ISHS Acta Horticulturae 538: Eucarpia symposium on Fruit Breeding and Genetics
LOW TEMPERATURE STORAGE AND CRYOPRESERVATION OF A VITIS VINIFERA L. GERMPLASM COLLECTION: FIRST RESULTS |
| Authors: | M.L. Miaja, I. Gribaudo, R. Vallania, L.F. Fernandez |
| Keywords: | grapevine, encapsulation-dehydration, bud, in vitro culture, sucrose, germplasm preservation. |
| Abstract:
An in vitro collection of clones of Piemonte, Valle d'Aosta and Liguria grapevine cultivars has been created and maintained since 1989. The possibility of genetic instability and the labour required to maintain field grown collections lead to the need for storing part of the collection at reduced temperature. Two experimental procedures were evaluated: cultivation at relatively low temperature (+11°C) of plantlets or buds, and cryopreservation of axillary buds. In both cases, explants were taken from in vitro plantlets cultured on a modified Murashige and Skoog (MMS) medium without growth regulators. In the first storage procedure, three kinds of cultures were grown for 4 months at +11°C, 10 mmol sec-1 m-2 light intensity and 16 hrs photoperiod: 1) freshly excised axillary buds, on MMS growth regulator-free medium; 2) freshly excised axillary buds, on MMS medium +3 mM BA; 3) rooted plantlets, on MMS growth regulator-free medium. At the end of the storage period, they were placed at +25°C and their subsequent growth was recorded after 10 more days. Culturing axillary buds at +11°C inhibited bud development, which was only partially resumed when the temperature was increased at +25°C. The recovery was higher if BA was present in the medium. Culture of rooted plantlets at +11°C slowed growth, and all the cultured explants survived the storage period. In the second procedure axillary buds were embedded in alginate beads and before immersion in liquid nitrogen were precultured in liquid culture media containing increasing concentrations of sucrose and then dehydrated under sterile air flow at +25°C for 4 hours. After dehydration the beads were transferred into cryovials for freezing. Samples were kept in liquid nitrogen for at least one day and then beads containing shoot-tips were thawed at +40°C in water bath and transferred to recovery medium. Plantlet recovery was observed in a low percentage of explants, while others remained green but did not develop further. | |
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