In Vitro Conservation and Cryopreservation of Ornamental Plants

  • Elif Aylin Ozudogru
  • Alberto Previati
  • Maurizio Lambardi
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 589)

Abstract

Today, the conservation of ornamental germplasm can take advantage of innovative techniques which allow preservation in vitro (slow growth storage) or in liquid nitrogen (cryopreservation) of plant material. Slow growth storage refers to the techniques enabling the in vitro conservation of shoot cultures in aseptic conditions by reducing markedly the frequency of periodic subculturing, without affecting the viability and regrowth of shoot cultures. Cryopreservation refers to the storage of explants from tissue culture at ultra-low temperature (−196°C). At such temperature, all the biological reactions within the cells are hampered, hence the technique makes available the storage of plant material for theoretically unlimited periods of time. An exhaustive review of papers dealing with the slow growth storage and the cryopreservation of ornamental species is reported here. Step-by-step protocols for the slow growth storage of rose germplasm, the production of synthetic seeds for the in vitro conservation of ornamentals, and the cryopreservation of Chrysanthemum morifolium are included.

Key words

Cryopreservation In vitro conservation Liquid nitrogen Slow growth storage Synthetic seeds 

References

  1. 1.
    Turner T (2005) Garden history : Philosophy and design 2000 BC–2000 AD. Abingdon, Spon 304 ppGoogle Scholar
  2. 2.
    Reed BM (1992) Cold storage of strawberries in vitro: A comparison of three storage systems. Fruit Varieties J 46:98–102Google Scholar
  3. 3.
    Lambardi M, De Carlo A (2003) Application of tissue culture to the germplasm conservation of temperate broad-leaf trees. In: Jain SM, Ishii K (eds) Micropropagation of woody trees and fruits. Kluwer, The Netherlands, pp 815–840Google Scholar
  4. 4.
    Lambardi M, Benelli C, De Carlo A, Fabbri A, Grassi S, Lynch PT (2002) Medium- and long-term in vitro conservation of olive germplasm (Olea europea L.). Acta Hortic 586:109–112Google Scholar
  5. 5.
    Withers LA, Engelmann F (1997) In vitro conservation of plant genetic resources. In: Altman A (ed) Biotechnology in agriculture. Marcel Dekker, New York, pp 57–88CrossRefGoogle Scholar
  6. 6.
    Ashmore SE (1997) Status report on the development and application of in vitro techniques for the conservation and use of plant genetic resources. International Plant Genetic Resources Institute, Rome 67 ppGoogle Scholar
  7. 7.
    Benson EE (2008) Cryopreservation theory. In: Reed BM (ed) Plant cryopreservation: A practical guide. Springer, USA, pp 15–32CrossRefGoogle Scholar
  8. 8.
    Fahy GM, MacFarlane DR, Angell CA, Meryman HT (1984) Vitrification as an approach to cryopreservation. Cryobiology 21:407–426CrossRefPubMedGoogle Scholar
  9. 9.
    Sakai A, Hirai D, Niino T (2008) Development of PVS-based vitrification and encapsulation-vitrification protocols. In: Reed BM (ed) Plant cryopreservation: A practical guide. Springer, USA, pp 33–57CrossRefGoogle Scholar
  10. 10.
    Lambardi M, Ozudogru EA, Benelli C (2008) Cryopreservation of embryonic cultures. In. Reed BM (ed) Plant cryopreservation: A practical guide. Springer, USA, pp 177–210Google Scholar
  11. 11.
