Investigation on Vitrification Protocols of Cryopreservation of Taiwan Native Plant of Nothapodytes nimmoniana (Graham) Mabberley

• Main Authors: Yen-Ling Peng, 彭燕玲
• Other Authors: 廖松淵
• Format: Others
• Language: zh-TW
• Published: 2012
• Online Access: http://ndltd.ncl.edu.tw/handle/8344bf

碩士 === 國立中興大學 === 生命科學系所 === 100 === This study was used in vitro-grown of Nothapodytes nimmoniana (Graham) Mabberley as experimental materials, and the excised shoot tips were cryopreserved by vitrification method. Investigated the injury of plant vitrification solutions and different preculture treatments by physiological analysis, and the effect on survival ratio by physiological state changed of shoot tip. 60-90 day-old plants were precultured and then excised 1-2 mm shoot tips for different experiments, treated with different duration of PVS2, LS and different precultured medium and days. In this study, the results demonstrated that the optimal cryopreservation protocol of Nothapodytes nimmoniana (Graham) Mabberley was precultured on 0.5 M sucrose medium for 3 days, then treated with LS for 90 min at room temperature and PVS2 for 90 min at 0℃ for long-term storage. Buds rewarmed rapidly after one week, then plated on petri dishes containing WPM medium, supplemented with BA 1mgL-1, and IBA 0.1 mgL-1. The survival ratio in this protocol could achieve 72.2% . In the cryopreservation protocol, the precultured treatment decreased the relative water content, water potential, and osmotic potential. The precultured treatment also increased the content of soluble protein and soluble sugar in shoot tips, and the soluble sugar content is the key factor affected osmotic potential. The content significantly increased to decrease the osmotic potential, regulated the physiological state of shoot tips with osmotic adjustment. Osmotic adjustment kept turgor pressure stably, and increased the drought and freezing tolerance, then plant could adapt osmotic stress during precultured treatment. Furthermore, LS and PVS2 were the key factors affected survival ratio of cryopreservation. If the exposure time of LS and PVS2 shorter than optimal, the dehydration and protection were not enough. If the time longer than optimal, the serious osmotic stress and toxicity were injured the shoot tips. The results indicated that shoot tips cultured on sugar enrich medium cooperated with suitable exposure time of LS and PVS2, which the dehydration and injury were balanced, and thus enhanced the survival ratio of Nothapodytes nimmoniana (Graham) Mabberley after cryopreservation.