ISHS Acta Horticulturae 1039: II International Symposium on Plant Cryopreservation
USING A DROPLET-VITRIFICATION METHOD TO PARTIALLY OVERCOME THE RECALCITRANCE OF CASSAVA TO CRYOSTORAGE |
| Authors: | R.H. Escobar, L. Muñoz, A. Rios, A. Núñez, J. Tohme |
| Keywords: | Manihot esculenta Crantz, cryopreservation, PVS2, black-box, long-term storage |
| DOI: | 10.17660/ActaHortic.2014.1039.29 |
| Abstract:
Cassava cryopreservation through droplet-vitrification has been implemented at CIAT to partially overcome the recalcitrance of this crop to cryobanking. A cassava clone is considered recalcitrant if its response (data obtained independently at least twice) after the freezing phase is less than 30% complete plant formation. One hundred clones that were proven to be recalcitrant using encapsulation-dehydration technique were selected for testing with an adapted droplet-vitrification method. Twenty shoot tips per clone were treated with loading solution for 2 h. Later, the tips were removed and transferred to Plant Vitrified Solution 2 for 30 min. Preliminary data showed that cassava is sensitive to the solution, so a critical point to consider with this method is the duration of each step before moving to the freezing phase. Treated tips were frozen quickly by immersion in liquid nitrogen. The recovery phase was carried out by successive changes in two different growth media, with the critical factors being the control of duration of exposure to the initial medium, light intensity, and the combination of kinetin and gibberellic acid, because these factors allow a slow recovery of shoot tip green-tissues and then plant conversion. No or minimal callus formation was observed in the regeneration phase. Some 70-75% of the clones tested surpassed the standard for full plant recovery (being considered as a successful cryo event), and overcame the previous recalcitrance to the encapsulation-dehydration method. Adjustments in time duration and recovery media are being tested to try to recover more clones after freezing. Cassava plants recovered from frozen tissues and from in vitro controls are transferred to the field for further evaluation and observation on parameters such as yield and morphological descriptors. | |
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