利用扣減法與基因微陣列技術篩選文心蘭假球莖中花序發育初期專一表現之基因

Abstract

文心蘭開花期的決定似乎與其營養狀態有相當大的關聯；而假球莖是文心蘭貯藏水分及養分的主要器官，本論文即以扣減式基因庫結合cDNA微陣列晶片技術，以二種不同發育階段：A時期，花軸2?3公分；及B時期，花軸7?8公分之文心蘭假球莖為材料進行扣減式篩選，期望能夠得到影響開花時間之基因。在A減B扣減式基因庫中，獲得了45個在A時期表現倍數大於B時期2倍的cDNA片段序列，在比對已知蛋白質序列的結果顯示，11個基因產物與光合作用相關，另外11個基因可能參與了蛋白質的合成分解及運送，其他還包括了似植物乳膠蛋白（latex-like proteins）、過氧化物?（peroxidases）、甘露糖結合活性的植物凝集素（mannose-binding lectins）等。本論文並對其中之一的過氧化物脢，在分類上屬於植物第三群（class III），初步探討其在開花過程中所扮演的角色。

The pseudobulb of Oncidium Gower Ramsey is the primary storage organ for water and nutrients preservation. The determination of flower trigger of Oncidium seems to have a strong relationship with the nutritional status of pseudobulbs. In order to obtain the genes specifically involved in the determination of flower trigger of Oncidium, we used PCR-select cDNA subtraction and cDNA microarray techniques to screen genes differentially expressed between two different developmental stages of pseudobulbs. Stage A represented the pseudobulbs with floral bud of 2?3 cm length, and stage B represented the pseudobulbs with floral bud of 7?8 cm length. We obtained 45 cDNA clones that expressed two folds higher in the A stage than those in the B stage, after screening the A minus B subtraction library. There are 11 clones found to be involved in photosynthesis process, and another 11 clones were found to be involved in protein synthesis and transportation. The others are latex-like proteins, peroxidases and mannose-binding lectins. On the other hand, we study further one of the peroxidases gene, OPOXl, which belongs to plant class III peroxidase. The gene expressions of OPOXl in Oncidium during different developmental stages were investigated, and the role of OPOXl involved in the flowering process has been discussed.