Propagation and cryopreservation of Ulleungdo hemlock (Tsuga ulleungensis) via somatic embryogenesis
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This study describes a method for the regeneration of Ulleungdo hemlock through somatic embryogenesis and a protocol for the cryopreservation of embryogenic cultures.
Ulleungdo hemlock (Tsuga ulleungensis) is a rare endemic forest species growing on Ulleung Island in Korea. Climate change and other factors such as insect pests and a low seed germination rate further endanger this species. Here, we established an in vitro propagation and cryopreservation system for Ulleungdo hemlock via somatic embryogenesis. Whole megagametophytes with zygotic embryos from immature Ulleungdo hemlock seeds were cultured on a modified Litvay medium supplemented with 9 µM 2,4-dichlorophenoxyacetic acid and 4.5 µM 6-benzylaminopurine. The cone collection date had a significant effect on the induction of embryogenic tissue (P < 0.001). Maturation medium containing Litvay salts, 50 g l−1 polyethylene glycol 4000, 30 g l−1 maltose, and abscisic acid (ABA) was used to test the effects of three different concentrations (30, 60, and 120 µM) of ABA on somatic embryo production. There were statistically significant effects of ABA concentration on somatic embryo production per 80 mg of embryogenic tissue (P < 0.001), and the most productive ABA concentration was 60 µM. We also conducted experiments to determine the effect of cryopreservation treatment on the maturation capacity of somatic embryos. Although the highest mean number of somatic embryo production per 80 mg of embryogenic tissue was obtained from a non-cryopreserved cell line (36.3 ± 3.5), no significant differences between the cryopreservation and non-cryopreservation treatment were observed (P = 0.093). Our results described here have great potential to contribute to the propagation and conservation of Ulleungdo hemlock in its native habitat.
KeywordsConifer Endangered species Embryogenic tissue initiation Cryopreservation Abscisic acid
This study was conducted with the support of ‘R&D Program for Forest Science Technology (Project no. FTIS 2018131B10-1820-BB01)’ provided by the Korea Forest Service (Korea Forestry Promotion Institute). This research was also supported by Grants from the Rural Development Administration, Republic of Korea [Next-Generation Bio-Green 21 Program (PJ01344401)]. We would like to thank Dr. Scott Merkle, Mr. Paul Montello, and Mr. Ryan Tull (University of Georgia, USA) for their technical assistance and advice.
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Conflict of interest
The authors declare that they have no conflicts of interest to disclose.
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