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Research Project: Innovative Strategies and Methods for Improving the Management, Availability, and Utility of Plant Genetic Resource Collections

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Title: Development of practical and successful Vitis shoot tip cryopreservation protocols

item BETTONI, JEAN - University Of Santa Catarina
item Bonnart, Remi
item Volk, Gayle

Submitted to: Cryobiology
Publication Type: Abstract Only
Publication Acceptance Date: 1/6/2019
Publication Date: 12/1/2019
Citation: Volk G. 2019. Development of practical and successful Vitis shoot tip cryopreservation protocols. Cryobiology 91:194-195.

Interpretive Summary: n/a

Technical Abstract: Cryopreservation provides a safe and cost-effective means to store plant germplasm of clonally propagated plants that are primarily maintained in vulnerable field collections. Effective cryopreservation procedures have been identified for Vitis using both droplet-vitrification (DV) and V cryo-plate procedures, which resulted in high levels of shoot tip regrowth after liquid nitrogen exposure. In these procedures, in vitro plants (2- to 3-month old) were used as source material for nodal sections. Nodal sections were cultured for 2 weeks and then shoot tips (1.0 mm) were then excised from nodal sections and cultured for three days at 25°C and cryopreserved using either DV on aluminum foil strips or by placement in calcium alginate gel in the wells of aluminium cryo-plates. Shoot tips were then treated for 20 min (DV) or 30 min (V cryo-plate) with loading solution composed of 2 M glycerol and 0.4 M sucrose. The optimum PVS2 exposure was 30 min at 22°C, and then full-strength PVS2 for 90 min at 0°C for the DV method and 30 min of full-strength PVS2 at 22°C using V cryo-plates. Shoot tips were warmed in unloading solution composed of 1.2 M sucrose and placed on recovery medium. Cultures regenerated from these techniques were high quality, and regrowth percentages reached at least of 43% in 13 genotypes representing 12 Vitis species with the DV method. Regrowth percentages =70% in 2 Vitis species with the V cryo-plate method. These procedures are now ready for implementation of Vitis cryostorage in genebanks. In addition to storage of genetic resources, the availability of robust Vitis cryopreservation methods may facilitate the use of cryotherapy methods to eradicate viruses, where vacuolated and differentiated cells that harbor viruses are eliminated because they do not survive LN exposure, producing virus-free plants.