    Walters C, Wesley-Smith J, Crane J, Hill LM, Chmielarz P, Pammenter NW, Berjak P (2008) Cryopreservation of recalcitrant (i.e. desiccation-sensitive) seeds. In: Reed BM (ed) Plant cryopreservation: A practical guide. Springer, USA, pp 465–484Google Scholar
  12. 12.
    Pritchard HW, Nadarajan J (2008) Cryopreservation of orthodox (desiccation tolerant) seeds. In: Reed BM (ed) Plant cryopreservation: A practical guide. Springer, USA, pp 485–501Google Scholar
  13. 13.
    Normah MN, Makeen AM (2008) Cryopreservation of excised embryos and embryonic axes. In: Reed BM (ed) Plant cryopreservation: A practical guide. Springer, USA, pp 211–240Google Scholar
  14. 14.
    Towill LE, Ellis DD (2008) Cryopreservation of dormant buds. In: Reed BM (ed) Plant cryopreservation: A practical guide. Springer, USA, pp 421–464Google Scholar
  15. 15.
    Sakai A, Kaboyashi S, Oiyama I (1990) Cryopreservation of nucellar cells of navel orange (Citrus sinensis Osb. var. brasiliensis Tanaka) by vitrification. Plant Cell Rep 9:30–33CrossRefGoogle Scholar
  16. 16.
    Fabre J, Dereuddre J (1990) Encapsulation-dehydration: A new approach to cryopreservation of Solanum shoot tips. CryoLetters 11:413–426Google Scholar
  17. 17.
    Panis B, Lambardi M (2006) Status of cryopreservation technologies in plants (crops and forest trees). In: Ruane J, Sonnino A (eds) The role of biotechnology in exploring and protecting agricultural genetic resources. Food and Agriculture Organization of the United Nations, Rome, Italy, pp 61–78Google Scholar
  18. 18.
    Murashige T (1977) Plant cell and organ cultures as horticultural practices. Acta Hortic 78:17–30Google Scholar
  19. 19.
    Lambardi M, Benelli C, Ozudogru EA, Ozden-Tokatli Y (2006) Synthetic seed technology in ornamental plants. In: Teixeira da Silva JA (ed) Floriculture, ornamental and plant biotechnology: Advances and topical issues, vol II. Global Science Books, UK, pp 347–354Google Scholar
  20. 20.
    Redenbaugh K, Fujii J, Slade D, Viss P, Kossler M (1991) Artificial seeds-Encapsulated somatic embryos. In: Bajaj YPS (ed) Biotechnology in agriculture and forestry, high-tech and micropropagation. Springer, Heidelberg, pp 395–416Google Scholar
  21. 21.
    Ballester A, Janeiro LV, Vieitez AM (1997) Cold storage of shoot cultures and alginate encapsulation of shoot tips of Camellia japonica L. and Camellia reticulata Lindley. Sci Hortic 71:67–78CrossRefGoogle Scholar
  22. 22.
    Reed BM, Okut N, Achino JD, Narver L, DeNoma J (2003) Cold storage and cryopreservation of hops (Humulus L.) shoot cultures through application of standard protocols. CryoLetters 24:389–396PubMedGoogle Scholar
  23. 23.
    Nunes EDC, Benson EE, Oltramari AC, Araujo PS, Righetto M, Viana M (2003) In vitro conservation of Cederla fissilis Vellozo (Meliaceae), a native tree of the Brazilian Atlantic Forest. Biodivers Conserv 12:837–848CrossRefGoogle Scholar
  24. 24.
    West TP, Ravindra MB, Preece JE (2006) Encapsulation, cold storage, and growth of Hibiscus moscheutos nodal segments. Plant Cell Tissue Organ Cult 87:223–231CrossRefGoogle Scholar
  25. 25.
    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 88:41–49CrossRefGoogle Scholar
  26. 26.
    Orlikowska T (1992) Effect of in vitro storage at 4°C on surviving and proliferation of two apple rootstocks. Plant Cell Tissue Organ Cult 31:1–7Google Scholar
