Abstract
Previous studies on the vitrification of peach palm (Bactris gasipaes Kunth) embryogenic cluster highlighted PVS3 as essential for survival, but toxic nonetheless. The objective of this work was to identify methods to improve embryogenic cluster survival through either modification to PVS3 or other changes to the established protocol. PVS3 reduced to 80% strength was less toxic to non-vitrified controls and showed no significant difference in vitrified embryogenic cluster at any incubation time (60, 120, 180, or 240 min) compared to full strength PVS3. The addition of 0.1 M KCl, MgCl2, MgSO4, or K2H(PO4) to 80% PVS3 increased vitrified embryogenic cluster regrowth. D2O had no significant effect on embryogenic cluster regrowth when used as a solvent for PVS3 compared to H2O. Larger embryogenic cluster showed greater regrowth than smaller ones. Regrowth increased with rising rewarming temperature up to 70 °C, the highest tested temperature. The duration of rewarming between 1, 2, or 3 min had no overall significant effect, suggesting that the most critical events in rewarming occur within the first minute. The most effective strips used for droplet vitrification were made from aluminum or silver, which were significantly more effective than polystyrene and expanded polystyrene. Droplet vitrification was initially more effective than conventional cryovial immersion vitrification at 60 min incubation in PVS3, but not significantly different at 120, 180 or 240 min incubations. Partial dehydration for 1 h in a laminar flow hood significantly increased regrowth, but longer dehydration times decreased regrowth when combines with longer PVS3 incubation times.
Key message
Inorganic ions in diluted PVS3 reduced damage to embryogenic clusters compared to PVS3. Partial dehydration was an effective pre-treatment. Regrowth improved with rising rewarming temperatures up to the maximum tested.
Similar content being viewed by others
References
Akyurt M, Zaki G, Habeebullah B (2002) Freezing phenomena in ice–water systems. Energy Convers Manag 43:1773–1789. https://doi.org/10.1016/S0196-8904(01)00129-7
Al Zoubi OM, Normah MN (2015) Ultrastructural response of embryonic axes of Fortunella polyandra to dehydration and cryopreservation. Cryo Lett 36:379–391
Ashby MF (1989) Overview No. 80: on the engineering properties of materials. Acta Metall 37:1273–1293. https://doi.org/10.1016/0001-6160(89)90158-2
Baek H-J, Kim H-H, Cho E-G, Chae Y-A, Engelmann F (2003) Importance of explant size and origin and of preconditioning treatments for cryopreservation of garlic shoot apices by vitrification. Cryo Lett 24:381–388
Berjak P, Sershen VB, Pammenter NW (2011) Cathodic amelioration of the adverse effects of oxidative stress accompanying procedures necessary for cryopreservation of embryonic axes of recalcitrant-seeded species. Seed Sci Res 21:187–203. https://doi.org/10.1017/S0960258511000110
Berjak P, Walker M, Mycock DJ, Wesley-Smith J, Watt P, Pammenter NW (2000) Cryopreservation of recalcitrant zygotic embryos. In: Cryopreservation tropical plant germplasm current research progress applications. Proceedings of an international workshop, Tsukuba, Japan, October 1998, p 140–155
Bernardin JD, Mudawar I (2004) A Leidenfrost point model for impinging droplets and sprays. J Heat Transf 126:272–278. https://doi.org/10.1115/1.1652045
Binder H, Zschörnig O (2002) The effect of metal cations on the phase behavior and hydration characteristics of phospholipid membranes. Chem Phys Lipids 115:39–61. https://doi.org/10.1016/S0009-3084(02)00005-1
Cañavate JP, Lubian LM (1995) Relationship between cooling rates, cryoprotectant concentrations and salinities in the cryopreservation of marine microalgae. Mar Biol 124:325–334. https://doi.org/10.1007/BF00347136
