Droplet vitrification of apical meristems: a cryopreservation protocol applicable to all Musaceae

One millimeter sized shoot-tips excised from rooted in vitro plants of the genera Musa and Ensete were successfully cryopreserved with the droplet vitrification technique. We show that the loading phase can be prolonged up to 7 h and that the optimal length of PVS2 treatment is 30-50 min at 0 °C. Ultra rapid freezing and thawing rates proved to be essential for high and reproducible post-thaw regeneration rates. This results in an increase of 23-46% compared to the normal cryovial freezing protocol. When the optimal procedure was applied to 56 accessions belonging to eight different genomic groups of Musa spp. and one Ensete spp., an average of 52.9% post-thaw regeneration was obtained. These results were relatively genotype independent. Only wild diploid Musa acuminata accessions proved to be somewhat more recalcitrant towards cryopreservation though an acceptable average regeneration rate of 39% was still obtained.