  27. 27.
    Hempel T, Hempel M (1984) Storage of gerbera propagated in vitro. In: Proceedings of International Symposium on “Plant Tissue and Cell Culture Application to Crop Improvement”, Czechoslovakia, pp 559–560Google Scholar
  28. 28.
    Previati A, Benelli C, Da Re F, Ozudogru A, Lambardi M (2008) Micropropagation and in vitro conservation of virus-free rose germplasm. Propagat Ornam Plants 8:93–98Google Scholar
  29. 29.
    Janeiro LV, Ballester A, Vieitez AM (1997) In vitro response of encapsulated somatic embryos of camellia. Plant Cell Tissue Organ Cult 51:119–125CrossRefGoogle Scholar
  30. 30.
    Pattnaik S, Chand PK (2000) Morphogenic response of the alginate-encapsulated axillary buds from in vitro shoot cultures of six mulberries. Plant Cell Tissue Organ Cult 60:177–185CrossRefGoogle Scholar
  31. 31.
    Ozden-Tokatli Y, De Carlo A, Gumusel F, Pignattelli S, Lambardi M (2008) Development of encapsulation techniques for the production and conservation of synthetic seeds in ornamental plants. Propagat Ornam Plants 8(1):17–22Google Scholar
  32. 32.
    Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant 15:473–479CrossRefGoogle Scholar
  33. 33.
    Quoirin M, LePoivre P (1977) Etude de millieux adaptés aux cultures in vitro de Prunus. Acta Hortic 78:437–442Google Scholar
  34. 34.
    Patel AV, Pusch I, Mix-Wagner G, Vorlop KD (2000) A novel encapsulation technique for the production of artificial seeds. Plant Cell Rep 19:868–874CrossRefGoogle Scholar
  35. 35.
    Standardi A, Micheli M, Piccioni E (1995) Incapsulamento in alginato di espianti micropropagati. Italus Hortus 2(1–2):46–48Google Scholar
  36. 36.
    Ipekci Z, Gozukirmizi N (2003) Direct somatic embryogenesis and synthetic seed production from Paulownia elongate. Plant Cell Rep 22:16–24CrossRefPubMedGoogle Scholar
  37. 37.
    Refouvelet E, Le Nours S, Tallon C, Daguin F (1998) A new method for in vitro propagation of lilac (Syringa vulgaris L.): Regrowth and storage conditions for axillary bud encapsulation in alginate beads, development of a pre-acclimatisation stage. Sci Hortic 74:233–241CrossRefGoogle Scholar
  38. 38.
    Halmagyi A, Fischer-Klüver G, Mix-Wagner G, Schumacher HM (2004) Cryopreservation of Chrysanthemum morifolium (Dendranthema grandiflora Ramat.) using different approaches. Plant Cell Rep 22:371–375CrossRefPubMedGoogle Scholar
  39. 39.
    Schäfer-Menuhr A, Mix-Wagner G, SchumacherHM (1997) Cryopreservation of potato cultivars-design of a method for routine application in genebanks. Acta Hortic 447:447–483Google Scholar
  40. 40.
    Miao N-H, Kaneko Y, Sugawara Y (2005) Ultrastructural implications of pretreatment for successful cryopreservation of Oncidium protocorm-like body. CryoLetters 26:333–340PubMedGoogle Scholar
  41. 41.
    Halmagyi A, Pinker I (2006) Cryopreservation of Rosa shoot tips: importance of preculture conditions. Acta Hortic 725:351–356Google Scholar
  42. 42.
    Martin C, González-Benito ME (2005) Survival and genetic stability of Dendranthema grandiflora Tzvelev shoot apices after cryopreservation by vitrification and encapsulation-dehydration. Cryobiology 51:281–289CrossRefPubMedGoogle Scholar
  43. 43.
