Articles
Cryopreservation of Paphiopedilum niveum (Rchb. f.) Stein calli using encapsulation-vitrification and vitrification methods
Article number
1167_8
Pages
55 – 62
Language
English
Abstract
Paphiopedilum niveum (Rchb. f.) Stein has been listed in the CITES Appendix I. To evaluate the success of calli cryopreservation of P. niveum, the encapsulation-vitrification and vitrification techniques were compared.
Six-month-old callus clumps were precultured in modified VW liquid medium containing various sucrose concentrations (0, 0.25, 0.50, 0.75 and 1.00 M) for 5 d with daily increasing sucrose concentration and dehydrated in PVS2 at different periods (0, 20, 40, 60, 80 and 100 min). Non-cryopreserved calli were used as a control.
The encapsulation-vitrification technique, which provided the best result, exhibited the highest viability absorbance value (0.237±0.011) and low moisture content (MC) at 27.34±0.96% when calli were precultured in 0.5 M sucrose followed by a 100-minute exposure to PVS2. Histological and histochemical observations also showed normal cell with a large nucleus, a dense cytoplasm, non-disrupted starch grains and protein.
Although the cryopreserved calli via vitrification presented high viability (0.216±0.009) and low moisture content (23.93±2.05%), the vitrification-based calli displayed the disruption of both starch grains and protein.
These cryopreserved calli exhibiting no change in ploidy level provided the survival rates at 29.63±10.31% (encapsulation-vitrification) and 22.22±7.85% (vitrification).
Six-month-old callus clumps were precultured in modified VW liquid medium containing various sucrose concentrations (0, 0.25, 0.50, 0.75 and 1.00 M) for 5 d with daily increasing sucrose concentration and dehydrated in PVS2 at different periods (0, 20, 40, 60, 80 and 100 min). Non-cryopreserved calli were used as a control.
The encapsulation-vitrification technique, which provided the best result, exhibited the highest viability absorbance value (0.237±0.011) and low moisture content (MC) at 27.34±0.96% when calli were precultured in 0.5 M sucrose followed by a 100-minute exposure to PVS2. Histological and histochemical observations also showed normal cell with a large nucleus, a dense cytoplasm, non-disrupted starch grains and protein.
Although the cryopreserved calli via vitrification presented high viability (0.216±0.009) and low moisture content (23.93±2.05%), the vitrification-based calli displayed the disruption of both starch grains and protein.
These cryopreserved calli exhibiting no change in ploidy level provided the survival rates at 29.63±10.31% (encapsulation-vitrification) and 22.22±7.85% (vitrification).
Authors
S. Chaireok, K. Thammasiri, U. Meesawat
Keywords
calli, encapsulation-vitrification, histochemistry, histology, Paphiopedilum, ploidy level, vitrification
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