Pre-treatment with salicylic acid improves plant regeneration after cryopreservation of grapevine (Vitis spp.) by droplet vitrification

Despite many reports on regeneration of Vitis after cryopreservation, there is no cryopreserved collection of its germplasm. Some Vitis genotypes are reported to be recalcitrant to cryopreservation. Droplet vitrification (DV), considered to be an emerging generic method of cryopreservation, has been applied only to a limited extent in Vitis. In the present study, we first tested the toxicity to vitrification solution of both axillary and apical buds in six diverse Vitis accessions. DV was then applied using 50 % predicted survival time of apical and axillary buds in vitrification solution after pre-treatment of in vitro cultured plantlets with salicylic acid (SA), a substance known to have a protective role in abiotic stress responses. Results showed that axillary buds are more tolerant of vitrification solution than apical buds and required longer treatment time. Pre-treatment of donor plants with 0.1 mM salicylic acid resulted in a significantly higher protection to cryopreserved buds, but serial dehydration in sucrose alone had little effect. SA treatment enabled successful cryopreservation of previously recalcitrant rootstock 41B, albeit at a low regeneration rate. For all other genotypes tested, cryopreservation of 6-11 explants will be sufficient to regenerate at least one plant at 95 % probability. This is the first report of successful cryopreservation of a set of diverse Vitis genotypes by DV, and we show that pre-treatment of donor plants with SA is critical for the success. This research will contribute to conservation of Vitis germplasm in a cost-effective way avoiding the risks associated with field-based collections.