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Cypripedium subtropicum embryo development and cytokinin requirements for asymbiotic germination

BACKGROUND: Cypripedium subtropicum is a unique, endangered lady’s slipper orchid with evergreen leaves on non-dormant shoots that is native to southwestern China. This study documents the major developmental events in C. subtropicum seed development from fertilization to seed maturity, determines t...

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Autores principales: Perner, Holger, Zhou, Rong, Perner, Wenqing, Jiang, Hong, Lee, Yung-I.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Springer Nature Singapore 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9525480/
https://www.ncbi.nlm.nih.gov/pubmed/36178517
http://dx.doi.org/10.1186/s40529-022-00359-4
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author Perner, Holger
Zhou, Rong
Perner, Wenqing
Jiang, Hong
Lee, Yung-I.
author_facet Perner, Holger
Zhou, Rong
Perner, Wenqing
Jiang, Hong
Lee, Yung-I.
author_sort Perner, Holger
collection PubMed
description BACKGROUND: Cypripedium subtropicum is a unique, endangered lady’s slipper orchid with evergreen leaves on non-dormant shoots that is native to southwestern China. This study documents the major developmental events in C. subtropicum seed development from fertilization to seed maturity, determines the optimum period for seed collection, and examines the cytokinin requirements for asymbiotic germination and protocorm survival. RESULTS: Structural studies revealed that embryo development proceeded after successful fertilization at 60 days after pollination (DAP). At 105 DAP, a globular embryo with the shrinking inner seed coat was observed, and seeds collected at this time point exhibited optimal germination. After 120 DAP, most seeds had a mature embryo within the capsule, and within the cells of the embryo proper, numerous proteins/lipid bodies were present as the main storage products. In addition, the inner seed coat had compressed into a thin layer that tightly enclosed the embryo, while the outer seed coat had progressively elongated, resulting in a hair-like appearance of the mature seed. Histochemical staining using Nile red and toluidine blue O (TBO) indicated that the lignified inner and outer seed coats may lead to coat-imposed dormancy. Seeds collected at this stage germinated poorly. Analyses of cytokinin preferences and optimal concentrations for germination and protocorm survival showed that both 6-(γ,γ-dimethylallylamino) purine (2iP) and 6-benzylaminopurine (BA) enhanced germination compared with the control, although higher concentrations of BA (4 and 8 μM) suppressed germination. The protocorm survival rate improved with increasing 2iP concentration. CONCLUSIONS: This study provides a reproducible procedure for culturing immature seeds of C. subtropicum based on a defined time schedule of seed development. In addition, the cytokinin 2iP was shown to improve germination and protocorm survival. This study provides a scientific basis for seedling establishment through asymbiotic seed culture for further reintroduction efforts.
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spelling pubmed-95254802022-10-02 Cypripedium subtropicum embryo development and cytokinin requirements for asymbiotic germination Perner, Holger Zhou, Rong Perner, Wenqing Jiang, Hong Lee, Yung-I. Bot Stud Original Article BACKGROUND: Cypripedium subtropicum is a unique, endangered lady’s slipper orchid with evergreen leaves on non-dormant shoots that is native to southwestern China. This study documents the major developmental events in C. subtropicum seed development from fertilization to seed maturity, determines the optimum period for seed collection, and examines the cytokinin requirements for asymbiotic germination and protocorm survival. RESULTS: Structural studies revealed that embryo development proceeded after successful fertilization at 60 days after pollination (DAP). At 105 DAP, a globular embryo with the shrinking inner seed coat was observed, and seeds collected at this time point exhibited optimal germination. After 120 DAP, most seeds had a mature embryo within the capsule, and within the cells of the embryo proper, numerous proteins/lipid bodies were present as the main storage products. In addition, the inner seed coat had compressed into a thin layer that tightly enclosed the embryo, while the outer seed coat had progressively elongated, resulting in a hair-like appearance of the mature seed. Histochemical staining using Nile red and toluidine blue O (TBO) indicated that the lignified inner and outer seed coats may lead to coat-imposed dormancy. Seeds collected at this stage germinated poorly. Analyses of cytokinin preferences and optimal concentrations for germination and protocorm survival showed that both 6-(γ,γ-dimethylallylamino) purine (2iP) and 6-benzylaminopurine (BA) enhanced germination compared with the control, although higher concentrations of BA (4 and 8 μM) suppressed germination. The protocorm survival rate improved with increasing 2iP concentration. CONCLUSIONS: This study provides a reproducible procedure for culturing immature seeds of C. subtropicum based on a defined time schedule of seed development. In addition, the cytokinin 2iP was shown to improve germination and protocorm survival. This study provides a scientific basis for seedling establishment through asymbiotic seed culture for further reintroduction efforts. Springer Nature Singapore 2022-09-30 /pmc/articles/PMC9525480/ /pubmed/36178517 http://dx.doi.org/10.1186/s40529-022-00359-4 Text en © The Author(s) 2022 https://creativecommons.org/licenses/by/4.0/Open AccessThis article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/ (https://creativecommons.org/licenses/by/4.0/) .
spellingShingle Original Article
Perner, Holger
Zhou, Rong
Perner, Wenqing
Jiang, Hong
Lee, Yung-I.
Cypripedium subtropicum embryo development and cytokinin requirements for asymbiotic germination
title Cypripedium subtropicum embryo development and cytokinin requirements for asymbiotic germination
title_full Cypripedium subtropicum embryo development and cytokinin requirements for asymbiotic germination
title_fullStr Cypripedium subtropicum embryo development and cytokinin requirements for asymbiotic germination
title_full_unstemmed Cypripedium subtropicum embryo development and cytokinin requirements for asymbiotic germination
title_short Cypripedium subtropicum embryo development and cytokinin requirements for asymbiotic germination
title_sort cypripedium subtropicum embryo development and cytokinin requirements for asymbiotic germination
topic Original Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9525480/
https://www.ncbi.nlm.nih.gov/pubmed/36178517
http://dx.doi.org/10.1186/s40529-022-00359-4
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