頁籤選單縮合
題 名 | 建立馬鈴薯多重聚合酶連鎖反應檢測技術=Establishment and Application of Multiplex Polymerase Chain Reaction for Simultaneous Detection of Transgenic Potatoes |
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作 者 | 杜元凱; 馮昱維; 陳烈夫; 陳述; 林彥君; 陳涵葳; | 書刊名 | 臺灣農業研究 |
卷 期 | 68:1 2019.03[民108.03] |
頁 次 | 頁16-27 |
分類號 | 434.32 |
關鍵詞 | 馬鈴薯; 多重聚合酶連鎖反應; 檢驗方法; Potato; mPCR; Detecting method; |
語 文 | 中文(Chinese) |
中文摘要 | 馬鈴薯(Solanum tuberosum L.)為我國常見的食用及加工作物,其塊莖在工業和飼料上具有很高的應用價值。本試驗以馬鈴薯物種專一性(taxon-specific)內生性基因尿苷二磷酸葡萄糖焦磷酸化酶(UDP-glucose pyrophosphorylase; UGPase)序列與基因轉殖馬鈴薯品項之專一性序列(event-specific sequence)建立多重聚合酶連鎖反應(multiplex polymerase chain reaction; mPCR)檢測技術。結果顯示,經多重聚合酶連鎖反應可正確增幅出目標片段,且混合2或3個不同轉殖品項之DNA樣本,亦能檢出專一性目標片段;單一基因轉殖品項之稀釋樣本檢測其最低檢測極限(limit of detection; LOD),本方法之檢測靈敏度可達0.5%。將本方法送至4個不同的試驗單位進行能力試驗,結果亦顯示具備良好穩定性與再顯性。另外,本試驗以快速馬鈴薯核酸萃取方法並利用毛細管電泳分析目標片段,可大幅縮減檢測流程所耗時間、人力成本,期望能應用於馬鈴薯檢測技術開發與基因改造(基改)馬鈴薯轉殖品項檢驗方法之參考。 |
英文摘要 | Potato (Solanum tuberosum L.) is known as one of the most important food crops in the world and has high application in industry and feed. In this study, we present a multiplex polymerase chain reaction (mPCR) method for simultaneous detection of genetically modified (GM) potatoes. Quadruplex PCR assay targeting three event-specific sequences and the taxon-specific endogenous reference gene (UDP-glucose pyrophosphorylase; UGPase) were established. All potato DNA samples generated the expected PCR products, and the detection method performed robustly and specifically with various combinations of mixed GM potato events. The limit of detection (LOD) of each single GM sample in our mPCR detection assay was 0.5%. Furthermore, a proficiency test returned from four research sectors also indicated that our detection method had good reproducibility and repeatability. We also developed a simple, rapid and cost-efficient nucleic acid extraction method coupled with multiplex PCR and followed capillary electrophoresis. These results suggest that our methods are suitable for detecting multiplex PCR targets and have the potential for screening unknown genetically modified potato samples through a simple procedure. |
本系統中英文摘要資訊取自各篇刊載內容。