Dumet D, Engelmann F, Chabrillange N, Duval Y (1993) Cryopreservation of oil palm (Elaeis guineensis Jacq.) somatic embryos involving a desiccation step. Plant Cell Rep 12:352–355. https://doi.org/10.1007/BF00237434
Elliott GD, Wang S, Fuller BJ (2017) Cryoprotectants: a review of the actions and applications of cryoprotective solutes that modulate cell recovery from ultra-low temperatures. Cryobiology 76:74–91. https://doi.org/10.1016/j.cryobiol.2017.04.004
Fahy GM, Wowk B (2015) Principles of cryopreservation by vitrification. In: Wolkers WF, Oldenhof H (eds) Cryopreservation and freeze-drying protocols. Springer, New York, NY, pp 21–82
Fernandez-Prini R, Harvey AH, Palmer DA (2004) Aqueous systems at elevated temperatures and pressures: physical chemistry in water, steam and hydrothermal solutions. Elsevier, Amsterdam
Friedman R (2018) Membrane-ion interactions. J Membr Biol 251:453–460. https://doi.org/10.1007/s00232-017-0010-y
Gerbeau P, Amodeo G, Henzler T, Santoni V, Ripoche P, Maurel C (2002) The water permeability of Arabidopsis plasma membrane is regulated by divalent cations and pH. Plant J 30:71–81. https://doi.org/10.1046/j.1365-313X.2002.01268.x
Han B, Bischof JC (2004) Direct cell injury associated with eutectic crystallization during freezing. Cryobiology 48:8–21. https://doi.org/10.1016/j.cryobiol.2003.11.002
He X, Park EYH, Fowler A, Yarmush ML, Toner M (2008) Vitrification by ultra-fast cooling at a low concentration of cryoprotectants in a quartz microcapillary: a study using murine embryonic stem cells. Cryobiology 56:223–232. https://doi.org/10.1016/j.cryobiol.2008.03.005
Heringer AS, Steinmacher DA, Fraga HPF, Vieira LN, Ree JF, Guerra MP (2013a) Global DNA methylation profiles of somatic embryos of peach palm (Bactris gasipaes Kunth) are influenced by cryoprotectants and droplet-vitrification cryopreservation. Plant Cell Tissue Organ Cult PCTOC 114:365–372. https://doi.org/10.1007/s11240-013-0331-1
Heringer AS, Steinmacher DA, Schmidt ÉC, Bouzon ZL, Guerra MP (2013b) Survival and ultrastructural features of peach palm (Bactris gasipaes, Kunth) somatic embryos submitted to cryopreservation through vitrification. Protoplasma 250:1185–1193. https://doi.org/10.1007/s00709-013-0500-4
Hughes ZE, Malajczuk CJ, Mancera RL (2013) The effects of cryosolvents on DOPC−β-sitosterol bilayers determined from molecular dynamics simulations. J Phys Chem B 117:3362–3375. https://doi.org/10.1021/jp400975y
Jena S, Horn J, Suryanarayanan R, Friess W, Aksan A (2017) Effects of excipient interactions on the state of the freeze-concentrate and protein stability. Pharm Res 34:462–478. https://doi.org/10.1007/s11095-016-2078-y
Kagawa R, Hirano Y, Taiji M, Yasuoka K, Yasui M (2013) Dynamic interactions of cations, water and lipids and influence on membrane fluidity. J Membr Sci 435:130–136. https://doi.org/10.1016/j.memsci.2013.02.006
Kartha KK, Fowke LC, Leung NL, Caswell KL, Hakman I (1988) Induction of somatic embryos and plantlets from cryopreserved cell cultures of white spruce (Picea glauca). J Plant Physiol 132:529–539. https://doi.org/10.1016/S0176-1617(88)80249-9
Kim H-H, Cho E-G, Baek H-J, Kim C-Y, Joachim Keller ER, Engelmann F (2004) Cryopreservation of garlic shoot tips by vitrification: effects of dehydration, rewarming, unloading and regrowth conditions. Cryo Lett 25:59–70
Kim HH, Yoon JW, Park YE, Cho EG, Sohn JK, Kim TK, Engelmann F (2006) Cryopreservation of potato cultivated varieties and wild species: critical factors in droplet vitrification. Cryo Lett 27:223–234
Kim H-H, Popova EV, Yi J-Y, Cho G-T, Park S-U, Lee S-C, Engelmann F (2010) Cryopreservation of hairy roots of Rubia akane (Nakai) using a droplet-vitrification procedure. Cryo Letters 31:473–484
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. Cryo Lett 33:506–517
