species of Ilex (aquifoliaceae)

Tropical Ilex species have recalcitrant seeds. This work describes experiments demonstrating the feasibility of long-term conservation of Ilex brasiliensis, I. brevicuspis, I. dumosa, I. intergerrima, I. paraguariensis, I. pseudoboxus, I. taubertiana, and I. theezans through cryopreservation of...

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Autores principales: Mroginski, Luis Amado, Sansberro, Pedro Alfonso, Scocchi, Adriana M., Luna, Claudia Verónica, Rey, Hebe Yolanda
Formato: Artículo
Lenguaje:Español
Publicado: Sociedad Latinoamericana de Microscopia Electrónica 2022
Materias:
Encapsulation-dehydration
Germplasm preservation
Liquid nitrogen
Llex
Acceso en línea:http://repositorio.unne.edu.ar/handle/123456789/48612
Aporte de:
RIUNNE - Repositorio Institucional de la Universidad Nacional del Nordeste (UNNE) de Universidad Nacional del Nordeste
id I48-R184-123456789-48612
record_format dspace
spelling I48-R184-123456789-486122024-09-27T12:28:16Z A cryopreservation protocol for immature zygotic embryos of species of Ilex (aquifoliaceae) Mroginski, Luis Amado Sansberro, Pedro Alfonso Scocchi, Adriana M. Luna, Claudia Verónica Rey, Hebe Yolanda Encapsulation-dehydration Germplasm preservation Liquid nitrogen Llex Tropical Ilex species have recalcitrant seeds. This work describes experiments demonstrating the feasibility of long-term conservation of Ilex brasiliensis, I. brevicuspis, I. dumosa, I. intergerrima, I. paraguariensis, I. pseudoboxus, I. taubertiana, and I. theezans through cryopreservation of zygotic rudimen- tary embryos at the heart developmental stage. The embryos were aseptically removed from the seeds and precultured (7 days) in the dark, at 27± 2oC on solidified (0.8% agar) 1/4MS medium, [consisting of quarter- strength salts and vitamins of Murashige and Skoog (1962) medium] with 3% sucrose and 0.1 mg/l Zeatin. The embryos were then encapsulated in 3% calcium alginate beads and pretreated at 24 h intervals in liquid medium supplemented with progressively increasing sucrose concentrations (0.5, 0.75 and 1 M). Beads were dehydrated for 5 h with silicagel to 25% water content (fresh weight basis) and then placed in sterile 5 ml cryovials. Then the beads were either plunged rapidly in liquid nitrogen were they were kept for 1 h (rapid cooling) or cooled at 1oC min-1 to -30oC. Then the beads were immersed in liquid nitrogen for 1 h (slow cooling). The beads were rewarmed by immersion of the cryovials for 1 min in a water bath thermostated at 30oC. Finally, beads were transferred onto culture medium (1/4MS, 3% sucrose, 0.1 mg/l zeatin, solidified with 0.8% agar) and incubated in a growth room at 27 ± 2oC under a 14 h light (116 μmol. m-2.s-1)/ 10 h dark photoperiod. Maximum recovery percentages between 15 and 83% (depending on de the species and the treatment) were obtained with the cryopreserved embryos. 2022-06-30T20:03:54Z 2022-06-30T20:03:54Z 2008-04 Artículo Mroginski, Luis Amado, et al., 2008. A cryopreservation protocol for immature zygotic embryos of species of Ilex (aquifoliaceae). Biocell. Mendoza: Sociedad Latinoamericana de Microscopia Electrónica, vol. 32, no. 1, p. 33-39. E-ISSN 1667-5746. 0327-9545 http://repositorio.unne.edu.ar/handle/123456789/48612 spa https://www.techscience.com/journal/biocell openAccess http://creativecommons.org/licenses/by-nc-nd/2.5/ar/ application/pdf application/pdf Sociedad Latinoamericana de Microscopia Electrónica Biocell, 2008, vol. 32, no. 1, p. 33-39.
institution Universidad Nacional del Nordeste
institution_str I-48
repository_str R-184
collection RIUNNE - Repositorio Institucional de la Universidad Nacional del Nordeste (UNNE)
language Español
topic Encapsulation-dehydration
Germplasm preservation
Liquid nitrogen
Llex
spellingShingle Encapsulation-dehydration
Germplasm preservation
Liquid nitrogen
Llex
Mroginski, Luis Amado
Sansberro, Pedro Alfonso
Scocchi, Adriana M.
Luna, Claudia Verónica
Rey, Hebe Yolanda
A cryopreservation protocol for immature zygotic embryos of species of Ilex (aquifoliaceae)
topic_facet Encapsulation-dehydration
Germplasm preservation
Liquid nitrogen
Llex
description Tropical Ilex species have recalcitrant seeds. This work describes experiments demonstrating the feasibility of long-term conservation of Ilex brasiliensis, I. brevicuspis, I. dumosa, I. intergerrima, I. paraguariensis, I. pseudoboxus, I. taubertiana, and I. theezans through cryopreservation of zygotic rudimen- tary embryos at the heart developmental stage. The embryos were aseptically removed from the seeds and precultured (7 days) in the dark, at 27± 2oC on solidified (0.8% agar) 1/4MS medium, [consisting of quarter- strength salts and vitamins of Murashige and Skoog (1962) medium] with 3% sucrose and 0.1 mg/l Zeatin. The embryos were then encapsulated in 3% calcium alginate beads and pretreated at 24 h intervals in liquid medium supplemented with progressively increasing sucrose concentrations (0.5, 0.75 and 1 M). Beads were dehydrated for 5 h with silicagel to 25% water content (fresh weight basis) and then placed in sterile 5 ml cryovials. Then the beads were either plunged rapidly in liquid nitrogen were they were kept for 1 h (rapid cooling) or cooled at 1oC min-1 to -30oC. Then the beads were immersed in liquid nitrogen for 1 h (slow cooling). The beads were rewarmed by immersion of the cryovials for 1 min in a water bath thermostated at 30oC. Finally, beads were transferred onto culture medium (1/4MS, 3% sucrose, 0.1 mg/l zeatin, solidified with 0.8% agar) and incubated in a growth room at 27 ± 2oC under a 14 h light (116 μmol. m-2.s-1)/ 10 h dark photoperiod. Maximum recovery percentages between 15 and 83% (depending on de the species and the treatment) were obtained with the cryopreserved embryos.
format Artículo
author Mroginski, Luis Amado
Sansberro, Pedro Alfonso
Scocchi, Adriana M.
Luna, Claudia Verónica
Rey, Hebe Yolanda
author_facet Mroginski, Luis Amado
Sansberro, Pedro Alfonso
Scocchi, Adriana M.
Luna, Claudia Verónica
Rey, Hebe Yolanda
author_sort Mroginski, Luis Amado
title A cryopreservation protocol for immature zygotic embryos of species of Ilex (aquifoliaceae)
title_short A cryopreservation protocol for immature zygotic embryos of species of Ilex (aquifoliaceae)
title_full A cryopreservation protocol for immature zygotic embryos of species of Ilex (aquifoliaceae)
title_fullStr A cryopreservation protocol for immature zygotic embryos of species of Ilex (aquifoliaceae)
title_full_unstemmed A cryopreservation protocol for immature zygotic embryos of species of Ilex (aquifoliaceae)
title_sort cryopreservation protocol for immature zygotic embryos of species of ilex (aquifoliaceae)
publisher Sociedad Latinoamericana de Microscopia Electrónica
publishDate 2022
url http://repositorio.unne.edu.ar/handle/123456789/48612
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