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題 名 | PCR-Mediated Detection of Ustilago Esculenta in Wateroat (Zizania Latifolia) by Ribosomal Internal Transcribed Spacer Sequences=利用內轉錄區間序列及PCR偵測茭白筍中之黑穗病菌 |
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作 者 | 陳瑞祥; 曾德賜; | 書刊名 | 植物病理學會刊 |
卷 期 | 8:4 1999.12[民88.12] |
頁 次 | 頁149-156 |
分類號 | 435.22 |
關鍵詞 | 茭白黑穗病菌; 聚合酵素連鎖反應; 內轉錄區間序列; 核糖體DNA; 偵測; Internal transcribed spacer; ITS; Ribosomal DNA; rDNA; Ustilago esculenta; Wateroat; Zizania latifolia turcz; PCR diagnosis; |
語 文 | 英文(English) |
中文摘要 | 本研究主要目的為篩選對茭白黑穗病菌具有專一性及敏感性之引子對,配合聚合酵素連鎖反應(PCR)增幅技術,開發於田間快速準確偵測茭白黑穗病菌之方法。首先利用相關真菌核糖體DNA研究中所發展所得之通用性引子,自茭白黑穗病菌DNA增幅獲得其核糖體DNA上之內轉錄區間(ITS)序列。經解序反應並與已發表的真菌核糖體DNA序列加以比對後,利用PC-Gene軟體設計偵測用引子對Ue10及Ue11。利用此引子對皆可自測試的16株茭白黑穗病菌DNA中獲得一208bp之增幅產物,而其他常見的植物病原真菌則無類似反應。進一步應用此引子對與茭白植株組織萃取所得DNA反應發現,可於生長點及葉鞘部位偵測到黑穗病菌,但在葉片則未見黑穗病菌的分布;而其偵測靈敏度經測試可達pg程度。本研究結果顯示此引子對的專一性及高靈敏性可實際應用於偵測田間感染茭白植株之黑穗病菌。 |
英文摘要 | PCR-mediated detection of Ustilago esculenta in wateroa (Zizania latifolia Turcu.) tissue have been developed. The internal transcribed spacer (ITS) region of the ribosomal DNA from the representative isolate of U. esculenta was amplified by PCR using conserved ITS primers. The entire ITS region was cloned into a pCRTM II vector, and sequenced. Two PCR primers, Ue10 and Ue11, were designed based on ITS 2 sequences, and tested for the detection of U. esculenta in wateroat tissue. The primer pair was subsequently shown to amplify predicted-size fragments from the DNA of all U. esculenta isolated from wateroats which were collected from the major growth area in Taiwan. On the contrary, this primer pair failed to amplified any fragments from the DNA extracted from other tested pathogenic fungi. Using this primer pair, it was successful to rapidly detect U. esculenta in DNA extracted from as few as 10 mg of diseased wateroat tissue. The amplification products could be deteced in the DNA only from growth tip and steath of diseased plant, but not in the leaves of diseased plants. The detection limit of the primer pair was tested and shown to amplify DNA at the pg level. |
本系統中英文摘要資訊取自各篇刊載內容。