2007-04-112024-05-17https://scholars.lib.ntu.edu.tw/handle/123456789/684330摘要：台灣商業栽培蘭花品種，由於高溫多溼，在作物栽培、運輸及貯藏時期經常發生細菌性軟腐病危害，造成植株水浸狀、組織柔軟液化及凹陷的情形，引起嚴重損失，故希望利用基因轉殖技術，培育出具抗病能力之蝴蝶蘭品種。另外，為避免基因轉殖作物所帶來環境安全的疑慮，減少基因轉殖蘭花在外銷及內銷市場遭受排斥，以及避免篩選標誌基因對轉殖細胞產生副作用等課題，因此選殖非抗生素耐受篩選標誌 EPSP 合成&#37238;基因及標誌基因去除之 Cre/loxP 序列，搭配目標基因，構築成無標誌通用載體，可應用於文心蘭、蝴蝶蘭等蘭科植物。本年度工作項目包括蝴蝶蘭癒傷組織軟腐病抗性基因 pelE-1 轉殖株之西方轉漬分析、蝴蝶蘭癒傷組織軟腐病抗性基因 pelZ 轉殖株之 GUS 活性組織化學染色、聚合&#37238;連鎖反應及南方氏雜交分析、選殖水稻 EPSP 合成&#37238; (5-enolpyruvylshikimate-3-phosphatesynthase) 基因並經點突變成為篩選標記基因、構築無標誌通用載體。預期本計畫選殖得到的 EPSP 合成&#37238;基因係非抗生素篩選基因，可提供作為非抗生素篩選載體之構築及轉殖；本計畫構築之通用載體，可提供不同目標基因建構無標誌基因之載體，應用於蘭花基因轉殖；轉殖軟腐病抗性基因pelE-1 和 pelZ，期望育出抗軟腐病之蝴蝶蘭品種。<br> Abstract: Bacterial soft rot disease always causes Phalaenopsis severe symptom and reduce the profit. Rresistant gene pectate lyase gene pelE-1 and pelZ isolated from Erwinia chrysanthemi were transformed into PhalaenopelE-1 and pelZ psis via Agrobacteriummediated transformation. The transformants will be confirmed by GUS staining, polymerase chain reaction, Southern analysis and Western analysis. On the other hand, to develop antibiotic-free selectable markers or the ability to excise selectable marker genes becomes important for plant genenetic transformation. The elimination system of selectable marker via Cre/loxP recombinase system will be established in orchid and positive selection marker EPSP synthase gene will be cloned.轉基因植物蝴蝶蘭無篩選標誌系統time細菌性軟腐病transgenic plantPhalaenopsismarker-free systembacterial soft rot