Cryopreservation of Orthodox (Desiccation Tolerant) Seeds

  • Hugh W. Pritchard
  • Jayanthi Nadarajan

Although there are several methods of ex situ plant conservation, seed banking is the most efficient for many species, particularly for ease of application and the amount of diversity conserved (Linington and Pritchard 2001). Indeed, seed storage is the main form of ex situ plant genetic resources (PGR) conservation globally, representing about 90% of all collections, the vast majority of which are crops, including cultivars (FAO 1996). More than half the world’s PGR accessions are held in medium-term or long-term storage conditions. For long-term storage, the international standards are drying at 10–25°C and 10–15% RH to 3–7% moisture content, followed by storage at about –18°C (FAO/IPGRI 1994). Whilst less than expected seed longevity at about –20°C is known for “intermediate” or Type II seeds (see Pritchard 2004), ‘orthodox’ Type I seeds can also age quicker at seed bank temperatures than predicted by the seed viability equations (for explanation see Pritchard and Dickie 2003). This was revealed by an elegant experiment in which orthodox Hordeum vulgare ssp distichium cv. Proctor seed, ageing at warm temperatures, was interrupted by transfer to –20°C, which indicated longevity parameters associated with storage at –6°C (Roberts and Ellis 1977). This observation, combined with comparisons between actual performance and extrapolation of longevity to sub-zero temperatures, suggests that the benefits of all sub-zero storage temperatures may be less than previously thought (Dickie et al. 1990; Pritchard 1995; Pritchard and Dickie 2003; Walters et al. 2004). Although the modelling of seed longevity at sub-zero temperatures is a challenge, such cooling generally enhances dry seed longevity (Dickie et al. 1990; Pritchard and Seaton 1993; Walters et al. 2004). Consequently, cryopreservation may be of particular importance for the long-term (10-100s years) storage of otherwise inherently short-lived orthodox seeds (Pritchard 1995; Pritchard and Seaton 1993; Pritchard et al. 1999b; Walters et al. 2004; Pritchard 2007).


