| Summary: | 碩士 === 國立中興大學 === 生命科學系所 === 102 === Cryopreservation of different explants has different dehydration process. To compare effectiveness of dehydration method and cryopreservation process on different
explants, we use the pollen of cucumber “Xiu Yan”, the shoot apical meristem of Phalaenopsis Aphrodite subsp. Formosana, and the buds of Cymbidium sinense as material to conduct cryopreservation. In addition, we use viral infected Phalaenopsis Sogo Yukidian’V3’ as material to test and improve the cryopreservation process in order to increase the success rate of virus elimination. The pollen of cucumber “Xiu Yan” was cryopreserved by desiccation dehydration approach. By putting fresh pollen into a glass gar, which contained a fixed number of grams of silica gel; treated with different desiccant dehydration duration and put into LN for a week afterward, and then warmed at 40 ℃ water bath, finally smeared on the germination medium with B & K, pH = 7.0, and 22% sucrose concentration. The results
indicated that after 45 minutes desiccation dehydration approach, the pollen have the best germination rate at about 65.1%, meanwhile the water content was 46.9%. Furthermore, all seeds germinated successfully after pollination properly. The Cymbidium sinense shoot of Taiwan-Yushan and the Cymbidium sinense shoot of Taiwan-Taitung was cryopreserved by vitrification method after high sucrose
medium precultured. The shoots induced by CYS medium from rhizomatous were pre-cultured with 0.4M、0.5M and 0.6M sucrose medium for 0-28 days, and then dealt with LS for 0-180 minutes and PVS2 for 0-240 minutes. After LN treatment for a week, the shoots were warmed with 40℃ water bath, treated with 1.2M sucrose solution for 20 minutes and then grew on the recovery medium. The result showed no survival rate after the experiment. The Phalaenopsis aphrodite subsp. formosana shoot was cryopreserved by vitrification method after high sucrose medium precultured. The shoots were
precultured with 0.4M sucrose medium for 7 days, and then followed with LS and PVS2 treatments for 120 and 240 minutes respectively. A week of LN treatment after, the shoots were warmed with 40℃ water bath, treated with 1.2M sucrose solution for 20 minutes and then grew on the recovery medium. The result showed a 25% survival rate after the experiment. The viral infected Phalaenopsis Sogo Yukidian’V3’ was eliminated virus by chemotherapy: First we sliced the Phalaenopsis shoot apical meristem to 2 mm as
explant, and precultured with the medium which contain virus inhibitor for a month. The result showed a 30.4% virus-free rate after the experiment. In addition, we used chemotherapy coordinated with cryopreservation to eliminate virus: firstly, the Phalaenopsis were cultured with the medium which contain virus inhibitor for a month, precultured with sucrose medium for 5 days, slicing the Phalaenopsis shoot apical meristem about 2 mm as explant to be, and treated with LN lastly. The result showed a 65.2% survival rate and 42.1% virus-free rate. After treating with proper dehydration and stress, most explants reduced the formation of ice crystals during the cryopreservation process, which lowered the degree of cell damage, moreover, the explants could still grow and increase their amount after rewarmed. In addition to this, using cryopreservation to eliminate virus successfully has much contribution on health seedling production and germplasm preservation.
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