    Tanaka D, Niino T, Isuzugawa K, Hikage T, Uemura M (2004) Cryopreservation of shoot apices of in-vitro grown gentian plants: comparison of vitrification and encapsulation-vitrification protocols. CryoLetters 25:167–176PubMedGoogle Scholar
  44. 44.
    Lambardi M, De Carlo A, Capuana M (2005) Cryopreservation of embryogenic callus of Aesculus hippocastanum L. by vitrification/one-step freezing. CryoLetters 26:185–192PubMedGoogle Scholar
  45. 45.
    Halmagyi A, Deliu C (2007) Cryopreservation of carnation (Dianthus caryophyllus L.) shoot tips by encapsulation-vitrification. Sci Hortic 113:300–306CrossRefGoogle Scholar
  46. 46.
    Lynch P, Harris WC, Chartier-Hollis JM (1996) The cryopreservation of shoot tips of Rosa multiflora. Plant Growth Regul 20:43–45CrossRefGoogle Scholar
  47. 47.
    Johnson JW, Harding K, Benson EE (2007) Antioxidant status and genotypic tolerance of Ribes in vitro cultures to cryopreservation. Plant Sci 172:524–534CrossRefGoogle Scholar
  48. 48.
    Fukai S, Goi M, Tanaka M (1991) Cryo-preservation of shoot tips of Chrysanthemum morifolium and related species native to Japan. Euphytica 54:201–204Google Scholar
  49. 49.
    Fukai S, Goi M, Tanaka M (1991) Cryopreservation of shoot tips of Carophyllaceae ornamentals. Euphytica 56:149–153CrossRefGoogle Scholar
  50. 50.
    Fukai S, Oe M (1990) Morphological observations of chrysanthemum shoot tips cultured after cryopreservation and freezing. J Jpn Soc Hort Sci 59:383–387CrossRefGoogle Scholar
  51. 51.
    Siebert M, Wetherbee PJ (1977) Increased survival and differentiation of frozen herbaceous plant organ cultures through cold treatment. Plant Physiol 59:1043–1046CrossRefGoogle Scholar
  52. 52.
    Halmagyi A, Pinker I (2006) Plant regeneration from Rosa shoot tips cryopreserved by a combined droplet vitrification method. Plant Cell Tissue Organ Cult 84:145–153CrossRefGoogle Scholar
  53. 53.
    Fukai S, Goi M, Tanaka M (1994) The chimeric structure of the apical dome of chrysanthemum (Dendranthema grandiflorum (Ramat.) Kitam.) is affected by cryopreservation. Sci Hortic 57:347–351CrossRefGoogle Scholar
  54. 54.
    Harding K (2004) Genetic integrity of cryopreserved plant cells: A review. CryoLetters 25:3–22PubMedGoogle Scholar
  55. 55.
    Park Y-G, Kwon G-S, Tay D (2005) Cryopreservation for gene conservation of Acer mono Max. Propagat Ornam Plants 5(2):78–83Google Scholar
  56. 56.
    Seo MJ, Shin JH, Sohn JK (2007) Cryo-preservation of dormant herbaceous peony (Paeonia lactiflora Pall.) shoot-tips by desiccation. CryoLetters 28:207–213PubMedGoogle Scholar
  57. 57.
    Ryyänen L, Aronen T (2007) Phenotypic expression of leaf variegation in two Betula pendula Roth genotypes following micropropagation, cryopreservation and grafting. Propagat Ornam Plants 7(1):23–28Google Scholar
  58. 58.
    Winkelmann T, Mußmann V, Serek M (2004) Cryopreservation of embryogenic suspension cultures of Cyclamen persicum Mill. Plant Cell Rep 23:1–8CrossRefPubMedGoogle Scholar
  59. 59.
    Scocchi A, Vila S, Mroginski L, Engelmann F (2007) Cryopreservation of somatic embryos of paradise tree (Melia azedarach L.). CryoLetters 28:281–290PubMedGoogle Scholar
  60. 60.
    Gale S, John A, Benson EE (2007) Cryo-preservation of Picea sitchensis (Sitka spruce) embryogenic suspensor masses. CryoLetters 28:225–239PubMedGoogle Scholar
  61. 61.