Seed Storage Plant Genetic Resource Saturated Salt Solution Seed Longevity Orthodox Seed 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Brown HT, Escombe F (1897-1898) Note on the influence of very low temperatures on the germinative power of seeds. Proc Roy Soc 62: 160-165Google Scholar
  2. Busse WF (1930) Effect of low temperatures on germination of impermeable seeds. Bot Gaz 89: 169-179CrossRefGoogle Scholar
  3. Busse WF, Burnham CR (1934) Some effects of low temperatures on seeds. Bot Gaz 90: 399-411CrossRefGoogle Scholar
  4. Crane J, Kovach D, Gardner C, Walters C (2006) Triacylglycerol phase and ‘intermediate’ seed storage physiology: A study of Cuphea carthagenesis. Planta 223: 1081-1089CrossRefPubMedGoogle Scholar
  5. Crane J, Miller AL, van Roekel JW, Walters CW (2003) Triacylglycerols determine the unusual storage physiology of Cuphea seed. Planta 217: 699-708CrossRefPubMedGoogle Scholar
  6. Darwin Initiative (2005) Cryoconservation Centre of Excellence for Sub-Sahara Africa.
  7. Daws MI, Davies J, Vaes E, van Gelder R, Pritchard HW (2007) Two-hundredyear seed survival of Leucospermum and two other woody species from the Cape Floristic region, South Africa. Seed Sci Res (in press)Google Scholar
  8. De Candolle A (1865) De la germination sous les degrees divers de temperature constante. Arch Sci Phys Nat 24: 243-282Google Scholar
  9. De Candolle C, Pictet R (1879) Recherches concernant l’action des basses temperatures sur la faculte germinative des graines. Arch Sci Phys Nat 2, 354: 629Google Scholar
  10. Dewar J, McKendrick JG (1892). On liquid air. Proc Roy Inst 12: 699Google Scholar
  11. Dickie JB, Ellis RH, Kraak HL, Ryder K, Tompsett PB (1990) Temperature and seed storage longevity. Ann Bot 65: 197-204Google Scholar
  12. Edwards M, Colin M (1834) De l’influence de la temperature sur la germination. Ann Sci Nat II, 1: 257-270Google Scholar
  13. Ellis RH, Hong TD, Roberts EH (1990) An intermediate category of seed storage behaviour? I. Coffee. J Exp Bot 41: 1167-1174CrossRefGoogle Scholar
  14. Ellis RH, Hong TD, Roberts EH (1991) Effect of storage temperature and moisture on the germination of papaya seeds. Seed Sci Res 1: 69-72Google Scholar
  15. European Union (2006) Cost Action 871. Cryopreservation of crop species in Europe.
  16. FAO (1996) The State of the World’s Plant Genetic Resources for Food and Agriculture. FAO, Rome, ItalyGoogle Scholar
  17. FAO/IPGRI (1994) Genebank Standards. FAO, Rome, ItalyGoogle Scholar
  18. Gonzalez-Benito ME, Fernandez-Llorente F, Perez-Garcia F (1998) Interaction between cryopreservation, rewarming rate and seed humidification on the germination of two Spanish endemic species. Ann Bot 82: 683-686CrossRefGoogle Scholar
  19. Hamilton KN, Ashmore SE, Drew RA (2005) Investigations on desiccation and freezing tolerance of Citrus australasica seed for ex situ conservation. pp 157-161 In: Adkins SW, Ainsley PJ, Bellairs SM, Coates DJ, Bell LC (eds) Proceedings of the 5th Australian Workshop on Native Seed Biology, Brisbane, Queensland, June 2004, ACMERGoogle Scholar
  20. Hor YL, Kim YJ, Ugap A, Chabrillange N, Sinniah UR, Engelmann F, Dussert S (2005) Optimal hydration status for cryopreservation of intermediate oily seeds: Citrus as a case study. Ann Bot 95: 1153-1161CrossRefPubMedGoogle Scholar
  21. Lambardi M, De Carlo A, Biricolti S, Puglia AM, Lombardo G, Siragusa M, De Pasquale F (2004) Zygotic and nucellar embryo survival following dehydration/cryopreservation of Citrus intact seeds. CryoLetters 25: 81-90PubMedGoogle Scholar
  22. Leprince O, van Aelst AC, Pritchard HW, Murphy DJ (1998) Oleosins prevent oil-body coalescence during seed imbibition as suggested by a lowtemperature scanning electron microscopy study of desiccation-tolerant and sensitive oilseeds. Planta 204: 109-119CrossRefGoogle Scholar
  23. Linington SH, Pritchard HW (2001) Gene banks. pp 165-181 In: Levin SA (editor in chief) Encyclopedia of Biodiversity. Vol 3. Academic Press, New YorkGoogle Scholar
  24. Lipman CB (1936) Normal viability of seeds and bacterial spores after exposure to temperatures near the absolute zero. Plant Physiol 11: 201-205CrossRefPubMedGoogle Scholar
  25. Lipman CB, Lewis GN (1934) Tolerance of liquid air temperatures by seed of higher plants for sixty days. Plant Physiol 9: 392-394CrossRefPubMedGoogle Scholar
  26. Pritchard HW (1995) Cryopreservation of seeds. pp 133-144 In: Day JG, McLellan MR (eds) Methods in Molecular Biology. Vol 38. Cryopreservation and Freeze-drying Protocols. Humana Press Inc., Totowa, NJGoogle Scholar
  27. Pritchard HW (2002) Cryopreservation and global warming! CryoLetters 23: 281-282PubMedGoogle Scholar
  28. Pritchard HW (2004) Classification of seed storage ‘types’ for ex situ conservation in relation to temperature and moisture. pp 139-161 In: Guerrant EO, Havens K, Maunder M (eds) Ex situ Plant Conservation: Supporting Species Survival in the Wild. Island Press, Washington DC, USAGoogle Scholar