Kirichek O, Soper A, Dzyuba B, Callear S, Fuller B (2015) Strong isotope effects on melting dynamics and ice crystallisation processes in cryo vitrification solutions. PLoS ONE. https://doi.org/10.1371/journal.pone.0120611
Klasczyk B, Knecht V, Lipowsky R, Dimova R (2010) Interactions of alkali metal chlorides with phosphatidylcholine vesicles. Langmuir 26:18951–18958. https://doi.org/10.1021/la103631y
Lang I, Sassmann S, Schmidt B, Komis G (2014) Plasmolysis: loss of turgor and beyond. Plants 3:583–593. https://doi.org/10.3390/plants3040583
Leunufna S, Keller ERJ (2003) Investigating a new cryopreservation protocol for yams (Dioscorea spp.). Plant Cell Rep 21:1159–1166. https://doi.org/10.1007/s00299-003-0652-3
Martino A, Songsasen N, Leibo SP (1996) Development into blastocysts of bovine oocytes cryopreserved by ultra-rapid cooling. Biol Reprod 54:1059–1069. https://doi.org/10.1095/biolreprod54.5.1059
Martınez MT, Ballester A, Vieitez AM (2003) Cryopreservation of embryogenic cultures of Quercus robur using desiccation and vitrification procedures. Cryobiology 46:182–189. https://doi.org/10.1016/S0011-2240(03)00024-5
Mathew L, McLachlan A, Jibran R, Burritt DJ, Pathirana R (2018) Cold, antioxidant and osmotic pre-treatments maintain the structural integrity of meristematic cells and improve plant regeneration in cryopreserved kiwifruit shoot tips. Protoplasma 255:1065–1077. https://doi.org/10.1007/s00709-018-1215-3
Matsumoto M, Saito S, Ohmine I (2002) Molecular dynamics simulation of the ice nucleation and growth process leading to water freezing. Nature 416:409–413. https://doi.org/10.1038/416409a
Mazur P (1990) Equilibrium, quasi-equilibrium, and nonequilibrium freezing of mammalian embryos. Cell Biophys 17:53–92. https://doi.org/10.1007/BF02989804
Mikuła A, Tomiczak K, Rybczyński JJ (2011) Cryopreservation enhances embryogenic capacity of Gentiana cruciata (L.) suspension culture and maintains (epi)genetic uniformity of regenerants. Plant Cell Rep 30:565–574. https://doi.org/10.1007/s00299-010-0970-1
Morel G, Wetmore RH (1951) Fern callus tissue culture. Am J Bot 38:141–143
Muldrew K, McGann LE (1994) The osmotic rupture hypothesis of intracellular freezing injury. Biophys J 66:532–541. https://doi.org/10.1016/S0006-3495(94)80806-9
Murashige T, Skoog F (1962) A revised medium for rapid growth and bio assays with tobacco tissue cultures. Physiol Plant 15:473–497
Mycock D (1999) Addition of calcium and magnesium to a glycerol and sucrose cryoprotectant solution improves the quality of plant embryo recovery from cryostorage. Cryo Lett 20:77–82
Mycock DJ, Berjak P, Pammenter NW, Vertucci CW (1997) Cryopreservation of somatic embryoids of Phoenix dactylifera. In: Ellis RH, Black M, Murdoch AJ, Hong TD (eds) Basic and applied aspects of seed biology: proceedings of the fifth international workshop on seeds, reading, 1995. Springer Netherlands, Dordrecht, pp 75–82
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
Nuc K, Marszałek M, Pukacki PM (2016) Cryopreservation changes the DNA methylation of embryonic axes of Quercus robur and Fagus sylvatica seeds during in vitro culture. Trees 30:1831–1841. https://doi.org/10.1007/s00468-016-1416-3
Panarese V, Laghi L, Pisi A, Tylewicz U, Rosa MD, Rocculi P (2012) Effect of osmotic dehydration on Actinidia deliciosa kiwifruit: a combined NMR and ultrastructural study. Food Chem 132:1706–1712. https://doi.org/10.1016/j.foodchem.2011.06.038
Panis B, Piette B, Swennen R (2005) Droplet vitrification of apical meristems: a cryopreservation protocol applicable to all Musaceae. Plant Sci 168:45–55. https://doi.org/10.1016/j.plantsci.2004.07.022
R Core Team (2014). R: A language and environment for statistical computing. R Foundation for Statistical Computing, Vienna, Austria. http://www.R-project.org/
