Abstract
In this study conservation of Castilleja levisecta Greenm., a globally endangered species was addressed through in vitro cryopreservation of shoot tips. In vitro cultures were successfully established using seedlings received from British Columbia, Canada. Shoot tips excised from in vitro propagated plants were cryopreserved using a droplet-vitrification method following optimization of individual protocol steps such as pre-culture, treatment with vitrification solutions, and unloading. The highest plant regrowth after cryopreservation (66%) was achieved when shoot tips were pre-cultured in 0.3 M sucrose for 17 h followed by 0.5 M sucrose for 4 h, incubated in an osmo-protectant solution (17.5% [v/v] glycerol and 17.5% [w/v] sucrose) for 20 min, exposed to vitrification solution A3 (37.5% [v/v] glycerol plus 15% [v/v] dimethylsulfoxide (DMSO) plus 15% [v/v] ethylene glycol (EG) plus 22.5% [w/v] sucrose) on ice for 40 min, and unloaded in 0.8 M sucrose solution for 30 min. Healthy plants were developed from cryopreserved shoot tips and propagated in vitro using nodal segments. Plants derived from in vitro culture and from cryopreserved tissues were successfully rooted and acclimated in a greenhouse with 100% survival rate. Acclimatized plants were reintroduced in a naturalized propagation area at the Conservation Nursery at Fort Rodd Hill, Canada. Twenty of 94 reintroduced plants (21%) survived the transit from lab to field and some had started to flower. This is the first report for cryopreservation of C. levisecta, an important step in conserving and re-introducing this critically imperiled species in nature.
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References
Ashmore SE, Hamilton KN, Offord CA (2011) Conservation technologies for safeguarding and restoring threatened flora: case studies from eastern Australia. In Vitro Cell Dev Biol —Plant 47(1):99–109. https://doi.org/10.1007/s11627-010-9320-9
Benson EE (2008) Cryopreservation theory. In: Reed BM (ed) Plant cryopreservation: a practical guide. Springer, Berlin, pp 15–32. https://doi.org/10.1007/978-0-387-72276-4_2
Caplow F (2004) Reintroduction plan for golden paintbrush (Castilleja levisecta). Washington Natural Heritage Program, Washington Department of Natural Resources Olympia, WA 98504–7014. Available from http://cascadiaprairieoak.org/documents/paintbrushreintroduction.pdf
Coelho N, González-Benito ME, Romano A (2014) Approaches for the cryopreservation of Plantago algarbiensis, a rare endemic species of the algarve. CryoLetters 35(6):521–529
Committee on the Status of Endangered Wildlife in Canada (COSEWIC) (2007) Assessment and update status report on the golden paintbrush Castilleja levisecta in Canada. Ottawa. Available from www.sararegistry.gc.ca/status/status_e.cfm
Coste A, Halmagyi A, Butiuc-Keul AL, Deliu C, Coldea G, Hurdu B (2012) In vitro propagation and cryopreservation of Romanian endemic and rare Hypericum species. Plant Cell Tissue Organ Cult 110(2):213–226. https://doi.org/10.1007/s11240-012-0144-7
Driver JA, Kuniyuki AH (1984) In vitro propagation of paradox walnut rootstock. HortSci 19:507–509
Dunwiddie PW, Martin RA, Jarisch MC (2013) Water and fertilizer effect on the germination and survival of direct-seeded golden paintbrush (Castilleja levisecta). Univ Wisconsin Press 31:10–12
Engelmann F (2004) Plant cryopreservation: progress and prospects. In Vitro Cell Dev Biol Plant 40(5):427–433. https://doi.org/10.1079/IVP2004541
Godt MJW, Caplow F, Hamrick JL (2005) Allozyme diversity in the federally threatened golden paintbrush, Castilleja levisecta (Scrophulariaceae). Conserv Gen 6(1):87–99. https://doi.org/10.1007/s10592-004-7746-5
González-Benito ME, Martín C (2011) In Vitro preservation of Spanish biodiversity. In Vitro Cell Dev Biol Plant 47(1):46–54. https://doi.org/10.1007/s11627-010-9333-4
González-Benito ME, Pérez C, Viviani AB (1997) Cryopreservation of nodal explants of an endangered plant species (Centaurium rigualii Esteve) using the encapsulation-dehydration method. Biodivers Conserv 6(4):583–590. https://doi.org/10.1023/A:1018337429589
