Cryopreservation of Protocorm-Like Bodies of Orchid Hybrid Dendrobium Kasem Boonchoo White by Vitrification

The present study was conducted to develop a protocol for long-term preservation of orchid germplasm. Protocorm-like bodies (PLBs) of orchid hybrid Dendrobium Kasem Boonchoo White were subjected to different treatments of preculture, cryoprotection and dehydration procedures of vitrification technique. The PLBs were precultured on Vacin and Went medium supplemented with 0, 0.06,0.1,0.3,0.5 and 0.7 M sucrose for 3 days. Preculturing with 0.06 to 0.5 M sucrose gave 3.7 to 23.3% viability after cryopreservation. No viable PLBs were found from 0 and 0.7 M sucrose treatments. None of the PLBs survived cryopreservation in all the sucrose concentrations tested. The PLBs were precultured with 0.3 M sucrose for 0, 1, 3, 5, 7 and 9 days. Viability ranged from 11.1 to 26.3% after cryopreservation from 1 to 9 days treatments. No viability was recorded after cryopreservation from treatment without sucrose preculture. There was no survival recorded in all treatments after cryopreservation. When precultured PLBs were cryoprotected with different loading solutions (LSI, LS2, LS3, LS4, LS5 and LS6) for 15 min, all treatments gave 7.4 to 34.4% viability after cryopreservation. All treatments gave 3.7 to 11.1% survival after cryopreservation except LS 1 and LS2 treatments. PLBs were cryoprotected with LS5 for 0, 5, 10, 15, 20, 25 and 30 min. Viability of 7.4 to 36.8% were recorded after cryopreservation when cryoprotected for 5 to 30 min. Without cryoprotection, viability was zero after cryopreservation. The survival rates of 11.1 to 30.3% were obtained after cryopreservation from 5 to 25 min treatments. Cryoprotection for 30 min and without cryoprotection did not give any survival at all after cryopreservation. Cryoprotected PLBs were dehydrated with different vitrification solutions (VS 1, VS2, VS3, VS4 and VS5) for 10 min. All vitrification solutions gave 11.1 to 39.2% viability after cryopreservation. Survival of 7.4 to 26.3% after cryopreservation were recorded in all treatments except VS 1.

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