Redondo-Morata L, Oncins G, Sanz F (2012) Force spectroscopy reveals the effect of different ions in the nanomechanical behavior of phospholipid model membranes: the case of potassium cation. Biophys J 102:66–74. https://doi.org/10.1016/j.bpj.2011.10.051
Ree JF, Polesi LG, Back F, Bertolazi AA, Silveira V, Guerra MP (2020) Aging peach palm (Bactris gasipaes Kunth) cultures lose embryogenic potential and metabolic cellular function due to continuous culture in hypoxic environments. Plant Cell Tissue Organ Cult PCTOC 140:49–67. https://doi.org/10.1007/s11240-019-01710-7
Reinhoud PJ, Iren F van, Kijne JW (2000) Cryopreservation of undifferentiated plant cells. In: Cryopreservation tropical plant germplasm current research progress applications. Proceedings of an international workshop, Tsukuba, Japan, October 1998, p 91–102
Ren L, Zhang D, Jiang X-N, Gai Y, Wang W-M, Reed BM, Shen X-H (2013) Peroxidation due to cryoprotectant treatment is a vital factor for cell survival in Arabidopsis cryopreservation. Plant Sci 212:37–47. https://doi.org/10.1016/j.plantsci.2013.07.011
Sanchez C, Martinez MT, Vidal N, San-Jose MC, Valladare S, Vieitez AM (2008) Preservation of Quercus robur germplasm by cryostorage of embryogenic cultures derived from mature trees and RAPD analysis of genetic stability. Cryo Lett 29:493–504
Sansinena M, Santos MV, Zaritzky N, Chirife J (2012) Comparison of heat transfer in liquid and slush nitrogen by numerical simulation of cooling rates for French straws used for sperm cryopreservation. Theriogenology 77:1717–1721. https://doi.org/10.1016/j.theriogenology.2011.10.044
Sansinena M, Santos MV, Chirife J, Zaritzky N (2018) In-vitro development of vitrified–warmed bovine oocytes after activation may be predicted based on mathematical modelling of cooling and warming rates during vitrification, storage and sample removal. Reprod Biomed Online 36:500–507. https://doi.org/10.1016/j.rbmo.2018.01.003
Santos MV, Sansinena M, Zaritzky N, Chirife J (2012) Assessment of external heat transfer coefficient during oocyte vitrification in liquid and slush nitrogen using numerical simulations to determine cooling rates. Cryo Lett 33:31–40
Sen A, Balamurugan V, Rajak KK, Chakravarti S, Bhanuprakash V, Singh RK (2009) Role of heavy water in biological sciences with an emphasis on thermostabilization of vaccines. Expert Rev Vaccines 8:1587–1602. https://doi.org/10.1586/erv.09.105
Sen-Rong H, Ming-Hua Y (2013) A simple cryopreservation protocol for in vitro-grown shoot tips of Chinese genuine red bud taro (Colocasia esculenta L. Schott Var. Cormosus CV. Hongyayu) by encapsulation-dehydration. Sci Hortic 162:226–233. https://doi.org/10.1016/j.scienta.2013.08.008
Sershen Berjak P, Pammenter NW, Wesley-Smith J (2012) Rate of dehydration, state of subcellular organisation and nature of cryoprotection are critical factors contributing to the variable success of cryopreservation: studies on recalcitrant zygotic embryos of Haemanthus montanus. Protoplasma 249:171–186. https://doi.org/10.1007/s00709-011-0275-4
Shabala S, Shabala L (2011) Ion transport and osmotic adjustment in plants and bacteria. Biomol Concepts 2:407–419. https://doi.org/10.1515/BMC.2011.032
Shimonishi K, Ishikawa M, Suzuki S, Oosawa K (2000) Cryopreservation of melon somatic embryos by desiccation method. In: Cryopreservation tropical plant germplasm current research progress applications. Proceedings of an international workshop, Tsukuba, Japan, October 1998, p 167–171
Song YS, Moon S, Hulli L, Hasan SK, Kayaalp E, Demirci U (2009) Microfluidics for cryopreservation. Lab Chip 9:1874–1881. https://doi.org/10.1039/B823062E
Song YS, Adler D, Xu F, Kayaalp E, Nureddin A, Anchan RM, Maas RL, Demirci U (2010) Vitrification and levitation of a liquid droplet on liquid nitrogen. Proc Natl Acad Sci 107:4596–4600. https://doi.org/10.1073/pnas.0914059107
