Welcome to International Network for Natural Sciences | INNSpub

Paper Details

Research Paper | December 1, 2014

| Download 1

Ultrastructural changes during cryopreservation of plumules and embryos of coconut (Cocos nucifera L.)

Oulo Alla-N'Nan, Kouakou Tiécoura, Sery Gonédelé Bi, Jean-Luc Verdeil, Bernard Malaurie

Key Words:

Int. J. Agron. Agri. Res.5(6), 103-115, December 2014


IJAAR 2014 [Generate Certificate]


This article aims to show the changes occurring during cryopreservation of embryos and plumules of coconut which are responsible of their death or survival. Embryos have been cryopreserved by preculture-dehydration for 24h, 28h, 30h, 34h, 38h and 48h on agar medium containing 600g/L of glucose combined with silica gel. The plumules were cryopreserved by encapsulation dehydration on solid medium containing 0.5 M, 0.75 M and 1M followed by dehydration with 40g of silica gel for different durations before rapid freezing. This study indicates that the damages undergone by seed samples can be divided into three types. The first stage of changes concerned the plasmolysis of cells with small vacuoles, condensation of chromatin, changing in the conformation of the DNA and the nucleus and stopping of mitosis. These types of changes are described in general in the context of a desiccation tolerance. The second degree of the changes was the retraction of the cytoplasm inside the cell, the increase in the periplasmic volume. The third degree of modification concerned the deformation of the walls, the invagination or the lysis of the plasma membrane resulting in the observation of distorted cells and and the bursting of the nucleus. These two types of modifications are irreversible and correspond to an absence of regrowth of the samples. Understanding the damage or changes that occur in cryopreserved cells is an important part of understanding how dehydration and frozen affect the viability of recalcitrant plants cells. These changes are made by dehydration and accentuated by freezing.


Copyright © 2014
By Authors and International Network for
Natural Sciences (INNSPUB)
This article is published under the terms of the Creative
Commons Attribution Liscense 4.0

Ultrastructural changes during cryopreservation of plumules and embryos of coconut (Cocos nucifera L.)

Assy Bah B, Durand-Gasselin T, Pannetier C. 1987. Use of zygotic embryo culture to collect germplasm of coconut (Cocos nucifera L.). Plant Genetic Resources Newsletter 71, 4-10.

Assy Bah B, Durand-Gasselin T, Engelmann F, Pannetier C. 1989. Culture in vitro d’embryons zygotiques de cocotier (Cocos nucifera L.). Méthode révisée et simplifiée d’obtention de plants de cocotier transférables en champ. Oléagineux 44(11), 515-523.

Assy Bah B. 1992. Utilisation de la culture d’embryons zygotiques pour la collecte et la conservation des ressources génétiques du cocotier (Cocos nucifera L.). Thèse de doctorat 3ème cycle, Université Pierre et Marie-Curie Paris VI. 159p.

Assy Bah B, Engelmann F. 1992. Cryopreservation of mature embryos of coconut (Cocos nucifera L.) and subsequent regeneration of plantlets. Cryo-Letters. 13, 117-126.

Bajaj YPS. 1984. Induction of growth in frozen embryos of coconut and ovules of Citrus. Current Science 53, 1215–1216.

Bandupriya HDD, Fernando SC, Verdeil JL, Malaurie B. 2010. Cryopreservation of encapsulated plumules of coconut: effect of transport/store conditions. Asia-Pacific journal of molecular biology and biotechnology 18, 135-137.

Bewley JD, Krochko JE. 1982. Desiccation-tolerance. In Lange O. L., Nobel P. S., Osmond CB, Zieler H (eds.). Physiology plant ecology II: Encyclopedia of Plant Physiology 12b, 325-378.

Buffard-Morel J, Verdeil JL, Pannetier C. 1992. Embryogenèse somatique du cocotier (Cocos nucifera L.)  à  partir  de  tissus  foliaires:  etude  histologique. Canadian journal of botany 70, 735-741.

Catesson AM. 1981. Le cycle saisonnier des cellules cambiales chez quelques feuillus. Bulletin de la Société Botanique de France, Actualités Botaniques 128, 43-52.

Chan JL, Sàenz L, Talavera C, Hornung R, Robert M, Oropeza C. 1998. Regeneration of coconut (Cocos nucifera L.) from plumule explants through somatic embryogenesis. Plant cell report 17, 515 – 521.

Chandel KPS, Chaudhury R, Radhamani J, Malik SK. 1995. Desiccation and freezing sensitivity in recalcitrant seeds of tea, cocoa and jackfruit. Annals of botany 76, 443-450.

Chin HF, Krishnapilla J. 1989. A note on the cryopreservation of coconut embryos from young coconuts (Cocos nucifera L.). Pertanika 12(2), 183-186.

Cottignies A. 1981. Arrêt saisonnier de la pré-synthèse de l’ADN en un point de restriction Go. Bulletin de la Société Botanique de France, Actualités Botaniques 128, 53-60.

Dereuddre J. 1981. Effets d’un traitement à 5°C sur l’organisation du réticulum endoplasmique. Bulletin de la Société Botanique de France, Actualités Botaniques 128, 83–89.

Drew PJ, Pammenter NW, Berjak P. 2000. Sub imbibed’ storage is not an option for extending longevity of recalcitrant seeds of tropical species, Trichilia dregeana Sond. Seed science research 10, 355 – 363.