  29. Pritchard HW (2006) Cryo network. Kew Scientist 30: 3 (
  30. Pritchard HW (2007) Cryopreservation of desiccation tolerant seeds. pp 183-199 In: Day JG, Stacey GN (eds) Cryopreservation and freeze-drying protocols. Humana Press Inc., Totowa, NJGoogle Scholar
  31. Pritchard HW, Dickie JB (2003) Predicting seed longevity: Use and abuse of seed viability equations. pp 653 -722 In: Smith RD, Dickie JB, Linington SH, Pritchard HW, Probert RJ (eds) Seed Conservation: Turning Science into Practice. Royal Botanic Gardens, Kew, UKGoogle Scholar
  32. Pritchard HW, Manger KR, Prendergast FG (1988) Changes in Trifolium arvense seed quality following alternating temperature treatment using liquid nitrogen. Ann Bot 62: 1-11Google Scholar
  33. Pritchard HW, Poynter AC, Seaton PT (1999a) Interspecific variation in orchid seed longevity in relation to ultra-drying and cryopreservation. Lindleyana 14: 92-101Google Scholar
  34. Pritchard HW, Seaton PT (1993) Orchid seed storage. Historical perspective, current status and future prospects. Selbyana 14: 89-104Google Scholar
  35. Pritchard HW, Wood CB, Amritphale D, Magill W, Benson EE (1999b) Freezinginduced dormancy in dried Carica papaya seeds: A new cryobiological syndrome? Cryobiology 38: 308Google Scholar
  36. Reed BM, Kovalchuk I, Kushnarenko S, Meier-Dinkel A, Schoenweiss K, Pluta S, Straczynska K, Benson EE (2004) Evaluation of critical points in technology transfer of cryopreservation protocols to international plant cryopreservation laboratories. CryoLetters 25: 341-352PubMedGoogle Scholar
  37. Roberts EH, Ellis RH (1977) Prediction of seed longevity at sub-zero temperatures and genetic resources conservation. Nature 268: 431-433CrossRefGoogle Scholar
  38. Roberts EH, Ellis RH (1989) Water and seed survival. Ann Bot 63: 39-52Google Scholar
  39. Sakai A, Noshiro M (1975) Some factors contributing to the survival of crop seeds cooled to the temperature of liquid nitrogen. pp 317-326 In: Frankel OH, Hawkes JG (eds) Crop Genetic Resources for Today and Tomorrow. Cambridge University Press, Cambridge, UKGoogle Scholar
  40. Selby AD (1901) Germination of the seeds of some common cultivated plants after prolonged immersion in liquid air. Bull Torrey Bot Club 28: 675-679CrossRefGoogle Scholar
  41. Stanwood PC (1985) Cryopreservation of seed germplasm for genetic conservation. pp 199-226 In: Kartha KK (ed) Cryopreservation of Plant Cells and Organs. CRC Press, Boca Raton, FloridaGoogle Scholar
  42. Stanwood PC, Bass LN (1978) Ultracold preservation of seed germplasm. pp 361- 371 In: Li PH, Sakai A (eds) Plant Cold Hardiness and Freezing Stress. Academic Press, New YorkGoogle Scholar
  43. Stushnoff C, Juntilla O (1978) Resistance to low temperature injury in hydrated lettuce seed by supercooling. pp 241-247 In: Li PH, Sakai A (eds) Plant Cold Hardiness and Freezing Stress: Mechanisms and Crop Implications, Academic Press, New YorkGoogle Scholar
  44. Sun WQ (2002) Methods for the study of water relations under desiccation stress. pp 47-91 In: Black M, Pritchard HW (eds) Desiccation and Survival in Plants: Drying Without Dying. CABI Publishing, Wallingford, UKGoogle Scholar
  45. Thiselton-Dyer W (1899) On the influence of the temperature of liquid hydrogen on the germinative power of seeds. Proc Roy Soc 65: 361-368CrossRefGoogle Scholar
  46. Vertucci CW (1989a) Effects of cooling rate on seeds exposed to liquid nitrogen temperatures. Plant Physiol 90: 1478-1485CrossRefPubMedGoogle Scholar
  47. Vertucci CW (1989b) Relationship between thermal transitions and freezing injury in pea and soybean seeds. Plant Physiol 90: 1121-1128CrossRefPubMedGoogle Scholar
  48. Volk GM, Crane J, Caspersen AM, Hill LM, Gardner C, Walters C (2006) Massive cellular disruption occurs during early imbibition of Cuphea seeds containing crystallized triacylglycerols. Planta 224: 1415-1426CrossRefPubMedGoogle Scholar
  49. Walters C, Wheeler L, Stanwood PC (2004) Longevity of cryogenically stored seeds. Cryobiology 48: 229-244CrossRefPubMedGoogle Scholar
  50. Wartman E (1860) Note relative a l’influence de froids excessifs sur la graines. Arch Sci Phys Nat 8: 227Google Scholar
  51. Wood C, Berjak P, Offord C (2005) Strengthening seed cryo-biology research. SAMARA 9 6
  52. Wood CB, Wood CB, Pritchard HW, Amritphale D (2000) Desiccation-induced dormancy in papaya (Carica papaya L.) is alleviated by heat shock. Seed Sci Res 10: 135-145Google Scholar

Copyright information

© Springer Science+Business Media, LLC 2008

Authors and Affiliations

  • Hugh W. Pritchard
    • 1
  • Jayanthi Nadarajan
    • 1
  1. 1.Seed Conservation DepartmentRoyal Botanic Gardens KewArdinglyUK

Personalised recommendations