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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
---|---|---|
dc.contributor.advisor | 闕玲玲(Ling-Ling Chueh) | |
dc.contributor.author | Ya-Ting Kuo | en |
dc.contributor.author | 郭雅婷 | zh_TW |
dc.date.accessioned | 2021-06-16T06:32:05Z | - |
dc.date.available | 2019-08-21 | |
dc.date.copyright | 2014-08-21 | |
dc.date.issued | 2014 | |
dc.date.submitted | 2014-08-06 | |
dc.identifier.citation | 黃瓊儀。犬傳染性呼吸道疾病之病毒核酸診斷與臨床病例分析。碩士論文。國立
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dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/56968 | - |
dc.description.abstract | 犬呼吸道感染常見於犬隻,其感染通常為多病原的,許多病毒及細菌已被報導為可能的病原菌,不同的呼吸道病原於呼吸道感染初期時所造成的臨床症狀相似而造成鑑別診斷上的困難。基因晶片可同時進行多個病原檢測。為了可以特異的敏感的進行犬呼吸道病原的偵測及鑑別,本研究建立了一個針對犬隻呼吸道感染的診斷基因晶片,偵測病原包括犬瘟熱病毒 (Canine distemper virus)、犬流感病毒 (Canine influenzavirus)、犬疱疹病毒 (Canine herpesvirus)、犬黴漿菌 (Mycoplasma cynos) 及支氣管敗血性博得氏菌 (Bordetella bronchiseptica)。此基因晶片除了偵測上述五種病原外,亦同時可進行犬瘟熱病疫苗毒株及野外毒株的區分。針對五種病原設計引子對建立多引子聚合酶鏈鎖反應,將多引子聚合酶鏈鎖反應的產物,基因晶片上依據各標定病原的特定序列設計的寡核苷酸探針進行雜交反應。以肉眼進行結果判讀,可見各病原的寡核苷探針皆能產生清晰且具特異性的雜交訊號。基因晶片於各標定病原的檢測極限約為10 copies到100 copies之間。於本研究所建立之犬呼吸道診斷基因晶片可能有潛力能成為未來的診斷工具之一。 | zh_TW |
dc.description.abstract | Canine respiratory infection is common in dogs worldwide. The infection appears to be multifactorial and a number of viruses and bacteria have been reported as potential pathogens. The clinical signs of early infection caused by different respiratory pathogens are similar and difficult in differentiation. Gene chip could analyze multiple pathogens at a time. To detect and differentiate canine respiratory pathogens with a higher sensitivity and specificity approach, a diagnostic DNA gene chip targeting at canine distemper virus, canine influenza virus, Mycoplasma cynos and Bordetella bronchiseptica was attempted. Besides these five specific pathogens, this assay could differentiate canine distemper virus vaccine strains from field strains. Multiplex RT-PCR using a set of primers for targeting all pathogens was established. The PCR products were then hybridized with the gene chip oligonucleotide probes based on the specific nucleotide sequences. Clear hybridization signals that could detect and differentiate the targeting pathogens were successfully developed and were read by naked eyes. The gene chip detection limits of all target pathogens were about 10 copie to 100 copies. The established gene chip would have the potential to be used as a diagnostic tool in the future. | en |
