SURVIVAL BY VITRIFICATION OF NUCELLAR CELLS OF NAVEL ORANGE (CITRUS-SINENSIS VAR BRASILIENSIS TANAKA) COOLED TO 196-DEGREES-C

The nucellar cells of navel orange (Citrus sinensis Osb. var. brasiliensis Tanaka) with potential for somatic embryo production have been successfully cryopreserved for 40 days by vitrification. In this method, cells were sufficiently dehydrated with a highly concentrated cryoprotective solution (designated PVS2) to be capable of vitrifying during rapid cooling in liquid nitrogen. The cells treated with PVS2 at 25-degrees-C for 3 min were transferred to a 1.8 mL plastic cryotube and then directly plunged into liquid nitrogen. The average rate of survival was about 90%. The revived cells resumed growth within 3 days and produced cotyledonary embryoids that developed into plants within 3 months of culture. This simple method has been successfully applied to six other species or cultivars with high rates of survival. The time used for the procedure is only several minutes. This novel method for cryopreservation seems promising as a practical method applicable to a wide range of cultured plant cells and meristems.