- 學位論文
台灣白及種子以矽膠乾燥前處理之超低溫保存及其應用
Cryopreservation of Bletilla formosana Seed by Silica Gel Desiccation and Its Application
摘要
蘭花遺傳資源的長期保存非常重要，是維繫台灣蘭花產業永續發展與維護生物多樣性極重要的盤石，本研究選定兼具藥用與觀賞價值的台灣原生蘭-台灣白及種子為材料，建立超低溫長期保存技術平台，並進一步應用至其他經濟蘭花種原保存。結果顯示，以矽膠乾燥或於室溫陰乾方式進行前處理24小時，再置入液態氮保存，有助於提高臺灣白及種子之活力率與發芽率。再者，台灣白及以黃綠色果莢、成熟度110天的種子為理想材料，配合矽膠乾燥前處理24小時，可使種子含水量降至5%以下，後續進行-196℃液態氮的超低溫保存，可以使保存24小時後的種子維持高發芽率(95.5%)，此證實台灣白及種子能夠忍受乾燥且不會失去種子活力，因此台灣白及種子應屬於正貯型種子(orthodox seed)。而且隨著超低溫保存期間延長，於液態氮中保存48個月(4年)後仍能維持87.7%的高發芽率，經播種後，種子可正常發芽、小苗生長發育正常，與未經超低溫保存者外表型並無不同。保存前經乾燥處理之低成熟度種子，於液態氮保存後發芽率有增加趨勢，經電子顯微鏡觀察，種皮受到破壞，可能因此提供水份進入的管道。本研究建議，常用的遺傳資源(種子與花粉)活力檢定法─TTC染色法(2, 3, 5-triphenyltetrazolium chloride)宜應用在檢定成熟度較高的蘭花種子，所得結果比較接近種子真實的活力。應用此結果進一步試驗紅花鶴頂蘭(Phaius tankervilleae)花粉、蝴蝶蘭(Phalaenopsis aphrodite subsp. formosana)與萬代蘭(Vanda lamellata與Vanda denisoniana)的種子，亦可以成功地以超低溫技術進行長期保存。
並列摘要
The development of a long-term conservation method is crucial for orchid germplasm preservation and the orchid industry in general. Taiwan native orchid-Bletilla formosana (Hayata) Schltr. seeds used as cryopreservation materials, established the platform for long-term preservation technology and further applied in other economic orchid species. The results showed that the viability and germination percentage of seeds desiccated by silica gel or air-drying for 24 h in laboratory conditions prior to cryopreservation was increased. Seeds in 110 DAP yellow-green capsule were suggested as materials and desiccated by silica gel for 24h prior to cryopreservation. Water content of seeds was decreased below 5%, but high germination percentage still remained (96.7%) after preservation for 48h. It can be seen that B. formosana seeds should be the orthodox seed preserved for long-term preservation because of high drought tolerance and high viability after cryostorage. Germination percentage can been still maintain high at least 48 months after cryopreservation. The plantlet from cryostoraged seeds could be grown and developed well. The plantlet from cryostoraged seeds could be grown and developed well. The seed vigor test adopting a TTC staining method was suitable for seeds with high maturity. The practical desiccation method could be applied for cryopreservation of Phaius tankervilleae polllen, Phalaenopsis aphrodite subsp. formosana , Vanda lamellate and Vanda denisoniana seeds successfully.