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Cryopreservation of 13 Commercial Cannabis sativa Genotypes Using In Vitro Nodal Explants

Cannabis has developed into a multi-billion-dollar industry that relies on clonal propagation of elite genetics with desirable agronomic and chemical phenotypes. While the goal of clonal propagation is to produce genetically uniform plants, somatic mutations can accumulate during growth and compromi...

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Autores principales: Downey, Cassandra D., Golenia, Gregory, Boudko, Ekaterina A., Jones, Andrew Maxwell P.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8470898/
https://www.ncbi.nlm.nih.gov/pubmed/34579326
http://dx.doi.org/10.3390/plants10091794
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author Downey, Cassandra D.
Golenia, Gregory
Boudko, Ekaterina A.
Jones, Andrew Maxwell P.
author_facet Downey, Cassandra D.
Golenia, Gregory
Boudko, Ekaterina A.
Jones, Andrew Maxwell P.
author_sort Downey, Cassandra D.
collection PubMed
description Cannabis has developed into a multi-billion-dollar industry that relies on clonal propagation of elite genetics with desirable agronomic and chemical phenotypes. While the goal of clonal propagation is to produce genetically uniform plants, somatic mutations can accumulate during growth and compromise long-term genetic fidelity. Cryopreservation is a process in which tissues are stored at cryogenic temperatures, halting cell division and metabolic processes to facilitate high fidelity germplasm preservation. In this study, a series of experiments were conducted to optimize various stages of cryopreservation and develop a protocol for long-term germplasm storage of Cannabis sativa. The resulting protocol uses a standard vitrification procedure to cryopreserve nodal explants from in vitro shoots as follows: nodes were cultured for 17 h in a pre-culture solution (PCS), followed by a 20-min treatment in a loading solution (LS), and a 60 min incubation in plant vitrification solution 2 (PVS2). The nodes were then flash frozen in liquid nitrogen, re-warmed in an unloading solution at 40 °C, and cultured on basal MS culture medium in the dark for 5 days followed by transfer to standard culture conditions. This protocol was tested across 13 genotypes to assess the genotypic variability. The protocol was successful across all 13 genotypes, but significant variation was observed in tissue survival (43.3–80%) and regrowth of shoots (26.7–66.7%). Plants grown from cryopreserved samples were morphologically and chemically similar to control plants for most major traits, but some differences were observed in the minor cannabinoid and terpene profiles. While further improvements are likely possible, this study provides a functional cryopreservation system that works across multiple commercial genotypes for long-term germplasm preservation.
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spelling pubmed-84708982021-09-27 Cryopreservation of 13 Commercial Cannabis sativa Genotypes Using In Vitro Nodal Explants Downey, Cassandra D. Golenia, Gregory Boudko, Ekaterina A. Jones, Andrew Maxwell P. Plants (Basel) Article Cannabis has developed into a multi-billion-dollar industry that relies on clonal propagation of elite genetics with desirable agronomic and chemical phenotypes. While the goal of clonal propagation is to produce genetically uniform plants, somatic mutations can accumulate during growth and compromise long-term genetic fidelity. Cryopreservation is a process in which tissues are stored at cryogenic temperatures, halting cell division and metabolic processes to facilitate high fidelity germplasm preservation. In this study, a series of experiments were conducted to optimize various stages of cryopreservation and develop a protocol for long-term germplasm storage of Cannabis sativa. The resulting protocol uses a standard vitrification procedure to cryopreserve nodal explants from in vitro shoots as follows: nodes were cultured for 17 h in a pre-culture solution (PCS), followed by a 20-min treatment in a loading solution (LS), and a 60 min incubation in plant vitrification solution 2 (PVS2). The nodes were then flash frozen in liquid nitrogen, re-warmed in an unloading solution at 40 °C, and cultured on basal MS culture medium in the dark for 5 days followed by transfer to standard culture conditions. This protocol was tested across 13 genotypes to assess the genotypic variability. The protocol was successful across all 13 genotypes, but significant variation was observed in tissue survival (43.3–80%) and regrowth of shoots (26.7–66.7%). Plants grown from cryopreserved samples were morphologically and chemically similar to control plants for most major traits, but some differences were observed in the minor cannabinoid and terpene profiles. While further improvements are likely possible, this study provides a functional cryopreservation system that works across multiple commercial genotypes for long-term germplasm preservation. MDPI 2021-08-28 /pmc/articles/PMC8470898/ /pubmed/34579326 http://dx.doi.org/10.3390/plants10091794 Text en © 2021 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Downey, Cassandra D.
Golenia, Gregory
Boudko, Ekaterina A.
Jones, Andrew Maxwell P.
Cryopreservation of 13 Commercial Cannabis sativa Genotypes Using In Vitro Nodal Explants
title Cryopreservation of 13 Commercial Cannabis sativa Genotypes Using In Vitro Nodal Explants
title_full Cryopreservation of 13 Commercial Cannabis sativa Genotypes Using In Vitro Nodal Explants
title_fullStr Cryopreservation of 13 Commercial Cannabis sativa Genotypes Using In Vitro Nodal Explants
title_full_unstemmed Cryopreservation of 13 Commercial Cannabis sativa Genotypes Using In Vitro Nodal Explants
title_short Cryopreservation of 13 Commercial Cannabis sativa Genotypes Using In Vitro Nodal Explants
title_sort cryopreservation of 13 commercial cannabis sativa genotypes using in vitro nodal explants
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8470898/
https://www.ncbi.nlm.nih.gov/pubmed/34579326
http://dx.doi.org/10.3390/plants10091794
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