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| 題 名 | 利用聚合酶連鎖反應-限制酶片段長度多型性快速鑑定變形桿菌屬菌種=Polymerase Chain Reaction-restriction Fragment Length Polymorphism for Rapid Identification of Proteus Species |
|---|---|
| 作 者 | 王冠傑; 何明隆; 蔡德君; 彭茹讌; 李明憲; 廖朝財; | 書刊名 | Journal of Biomedical & Laboratory Sciences |
| 卷 期 | 36:3 2024.09[民113.09] |
| 頁 次 | 頁122-132 |
| 分類號 | 415.12 |
| 關鍵詞 | 變形桿菌; recA基因; 聚合酶連鎖反應; 限制酶片段長度多型性; Proteus; recA genes; PCR; Restriction fragment length polymorphism; |
| 語 文 | 中文(Chinese) |
| 中文摘要 | 本研究我們以 recombinase A gene (recA) 為目標基因,開發出 PCR-RFLP 快速鑑定變形桿菌屬菌種之分 子鑑定的方法。首先我們以 GeneBank 中 Proteus mirabilis recA 基因 (accession no. X14870) 全長 1348 bp 的 DNA 進行 BLASTN 比對,BLASTN 比對結果顯示第 328 核苷酸位置的 G 至第 937 核苷酸位置的 A 共 610 個核苷酸區域與許多細菌之 recA 基因有很高的相同性 (identity)。接著我們在此一區域設計了 對變形桿菌屬 recA 基因的退化性引子 (degenerate primers) recA5F 與 recA3R,以此二條退化性引子進 行 PCR 反應可將四種變形桿菌屬菌種之 441 bp recA 基因部分片段增幅。然後我們根據限制酶圖譜分析 之結果,將四種變形桿菌屬菌種之 441 bp recA DNA 利用 TaqⅠ剪切,電泳結果顯示四種變形桿菌屬菌種 之 441 bp recA DNA 利用 TaqⅠ剪切後呈現出不同長度的 DNA 片段的多型性。總而言之,本研究結果顯示 以 441 bp recA DNA 當作目標基因之 PCR-RFLP 方法可快速且正確的區別鑑定四種變形桿菌屬菌種。 |
| 英文摘要 | In this study, we developed a PCR-RFLP method for rapid molecular identification of bacteria within the Proteus genus, targeting the recombinase A gene (recA). Initially, we performed a BLASTN comparison using the full-length 1348 bp DNA sequence of the Proteus mirabilis recA gene (accession no. X14870) obtained from GeneBank. The BLASTN results indicated a high degree of identity between a region spanning from the 328th nucleotide position (G) to the 937th nucleotide position (A), encompassing 610 nucleotides, and the recA genes of many other bacteria. Next, we designed a pair of degenerate primers, recA5F and recA3R, targeting the recA gene region within the Proteus genus. Using these two degenerate primers in a PCR reaction allowed us to amplify a 441 bp partial segment of the recA gene in four different Proteus species. We digested the amplified 441 bp recA PCR products with the restriction enzymes Taq I. The analysis showed that the RFLP profiles of PCR products from each species of Proteus were quite distinguishable. In conclusion, the results of this study demonstrate that the PCR-RFLP method targeting the 441 bp recA DNA can effectively and accurately differentiate and identify the four Proteus species. |
本系統中英文摘要資訊取自各篇刊載內容。