Articles
Application of differential thermal analysis for the improved regeneration after cryopreservation of vegetatively-propagated plants
Article number
1234_7
Pages
57 – 64
Language
English
Abstract
Three critical key parameters for cryoprotocol improvement resulting in increasing the regeneration rate after cryopreservation can be determined by thermal analysis: 1) the content of frozen water.
The boundary content of frozen water affected by dehydration over desiccants or a mixture of cryoprotectants can be determined by differential scanning calorimetry; in the case of cryopreservation of garlic shoot tips, by dehydration in different concentrations of Plant Vitrification Solution No 3 (PVS3). Particular concentrations of cryoprotectants can regulate the final frozen water content.
Although the regeneration of garlic shoot tips after their exposure to diluted PVS3 (i.e., sucrose/glycerol components 45/45) was similar to PVS3 (sucrose/glycerol 50/50), the use of less concentrated PVS3 reduces the risk of dehydration damage before ultra-low temperature application; 2) the content of unfreezable water.
The intersection of the content of frozen water and the absolute content of water can be determined gravimetrically in shoot tips at different levels of dehydration and the content of unfreezable water derived.
The unfreezable water content can be the limit of the minimum water content below which the shoot tips cannot survive dehydration; and 3) glass transition temperature.
Determination of the glass transition temperature is important for long-term cryopreservation without undesirable alterations to the sample.
A pure cryoprotectant, added usually in an excessive amount, has a constant Tg; whereas, the Tg of shoot tips increases as the content of water removed with the cryoprotective solution decreases.
The highest recovery level after cryopreservation treatment should be achieved at the highest obtained sample glass transition temperature.
The three above-mentioned parameters define the threshold dehydration level of shoot tips important for a high regeneration rate after cryopreservation.
The boundary content of frozen water affected by dehydration over desiccants or a mixture of cryoprotectants can be determined by differential scanning calorimetry; in the case of cryopreservation of garlic shoot tips, by dehydration in different concentrations of Plant Vitrification Solution No 3 (PVS3). Particular concentrations of cryoprotectants can regulate the final frozen water content.
Although the regeneration of garlic shoot tips after their exposure to diluted PVS3 (i.e., sucrose/glycerol components 45/45) was similar to PVS3 (sucrose/glycerol 50/50), the use of less concentrated PVS3 reduces the risk of dehydration damage before ultra-low temperature application; 2) the content of unfreezable water.
The intersection of the content of frozen water and the absolute content of water can be determined gravimetrically in shoot tips at different levels of dehydration and the content of unfreezable water derived.
The unfreezable water content can be the limit of the minimum water content below which the shoot tips cannot survive dehydration; and 3) glass transition temperature.
Determination of the glass transition temperature is important for long-term cryopreservation without undesirable alterations to the sample.
A pure cryoprotectant, added usually in an excessive amount, has a constant Tg; whereas, the Tg of shoot tips increases as the content of water removed with the cryoprotective solution decreases.
The highest recovery level after cryopreservation treatment should be achieved at the highest obtained sample glass transition temperature.
The three above-mentioned parameters define the threshold dehydration level of shoot tips important for a high regeneration rate after cryopreservation.
Authors
J. Zámečník, M. Faltus, A. Bilavčík
Keywords
differential scanning calorimeter, frozen water content, unfreezable water content, glass transition temperature, plant vitrification solution
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