低溫會導致蝴蝶蘭葉子黃化，影響生長和發育，甚至導致整株植物死亡，影響產值。因此我們利用cDNA篩減方法 (cDNA subtraction) 來選殖與蝴蝶蘭低溫表現相關的基因，從而瞭解蘭花在低溫下的分子機制。
我們將耐寒的蝴蝶蘭TS97及不耐寒的蝴蝶蘭TS365，分別經4℃黑暗處理 (TS97-4℃ & TS365-4℃) 和25℃黑暗處理 (TS97-25℃ & TS365-25℃) 而控制組則不經任何處理 (TS97-L & TS365-L)。兩天之後，取其葉子的total RNA來進行刪減篩選。所篩選的cDNA片段中，發現含有catalase、glyceraldehyde-3-phosphate dehydrogenase、ATP/ADP translocator、及polyubiquitin之部分基因。前人研究指出，低溫會誘導抗氧化酵素的表現，因此我們針對catalase探討其於低溫下是否亦有增加的現象。

以北方印跡法 (Northern blot) 來分析catalase在不同條件處理下的表現量，結果發現TS365 catalase的mRNA在4℃黑暗中會顯著減少，而且在4℃黑暗中，TS365 catalase mRNA之表現量比TS97者少。再以catalase 蛋白質活性分析，亦發現在黑暗中4℃ TS365的活性明顯比TS97者少。雖然catalase在4℃並未被誘導增加，但不耐寒的TS365在4℃下比TS97的表現明顯降低，不耐寒植株catalase在低溫時表現的下降與寒害的形成可能有關。

Phalaenopsis frequently suffer from chilling injury under low temperature. The chilling injury may cause leaf chlorosis and growth retardation. Chilling may also result in plant death and economic loss. To gain more insights of molecular mechanisms of chilling tolerance, genes possibly associate with low temperature condition in Phalaenopsis were isolated using the cDNA subtraction method. Chilling-resistant TS97 and chilling-sensitive TS365 plants were treated with either 4℃/dark (TS97-4℃ & TS365-4℃) or 25℃/dark (TS97-25℃ & TS365-25℃), but control plants were in normal condition (TS97-L & TS365-L). After two days, total RNA was isolated from leaves and used for cDNA subtraction. Partial cDNA fragments of catalase、polyubiquitin glyceraldehyde-3-phosphate dehydrogenase and ATP/ADP translocator genes were obtained. It was reported that some antioxidant enzymes were induced at low temperature. So we focused on the expression of catalase in different conditions.
The catalase mRNA levels were analyzed with Northern blots. We found that the expression of TS365 catalase mRNA was significant reduced at 4℃ in the dark. At 4℃ in the dark TS365 catalase mRNA level was also less than that of TS97. Moreover, the catalase activity in TS365 was lower than that in TS97 at 4℃ in the dark. Although catalase was not induced by low temperaure, a significant reduction was found in the expression of TS365 at 4℃. The reduction of catalase gene expression in TS365 maybe related to chilling injury.

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名詞縮寫-----------------------------------------iv

前言----------------------------------------------1

材料與方法---------------------------------------10

結果---------------------------------------------20

討論---------------------------------------------25

參考文獻-----------------------------------------31

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附錄---------------------------------------------56

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