CRYOPRESERVATION OF WOODY PLANTS BY AXILLARY BUD VITRIFICATION: A FIRST APPROACH WITH POPLAR

Abstract

Axillary buds from in vitro-grown stock plants of White Poplar (Populus alba L.) were utilized to test the efficiency of the vitrification technique, as cryoprotective during direct freezing in liquid nitrogen. Excised axillary buds were precultured for 2 days at 5°C on a gelled MS medium, loaded for 20 min at 25°C with a cryoprotectant (2 M glycerol and 0.4 M sucrose, 1: 1), and then exposed to a concentrated vitrification solution (PVS2: 30% glycerol, 15% ethylene glycol, 15% DMSO in MS medium). After vitrification, the buds were immersed directly into liquid nitrogen for at least one hour. Successfully vitrified buds remained green after plating, and resumed growth within 2 weeks. The highest percentage of bud survival (90%) was obtained when the buds were precultured on MS medium containing 0.09 M sucrose, and vitrified by exposure to PVS2 for 60 min at 0°C. The average rate of healthy shoot development from the surviving buds amounted to about 60%. Vitrification procedure can be proposed as an useful technique for germplasm cryopreservation of woody plants.