In vitro propagation and cryopreservation of Thuja koraiensis Nakai via somatic embryogenesis
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Korean arbor vitae (KAV; Thuja koraiensis Nakai) is a critically endangered coniferous tree in Korea. Here, we report the somatic embryogenesis (SE) and cryopreservation system that can be used for micropropagation of KAV and long-term storage of KAV cultures. To induce SE in KAV, the influence of the developmental stage of zygotic embryos and the effect of basal medium on embryogenesis induction were examined. The developmental stage of zygotic embryos had a significant effect on the embryogenesis induction (P < 0.0001). The highest frequency of embryogenesis induction occurred in megagametophytes with zygotic embryos at precotyledonary (P) and late embryogeny (L1) stage (36%). The highest frequency of embryogenesis induction was obtained on initiation medium containing IM basal salts with 2.2 μM 6-benzylaminopurine and 4.5 μM 2,4-dichlorophenoxyacetic acid (35%). The effect of abscisic acid (ABA) on production of somatic embryos was tested. The highest number of somatic embryos per 50 mg of embryogenic tissue was achieved on maturation medium with levels of 100 μM ABA (24.0 ± 2.4). The effect of cryopreservation treatment to embryogenic tissues on the maturation capacity of somatic embryos was also tested. No significant differences between noncryopreservation and cryopreservation treatment were observed (P = 0.1896), and the highest mean number of somatic embryo per 50 mg of embryogenic tissues was obtained in noncryopreserved cell line (28.17 ± 5.66). Finally, the genetic identities of the plantlets regenerated from non- and cryopreserved embryogenic cell lines were verified and there was no genetic variation in the regenerated plantlets from cryostored embryogenic cell lines. This study is the first report on SE and the successful cryopreservation of embryogenic culture of the genus Thuja.
KeywordsCupressaceae Endangered conifer Embryogenic tissue Cryostorage Genetic fidelity
This research was carried out with the support of ‘R&D Program for Forest Science Technology (Project No. FTIS 2018131B10-1820-BB01) provided by Korea Forest Service (Korea Forestry Promotion Institute). We would like to thank the National Park Research Institute of Korea and Korea National Arboretum. We also thank to Dr. Hyo-In Lim (Korea Forest Research Institute) for the assistance.
YEC and CHA designed all experiments and wrote all drafts of the manuscript. CHA conducted the experiments and data analysis. KH carried out cytological analysis. HSP collected KAV cones for culture initiation and provided some photos.
Compliance with ethical standards
Conflict of interest
The authors declare that they have no conflict of interest.
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