Kim HH, Lee YG, Park SU, Lee SC, Baek HJ, Cho EG, Engelmann F (2009a) Development of alternative loading solutions in droplet-vitrification procedures. CryoLetters 30(4):291–299
Kim HH, Lee YG, Shin DJ, Ko HC, Gwag JG, Cho EG, Engelmann F (2009b) Development of alternative plant vitrification solutions in droplet-vitrification procedures. CryoLetters 30(5):320–334
Kim HH, Lee SC (2012) Personalisation of droplet-vitrification protocols for plant cells: a systematic approach to optimizing chemical and osmotic effects. CryoLetters 33(4):271–279
Kim HH, Popova EV, Shin DJ, Bae CH, Baek HJ, Park SU, Engelmann F (2012) Development of a droplet-vitrification protocol for cryopreservation of Rubia akane (Nakai) hairy roots using a systematic approach. CryoLetters 33(6):506–517
Lawrence BA, Kaye TN (2009) Reintroduction of Castilleja levisecta: effects of ecological similarity, source population genetics, and habitat quality. Restor Ecol 19:166–176
Lee YG, Popova E, Cui HY, Kim HH, Park SU, Bae CH, Lee SC, Engelmann F (2011) Improved cryopreservation of chrysanthemum (Chrysanthemum morifolium) using droplet-vitrification. CryoLetters 32(6):487–497
Lin L, Yuan B, Wang D, Li W (2014) Cryopreservation of adventitious shoot tips of Paraisometrum mileense by droplet vitrification. CryoLetters 35(1):22–28
Ma X, Bucalo K, Determann RO, Cruse-Sanders JM, Pullman GS (2012) Somatic embryogenesis, plant regeneration, and cryopreservation for Torreya taxifolia, a highly endangered coniferous species. In Vitro Cell Dev Biol Plant 48(3):324–334. https://doi.org/10.1007/s11627-012-9433-4
Mallón R, Barros P, Luzardo A, González ML (2007) Encapsulation of moss buds: an efficient method for the in vitro conservation and regeneration of the endangered moss Splachnum ampullaceum. Plant Cell Tissue Organ Cult 88(1):41–49. https://doi.org/10.1007/s11240-006-9176-1
Mallón R, Bunn E, Turner SR, González ML (2008) Cryopreservation of Centaurea ultreiae (Compositae) a critically endangered species from Galicia (Spain). CryoLetters 29(5):363–370
Mandal BB, Dixit-Sharma S (2007) Cryopreservation of in vitro shoot tips of Dioscorea deltoidea wall, an endangered medicinal plant: effect of cryogenic procedure and storage duration. CryoLetters 28:461–470
Menon A, Funnekotter B, Kaczmarczyk A, Bunn E, Turner S, Mancera RL (2014) Cold-induced changes affect survival after exposure to vitrification solution during cryopreservation in the south-west Australian Mediterranean climate species Lomandra sonderi (Asparagaceae). Plant Cell Tissue Organ Cult 119(2):347–358. https://doi.org/10.1007/s11240-014-0538-9
Muldrew K, Acker JP, Elliott JAW, McGann LE (2004) The water to ice transition: implications for living cells. In: Fuller BJ, Lane N, Benson EE (eds) Life in the frozen state. CRC Press, Baca Raton, pp 67–108
Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant 15(3):473–497. https://doi.org/10.1111/j.1399-3054.1962.tb08052.x
Natureserve. Available from http://explorer.natureserve.org/servlet/NatureServe?searchName=Castilleja+levisecta Accessed on 13.09.2016
Nishizawa S, Sakai A, Amano Y, Matsuzawa T (1993) Cryopreservation of asparagus (Asparagus officinalis L.) embryogenic suspension cells and subsequent plant regeneration by vitrification. Plant Sci 91(1):67–73. https://doi.org/10.1016/0168-9452(93)90189-7
Panis B, Strosse H, Van Den Hende S, Swennen R (2002) Sucrose preculture to simplify cryopreservation of banana meristem cultures. CryoLetters 23(6):375–384
Pearson S, Dunwiddie P (2006) Experimental seeding and outplanting of golden paintbrush (Castilleja levisecta) at Glacial Heritage, Mima Mounds and Rocky Prairie, Thurston County, WA. Final Report, February 2006. 19 pp
Pence VC (2010) The possibilities and challenges of in vitro methods for plant conservation. Kew Bull 65(4):539–547. https://doi.org/10.1007/s12225-011-9245-4
