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|標題:||耐寒性相關基因(MF9 及 MF33)轉殖到青花菜及甘藍之研究|
Studies on Genetic Transformation of Broccoli and Cabbage with Chilling-Resistant Related Genes, MF9 and MF33
Plant Gene Transformation
Mefluidide 是一種人工合成的植物生長調節劑，能保護低溫敏感植物，例如：番茄、玉米、胡瓜、及水稻等作物免於寒害。本實驗室自玉米篩選出2個mefluidide誘導的特有選殖系，MF9與MF33。MF9為植物過敏性相關基因，MF33為過氧化酵素基因。本實驗將MF9與MF33基因構築到以CaMV 35S及rbcS啟動子之同義 (sense) 及反義 (antisense) 基因之轉殖載體，並利用農桿菌將此二基因轉殖至甘藍及青花菜。其目的為探討這二個基因在增加植物耐寒性所扮演的角色，並探討藉由基因轉移來增加植物耐寒性或其他逆境之可行性。
將MF9與MF33基因所構築之載體，以農桿菌法將基因轉移至青花菜(綠王)與甘藍(初秋)的下胚軸，並再生青花菜及甘藍之轉殖植株。以PCR及南方墨點分析轉殖植株之結果顯示，MF9與MF33基因已轉移到青花菜及甘藍。北方墨點之分析結果顯示，轉移同義或反義MF9與MF33 基因的再生青花菜及甘藍植株，均有MF9與MF33 RNA的表現。共同轉殖同義與反義MF9或MF33 基因的再生青花菜及甘藍植株之MF9或MF33 RNA的表現量有減少或全無的現象。只有在轉殖同義MF33基因的再生植株才有獨特之過氧化酵素同功異構酵素出現，轉殖反義或共同轉殖同義與反義基因MF33之再生植株則無。轉殖同義MF9或MF33基因之青花菜及甘藍植株可耐低溫處理 (0℃，7天)。轉殖同義MF9或MF33基因之甘藍可耐高溫處理 (42℃，2天)。|
Abstract Mefluidide is a synthetic plant growth regulator, which could protect chilling sensitive plants such as tomato, corn, cucumber and rice form chilling injury. However, the protection mechanism is not clear. A cDNA library was constructed from the poly-A RNA isolated from mefluidide-treated corn leaves. Two mefluidide-induced clones (MF9 and MF33) were selected by subtractive screening. MF9 and MF33 were identified as hypersensitive-related gene and peroxidase gene, respectively, by comparing with the database of Gene Bank. To explore the functions of these two genes in plant, MF9 and MF33 genes driven by CaMV 35S or rbcS promoter in sense and/or anti-sense orientation were transferred into the cabbage and broccoli via Agrobacterium-mediated transformation. Results of PCR and Southern analysis confirmed the MF9 and MF33 genes had been incorporated into the genome of the broccoli and cabbage. High expressions of MF9 and MF33 RNA were found in the transgenic plants transformed with sense or anti-sense MF9 or MF33 genes. Whereas, none or trace of MF9 or MF33 RNA was found in the transgenic plants co-transformed with sense and anti-sense MF9 or MF33 genes. A unique peroxidase isozyme was only found in the transgenic plants transformed with sense MF33 genes. Chilling (0℃, 7 days) as well as heat (42℃, 2 days) resistance were found in the transgenic plants transformed with sense MF9 or MF33 genes, but not in the transgenic plants transformed with anti-sense or co-transformed with sense and anti-sense MF9 or MF33 genes. These results suggest that MF9 and MF33 genes may play an important role in the stress resistance.
|Appears in Collections:||分子生物學研究所|
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