Engelmann F. 1997. Importance of desiccation for the cryopreservation of recalcitrant seed and vegetatively propagated species. Plant genetic resources newsletter 112, 9-18.

Fisher DB. 1968. Protein staining of ribbonedepon sections for light microscopy. Histochemie 16, 92-96.

Golovina EA, Hoekstra FA, Van Aelst AC. 2001. The competence to acquire cellular desiccation tolerance is independent of seed morphological development. Journal of experimental botany 52 (358), 1015-1027.

Helliot B, Swennen R, Poumay Y, Frison E. 2003. Ultrastructural changes associated with cryopreservation of banana (Musa spp.) highly proliferating meristems. Plant cell Reports 21, 690 – 698.

Hornung R, Domas R, Lynch PT. 2001. Cryopreservation of plumulars explants of coconut (Cocos nucifera L.) to support programmes for mass clonal propagation through somatic embryogenesis. Cryo-Letters 2, 211-220.

Kaczmarczyk A, Veli-MattiRokka V, Joachim Keller ER. 2011. Potato Shoot Tip Cryopreservation. A Review. Potato research 54, 45-79.

Karun A, Sajini KK, Parthasarathy VA. 2005. Cryopreservation of mature zygotic embryos of coconut by desiccation methods. Cord 21, 13–19.

Kushhnarenko S, Kovalchuk I, Mukhidinoval Z, Rakhimoval E. 2010. Ultrastructure study of apple meristem cells during cryopreservation. Asian and Australasian journal of plant science and.biotechnology. Global Science Books, 10-20.

Malaurie B, Bandupriya HDD, Fernando SC, Verdeil JL. 2006. Optimisation du procédé de cryoconservation de la plumule de cocotier. Les Actes du BR 6, 449-468.

Mari S, Engelmann F, Chabrillange N, Huet C, Michaux-Ferrière N. 1995. Histo-cytological study of apices of (Coffea racemosa and C. sessiflora) in vitro plantlets during their cryopreservation using the encapsulation – dehydration technique. Cryo-Letters 16, 289-306.

Michaux-Ferrière N. 1981. Le méristème caulinaire de Isoètes Setacea Lam: au cours de son cycle annuel. Bulletin de la Société Botanique de France, Actualités Botaniques 128, 73-82.

N’Nan O, Hocher V, Verdeil JL, Konan JL, Ballo K, Mondeil F, Malaurie B. 2008. Cryopreservation by encapsulation-dehydration of plumules of coconut (Cocos nucifera L.). Cryo-Letters 29(4), 339-350.

N’Nan O, Borges M, Konan KJL, Hocher V, Verdeil JL, Tregear J, N’guetta ASP, Malaurie B. 2012. Simple cryopreservation process of zygotic embryos of ten accessions of coconut (Cocos nucifera L.). In vitro cellular developmental and Biology Plant 48, 160-166.

Pammenter NW, Berjak P. 1999. A review of recalcitrant seed physiology in relation to desiccation tolerance mechanisms. Seed science research 9, 13-37.

Pammenter NW, Greggains V, Kioko JI, Wesley-Smith J, Berjak P, Finch-Savage WE. 1999. Effects of differential drying rates on viability retention of recalcitrant seeds of Ekebergia capensis. Seed science research 8, 463-471.

Reynolds ES. 1963. The use of lead citrate at high pH as on electron opaque stain in electron microscopy. Journal of cell biology 17, 208-212.

Sajini KK, Karun A, Kumaran PM. 2006. Cryopreservation of coconut zygotic embryos by pre growth desiccation. Journal of plant crops 34, 576– 581.

Spurr R. 1969. A low viscosity epoxy resin embedding medium for electron microscopy. Journal of ultrastructure research 26, 31-43.

Verdeil JL, Huet C, Grosdemange F, Buffard-Morel J. 1994. Plant regeneration from cultured immature inflorescences of coconut (Cocos nucifera L.): evidence for somatic embryogenesis. Plant cell reports 13, 218-221.

Verdeil JL, Hocher V, Huet C, Grosdemange F, Escoute J, Ferriere N, Nicole M. 2001. Ultrastructural Changes in Coconut Calli Associated with the Acquisition of Embryogenic Competence. Annals of botany 88, 9-18.

Wesley SJ, Berjak P, Pammenter NW, Vertucci CW. 1995. Ultrastructural evidence for the effects of freezing in embryonic axes of Pisum sativum L. at various water content. Annals of botany 76, 59-64.

Wesley SJ, Pammenter NW, Berjak P, Walters C.  2001.  The  effects  of  two  drying  rates  on  the desiccation tolerance of embryonic axes of recalcitrant jackfruit (Artocarpus heterophyllus Lamk.) seeds. Annals of botany 88, 653 – 664.

Wilkinson T, Wetten A, Prychid C, Fay MF. 2003. Suitability of cryopreservation for long term storage of rare and endangered plant species: a case history for Cosmos astrosanguineus. Annals of botany 91, 65-74.

Withers LA, Engels JM. 1990. The test tube genebank -a safe alternative to field conservation, IPGRI Newsletter for Asia and Pacific 3, 1 – 2.

Xu CX, Li Y, Deng XF, Chen HB, Panis B. 2009. ultrastructural changes in suspension cultures of banana (musa spp) during cryopreservation by vitrification. ISHS ActaI International Symposium on Cryopreservation in Horticultural Species Horticulturae 908, 73.


Style Switcher

Select Layout
Chose Color
Chose Pattren
Chose Background