    Towill LE, Widrlechner M (2004) Cryopreser-vation of Salix species using sections from winter vegetative scions. CryoLetters 25:71–80PubMedGoogle Scholar
  62. 62.
    Sant R, Taylor M, Tyagi A (2006) Cryopreserva-tion of in vitro-grown shoot-tips of tropical taro (Colocasia esculenta var. Esculenta) by vitrification. CryoLetters 27:133–142PubMedGoogle Scholar
  63. 63.
    Thammasiri K (2000) Cryopreservation of seeds of a Thai orchid (Doritis pulcherrima Lindl.) by vitrification. CryoLetters 21:237–244PubMedGoogle Scholar
  64. 64.
    Zaimah NA, Benson EE, Marzalina M (2007) Viability of Elateriospermum tapos (Perah) embryo after storage in liquid nitrogen. J Trop For Sci 19(1):1–5Google Scholar
  65. 65.
    Matsumoto T, Takahashi C, Sakai A, Nako Y (1998) Cryopreservation of in vitro-grown apical meristems of hybrid statice by three different procedures. Sci Hortic 76:105–114CrossRefGoogle Scholar
  66. 66.
    Yang B-H, Kim H-T, Park J-Y, Park Y-G (2006) Cryopreservation of in vitro-cultured axillary shoot tips of Japanese bead tree (Melia azedarach) using vitrification technique. Korean J Plant Res 19(3):385–391Google Scholar
  67. 67.
    Hirano T, Ishikawa K, Mii M (2005) Cryopreservation of immature seeds of Ponerorchis graminifolia var. suzukiana by vitrification. CryoLetters 26:139–146PubMedGoogle Scholar
  68. 68.
    Nadarajan J, Staines HJ, Benson EE, Mansor M, Krishnapillay B, Harding K (2007) Optimization of cryopreservation for Sterculia cordata zygotic embryos using vitrification techniques. J Trop For Sci 19(2):79–85Google Scholar
  69. 69.
    Kim H-H, Cha Y-S, Baek H-J, Cho E-G, Chae Y-A, Engelmann F (2002) Cryopreservation of tea (Camellia sinensis L.) seeds and embryonic axes. CryoLetters 23:209–216PubMedGoogle Scholar
  70. 70.
    Wood CB, Pritchard HW, Miller AP (2000) Simultaneous preservation of orchid seed and its fungal symbiont using encapsulation-dehydration is dependent on moisture content and storage temperature. CryoLetters 21:125–136PubMedGoogle Scholar
  71. 71.
    Fukai S, Togashi M, Goi M (1994) Cryopreservation of in vitro-grown Dianthus by encapsulation-dehydration. Tech Bull Fac Agric Kagawa Univ 46:101–107Google Scholar
  72. 72.
    Flachsland E, Terada G, Scocchi A, Rey H, Mroginski L, Engelmann F (2006) Cryopreservation of seeds and in vitro-cultured protocorms of Oncidium bifolium Sims. (Orchidaceae) by encapsulation-dehydration. CryoLetters 27:235–242Google Scholar
  73. 73.
    Verleysen H, Bockstaele EV, Debergh P (2005) An encapsulation-dehydration protocol for cryopreservation of the azalea cultivar ‘Nordlicht’ (Rhododendron simsii Planch.). Sci Hortic 106:402–414CrossRefGoogle Scholar
  74. 74.
    Driwer JA, Kuniyuku AH (1984) In vitro propagation of paradox walnut rootstock. HortScience 19:507–509Google Scholar

Copyright information

© Humana Press, a part of Springer Science+Business Media, LLC 2010

Authors and Affiliations

  • Elif Aylin Ozudogru
    • 1
  • Alberto Previati
    • 2
  • Maurizio Lambardi
    • 3
  1. 1.Department of Biology, Laboratory of Plant Tissue CultureGebze Institute of TechnologyKocaeliTurkey
  2. 2.Centro Sperimentale Ortofloricolo “Po di Tramontana”, Veneto AgricolturaRosolinaItaly
  3. 3.IVALSA/Istituto per la Valorizzazione del Legno e delle Specie Arboree, National Research Council (CNR)Sesto, FiorentinoItaly

Personalised recommendations