dc.description.provenance | Made available in DSpace on 2021-06-16T06:32:05Z (GMT). No. of bitstreams: 1 ntu-103-R01629012-1.pdf: 1948032 bytes, checksum: 9993dbb197272b32cbaded1baad84505 (MD5) Previous issue date: 2014 | en |
dc.description.tableofcontents | 誌謝 i
摘要 ii Abstract iii 目錄 v 表目錄 viii 圖目錄 ix 第一章 序言 1 第二章 文獻探討 3 第一節 犬呼吸道感染疾病 3 第二節 犬瘟熱病毒 (Canine distemper virus) 4 2.2.1 犬瘟熱病毒之歷史背景、宿主及流行病學 4 2.2.2 犬瘟熱病毒之病毒特性 5 2.2.3 犬瘟熱病毒之致病機轉與臨床症狀 6 第三節 犬流感病毒 7 2.3.1 犬流感病毒之歷史背景、宿主及流行病學 7 2.3.2 犬流感病毒之病毒特性 8 2.3.3 犬流感病毒之致病機轉與臨床症狀 8 第四節 犬疱疹病毒 9 2.4.1 犬疱疹病毒之歷史背景、宿主及流行病學 9 2.4.2 犬疱疹病毒之病毒特性 9 2.4.3 犬疱疹病毒之致病機轉與臨床症狀 10 第五節 Mycoplasma cynos 11 2.5.1 Mycoplasma cynos之歷史背景、宿主及流行病學 11 2.5.2 Mycoplasma cynos之生物學性狀特徵 12 2.5.3 Mycoplasma cynos之致病機轉與臨床症狀 12 第六節 支氣管敗血性博德氏菌 (Bordetella bronchiseptica) 13 2.6.1 支氣管敗血性博得氏菌之歷史背景、宿主及流行病學 13 2.6.2 支氣管敗血性博得氏菌之細菌特性 13 2.6.3 支氣管敗血性博得氏菌之致病機轉與臨床症狀 13 第三章 材料與方法 15 第一節 實驗設計與流程圖 15 第二節 實驗陽性檢體來源 16 第三節 寡核苷酸引子與探針之選擇與設計 17 3.3.1 寡核苷酸引子之選擇 17 3.3.2 寡核苷酸探針之設計 18 第四節 單引子(反轉錄)聚合酶鏈鎖反應之建立 19 3.4.1 DNA病毒核酸與細菌核酸之萃取 19 3.4.2 RNA病毒核酸之萃取 20 3.4.3 反轉錄反應 (Reverse transcription) 20 3.4.4 單引子聚合酶鏈鎖反應之建立 21 3.4.5 單引子聚合酶鏈鎖反應之專一性分析 22 第五節 聚合酶鏈鎖反應陽性控制組之建立 22 3.5.1 各病原標的基因萃取與純化 22 3.5.2 TA轉殖接合反應 (TA cloning) 23 3.5.3 質體之轉型作用 (Transformation) 23 3.5.4 重組質體之挑選與確認 23 第六節 多引子聚合酶鏈鎖反應之建立 24 第七節 偵測分型基因晶片之建立 25 3.7.1 基因晶片(Gene chip)之置備 25 3.7.2 產物與基因晶片之雜交反應 (Chip Hybridization) 25 3.7.3 偵測分型基因晶片之條件測試 26 3.7.4 多引子聚合酶鏈鎖反應及基因晶片與犬體細胞及鼻腔可能常在病原之測試 26 3.7.5 基因晶片之特異性分析 27 3.7.6 多引子聚合酶鏈產物於電泳及基因晶片之偵測極限 27 3.7.7 基因晶片於臨床檢體及標準品之測試 28 第四章 結果 29 第一節 單引子(反轉錄)聚合酶鏈鎖反應之建立 29 第二節 多引子聚合酶鏈鎖反應之建立 29 第三節 偵測分型基因晶片之建立 30 4.3.1 以基因晶片確認多引子聚合酶鏈鎖反應產物並分型 30 4.3.2 多引子聚合酶鏈鎖反應及基因晶片與犬體細胞及鼻腔可能常在病原之測試 31 4.3.3 多引子聚合酶鏈鎖反應及基因晶片之特異性試驗 31 4.3.4 多引子聚合酶鏈鎖反應之偵測極限 31 4.3.5 基因晶片之偵測極限 32 4.3.6 基因晶片於臨床檢體及標準品之測試 33 第五章 討論 34 附錄-臨床檢體之感染病原確認 54 參考文獻 58 | |
dc.language.iso | zh-TW | |
dc.title | 犬呼吸道病原偵測及分型基因晶片之研發 | zh_TW |
dc.title | The Detection and Differentiation of Canine Respiratory Pathogens Using Gene Chip | en |
dc.type | Thesis | |
dc.date.schoolyear | 102-2 | |
dc.description.degree | 碩士 | |
dc.contributor.coadvisor | 王儷蒨(Lih-Chiann Wang) | |
dc.contributor.oralexamcommittee | 陳健尉,張長泉,張雅君,李雅珍 | |
dc.subject.keyword | 基因晶片,犬呼吸道感染,犬瘟熱病毒,犬流感病毒,犬?疹病毒,支氣管敗血性博得氏菌,犬黴漿菌,多引子聚合?鏈鎖反應, | zh_TW |
dc.subject.keyword | Gene chip,canine respiratory infection,canine distemper virus,canine influenza virus,canine herpesvirus,Mycoplasma cynos,Bordetella bronchiseptica,multiplex PCR, | en |
dc.relation.page | 72 | |
dc.rights.note | 有償授權 | |
dc.date.accepted | 2014-08-06 | |
dc.contributor.author-college | 獸醫專業學院 | zh_TW |
dc.contributor.author-dept | 獸醫學研究所 | zh_TW |
顯示於系所單位: | 獸醫學系 |
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