Peters P (2000) Tetrazolium Testing Handbook, Contribution No. 29. The Handbook on Seed Testing. Prepared by the Tetrazolium Subcommittee of the Association of Official Seed Analysts. Part 2. Lincoln, Nebraska
Popova E, Shukla M, Kim HH, Saxena PK (2015) Plant cryopreservation for biotechnology and breeding. In: Al-Khayri JM et al (eds) Advances in Plant Breeding Strategies: Breeding, Biotechnology and Molecular Tools. Springer International Publishing, Switzerland, pp 63–93
Rabba’a MM, Shibli RA, Shatnawi MA (2012) Cryopreservation of Teucrium polium L. shoot-tips by vitrification and encapsulation-dehydration. Plant Cell Tissue Organ Cult 110(3):371–382. https://doi.org/10.1007/s11240-012-0158-1
Rathwell R, Popova E, Shukla MR, Saxena PK (2016) Development of cryopreservation methods for cherry birch (Betula lenta L.), an endangered tree species in Canada. Can J For Res 46:1–9
Reed BM, Sarasan V, Kane M, Bunn E, Pence VC (2011) Biodiversity conservation and conservation biotechnology tools. In Vitro Cell Dev Biol Plant 47(1):1–4. https://doi.org/10.1007/s11627-010-9337-0
Ružić D, Vujović T, Cerović R (2013) Cryopreservation of cherry rootstock Gisela 5 (Prunus cerasus × Prunus canescens) shoot tips by droplet-vitrification technique. J Hort Res 21:79–85
Sakai A, Engelmann F (2007) Vitrification, encapsulation-vitrification and droplet vitrification: a review. CryoLetters 28(3):151–172
Turner S, Senaratna T, Touchell D, Bunn E, Dixon K, Tan B (2001a) Stereochemical arrangement of hydroxyl groups in sugar and polyalcohol molecules as an important factor in effective cryopreservation. Plant Sci 160(3):489–497. https://doi.org/10.1016/S0168-9452(00)00420-9
Turner SR, Senaratna T, Bunn E, Tan B, Dixon KW, Touchell DH (2001b) Cryopreservation of shoot tips from six endangered Australian species using a modified vitrification protocol. Ann Bot 87(3):371–378. https://doi.org/10.1006/anbo.2000.1346
U.S. Fish and Wildlife Service (2007) Golden Paintbrush (Castilleja levisecta) 5-year review summary and evaluation. Western Washington Fish and Wildlife Office, Lacey, Washington 98503. Available from http://ecos.fws.gov/docs/five_year_review/doc1764.pdf
Uchendu EE, Shukla MR, Reed BM, Saxena PK (2013) Melatonin enhances the recovery of cryopreserved shoot tips of American elm (Ulmus americana L.) J Pineal Res 55(4):435–442. https://doi.org/10.1111/jpi.12094
Volk GM, Walters C (2006) Plant vitrification solution 2 lowers water content and alters freezing behavior in shoot tips during cryoprotection. Cryobiology 52(1):48–61. https://doi.org/10.1016/j.cryobiol.2005.09.004
Wang B, Wang RR, Cui ZH, Bi WL, Li JW, Li BQ, Ozudogru EA, Volk GM, Wang QC (2014) Potential applications of cryogenic technologies to plant genetic improvement and pathogen eradication. Biotechnol Adv 32(3):583–595. https://doi.org/10.1016/j.biotechadv.2014.03.003
Wentworth JB (1994) The demography and population dynamics of Castilleja levisecta, an endangered perennial [thesis]. University of Washington, Seattle
Whiteley SE, Bunn E, Menon A, Mancera RL, Turner SR (2016) Ex situ conservation of the endangered species Androcalva perlaria (Malvaceae) by micropropagation and cryopreservation. Plant Cell Tissue Organ Cult 125(2):341–352. https://doi.org/10.1007/s11240-016-0955-z
Wilkinson T, Wetten A, Prychid C, Fay MF (2003) Suitability of cryopreservation for the long-term storage of rare and endangered plant species: a case history for Cosmos atrosanguineus. Ann Bot 91(1):65–74. https://doi.org/10.1093/aob/mcg009
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This research was supported by grants from the Gosling Foundation, Guelph, Canada, to the Gosling Research Institute for Plant Preservation (GRIPP).
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Salama, A., Popova, E., Jones, M.P. et al. Cryopreservation of the critically endangered golden paintbrush (Castilleja levisecta Greenm.): from nature to cryobank to nature. In Vitro Cell.Dev.Biol.-Plant 54, 69–78 (2018). https://doi.org/10.1007/s11627-018-9888-z
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DOI: https://doi.org/10.1007/s11627-018-9888-z