Steinmacher DA, Krohn NG, Dantas ACM, Stefenon VM, Clement CR, Guerra MP (2007) Somatic embryogenesis in peach palm using the thin cell layer technique: induction, morpho-histological aspects and AFLP analysis of somaclonal variation. Ann Bot 100:699–709
Stott SL, Karlsson JOM (2009) Visualization of intracellular ice formation using high-speed video cryomicroscopy. Cryobiology 58:84–95. https://doi.org/10.1016/j.cryobiol.2008.11.003
Takahashi D, Uemura M, Kawamura Y (2018) Freezing tolerance of plant cells: from the aspect of plasma membrane and microdomain. In: Iwaya-Inoue M, Sakurai M, Uemura M (eds) Survival strategies in extreme cold and desiccation: adaptation mechanisms and their applications. Springer, Singapore, pp 61–79
Teixeira AS, Faltus M, Zámečník J, González-Benito ME, Molina-García AD (2014a) Glass transition and heat capacity behaviors of plant vitrification solutions. Thermochim Acta 593:43–49. https://doi.org/10.1016/j.tca.2014.08.015
Teixeira AS, González-Benito ME, Molina-García AD (2014b) Measurement of cooling and warming rates in vitrification-based plant cryopreservation protocols. Biotechnol Prog 30:1177–1184. https://doi.org/10.1002/btpr.1938
Valentovic P, Luxova M, Kolarovic L, Gasparikova O (2006) Effect of osmotic stress on compatible solutes content, membrane stability and water relations in two maize cultivars. Plant Soil Environ 52:186–191
Valladares S, Toribio M, Celestino C, Vieitez AM (2004) Cryopreservation of embryogenic cultures from mature Quercus suber trees using vitrification. Cryo Lett 25:177–186
Volk GM, Harris JL, Rotindo KE (2006) Survival of mint shoot tips after exposure to cryoprotectant solution components. Cryobiology 52:305–308. https://doi.org/10.1016/j.cryobiol.2005.11.003
Wang T, Zhao G, Tang HY, Jiang ZD (2015) Determination of convective heat transfer coefficient at the outer surface of a cryovial being plunged into liquid nitrogen. Cryo Lett 36:285–288
Wen B, Cai C, Wang R, Tan Y, Lan O (2010) Critical moisture content windows differ for the cryopreservation of pomelo (Citrus grandis) seeds and embryonic axes. Cryo Lett 31:29–39
Wesley-Smith J, Walters C, Pammenter NW, Berjak P (2001) Interactions among water content, rapid (Nonequilibrium) cooling to −196 °C, and survival of embryonic axes of Aesculus hippocastanum L. Seeds Cryobiol 42:196–206. https://doi.org/10.1006/cryo.2001.2323
Wowk B (2010) Thermodynamic aspects of vitrification. Cryobiology 60:11–22. https://doi.org/10.1016/j.cryobiol.2009.05.007
Zhang D, Ren L, Chen G, Zhang J, Reed BM, Shen X (2015) ROS-induced oxidative stress and apoptosis-like event directly affect the cell viability of cryopreserved embryogenic callus in Agapanthus praecox. Plant Cell Rep 34:1499–1513. https://doi.org/10.1007/s00299-015-1802-0
Acknowledgements
The authors thank the National Council for Scientific and Technological Development (CNPq, Proc. 407974/2018-0, and 302798/2018-8) and CAPES (Coordenação de Aperfeicoamento de Pessoal de Nível Superior) of Brazil for funding. The authors would also like to thank Dr. Angelo Schaubb Heringer for sharing his expertise in cryopreservation techniques.
Author information
Authors and Affiliations
Contributions
Each author contributed equally.
Corresponding author
Ethics declarations
Conflict of interest
The authors have no conflict of interest to report.
Additional information
Communicated by Qiao-Chun Wang.
Publisher's Note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Rights and permissions
About this article
Cite this article
Ree, J.F., Guerra, M.P. Exogenous inorganic ions, partial dehydration, and high rewarming temperatures improve peach palm (Bactris gasipaes Kunth) embryogenic cluster post-vitrification regrowth. Plant Cell Tiss Organ Cult 144, 157–169 (2021). https://doi.org/10.1007/s11240-020-01852-z
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s11240-020-01852-z