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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 王錦堂 | |
| dc.contributor.author | Yuan-Hsin Lu | en |
| dc.contributor.author | 呂元馨 | zh_TW |
| dc.date.accessioned | 2021-06-13T07:59:13Z | - |
| dc.date.available | 2005-08-02 | |
| dc.date.copyright | 2005-08-02 | |
| dc.date.issued | 2005 | |
| dc.date.submitted | 2005-07-22 | |
| dc.identifier.citation | 1.Alm R. A., Mattick J. S. 1997. Gene involved in the biogenesis and function of type-4 fimbriae in Pseudomonas aeruginosa. Gene. 192:89-98.
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| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/36391 | - |
| dc.description.abstract | 克雷伯氏肺炎桿菌 (Klebsiella pneumoniae)屬於腸內桿菌科，是一種伺機性感染病原菌。近二十年來，台灣出現一種新型侵襲性克雷伯氏肺炎桿菌感染症，會造成原發性肝膿瘍，有時合併轉移性眼內炎或腦膜炎。比較侵襲性克雷伯氏肺炎桿菌NTUH-K2044及非侵襲性克雷伯氏肺炎桿菌菌株MGH 78578的基因組DNA序列，發現NTUH-K2044的基因組DNA比MGH 78578多出了11個大片段。其中一個片段帶有和纖毛 (fimbriae) 合成相關的基因叢集 (gene cluster)，包括KP0596-KP0601。觀察KP0596-KP0601在臨床分離菌株中的分佈情形，發現此片段在侵襲性菌株中的普遍率比非侵襲性菌株中的普遍率來的高(34/46：1/34)。為了瞭解此片段對於NTUH-K2044致病能力的影響，將其中合成纖毛基座 (usher)的基因KP0600剔除，構築KP0600基因剔除突變株，以胃內感染 (intragastrical infection)的方式感染鼷鼠，結果其50﹪致死劑量 (LD50)和野生株並無差別 (105 CFU)。將KP0596-KP0602纖毛基因叢集送入無纖毛大腸桿菌HB101表現，以穿透式電子顯微鏡觀察並沒有看到纖毛產生。利用西方轉漬法發現感染克雷伯氏肺炎桿菌而造成肝膿瘍的恢復期病人血清中並沒有抗纖毛單元(fimbrial subunit) KP0597抗體的存在。因此認為在KP0596-KP0601基因片段之外，克雷伯氏肺炎桿菌應該有其他的纖毛基因。 | zh_TW |
| dc.description.abstract | Klebsiella pneumoniae is a pathogen of opportunistic infection. Over the past two decades, a new type of invasive K. pneumoniae disease causing primary liver abscess has emerged in Taiwan. Comparing the difference between the genome of tissue-invasive K. pneumoniae NTUH-K2044 and of noninvasive strain MGH-78578, we found that there are eleven large DNA fragments exist in the genome of NTUH-K2044 but not in the genome of MGH-78578. One of the fragments contains a gene cluster including KP0596-KP0601 that may responsible for the expression of fimbriae. We analyzed the distribution of KP0596-KP0601 gene cluster in clinical K. pneumoniae strains. The prevalence rate of KP0596-KP0601 fragment in invasive strains was higher than that in non-invasive strains (34/46 vs. 1/34). To investigate whether the fragment participate in the pathogenicity of NTUH-K2044, we knockout KP0600 gene which encode usher protein. The LD50 of the mutant is the same as that of the wild type strain (105 CFU) in intragastrical inoculation of BALB/c mice. Non-fimbriae Escherichia coli HB101 was transformed with a recombinant plasmid containing fimbrial gene cluster to expression the fimbriae. We did not find the expression of fimbriae with TEM. The purified fimbrial subunit protein was detected by serum of convalescent-phase patients, and there was no anti-subunit antibody in the serum.Therefore, there are probably other gene clusters of fimbriae in NTUH-K2044. | en |
| dc.description.provenance | Made available in DSpace on 2021-06-13T07:59:13Z (GMT). No. of bitstreams: 1 ntu-94-R92445116-1.pdf: 1666793 bytes, checksum: aa1f6d572b11d9d6d314ab9c615c6228 (MD5) Previous issue date: 2005 | en |
| dc.description.tableofcontents | 中文摘要 …………………………………………………………………6
英文摘要 …………………………………………………………………7 第一章 緒論 …………………………………………………………… 8 第二章 材料 ……………………………………………………………14 2.1 克雷伯氏肺炎桿菌菌株 (Klebsiella pneumoniae strains)…14 2.2 大腸桿菌菌株 (Escherichia coli strains) …………………15 2.3 質體 (plasmids) …………………………………………………16 2.4 引子 (primers) …………………………………………………16 2.5 培養基 (medium) …………………………………………………18 2.6 抗生素 (antibiotics) …………………………………………18 2.7 限制酶 (restriction enzyme) …………………………………19 2.8 血清 (sera) ………………………………………………………19 第三章 方法 ……………………………………………………………20 3.1 KP0596-KP0606的生物資訊學分析……………………………… 20 3.2 KP0596-KP0601基因叢集在臨床分離的克雷伯氏肺炎桿菌菌株中的普遍程度 …………………………………………………………………20 3.2.1 聚合酶連鎖反應(polymerase chain reaction) ……………20 3.3 克雷伯氏肺炎桿菌基因剔除突變株的建構 ………………………21 3.3.1克雷伯氏肺炎桿菌基因體DNA的萃取 ……………………………21 3.3.2 TA選殖 (TA cloning) …………………………………………22 3.3.3 KP0600基因剔除突變株的建構 …………………………………23 3.3.4 KP0597基因剔除突變株的建構 …………………………………24 3.3.5 南方轉漬法 (Southern blot) …………………………………25 3.4 動物致病模式 (animal model) …………………………………27 3.5利用穿透式電子顯微鏡觀察纖毛表現 ……………………………28 3.5.1 觀察克雷伯氏肺炎桿菌纖毛的表現 ……………………………28 3.5.2 觀察無纖毛大腸桿菌HB101表現克雷伯氏肺炎桿菌KP0596-KP0602基因叢集 …………………………………………………………28 3.6 KP0597重組蛋白質之表現與純化 …………………………………29 3.6.1 KP0597基因表現載體的建構 ……………………………………29 3.6.2 KP0597蛋白質的表現 ……………………………………………29 3.6.3 KP0597重組蛋白質之純化 ………………………………………30 3.6.4 蛋白質濃度的測定 ………………………………………………32 3.7偵測病人血清中抗KP0597的抗體 …………………………………32 3.7.1 SDS-聚丙烯醯胺膠體電泳 (SDS-PAGE)…………………………32 3.7.2 西方轉漬法 (Western blot) …………………………………33 第四章 結果 ……………………………………………………………34 4.1 KP0596-KP0606的生物資訊學分析 ……………………………… 34 4.2 KP0596-KP0601基因叢集在臨床分離的克雷伯氏肺炎桿菌菌株中的普遍程度 ……………………………………………………………… 35 4.3 克雷伯氏肺炎桿菌基因剔除突變株的建構 ………………………36 4.3.1 KP0600基因剔除突變株的建構 ……………………………… 36 4.3.2 KP0597基因剔除突變株的建構 ……………………………… 37 4.4 動物致病模式 …………………………………………………… 37 4.5 利用穿透式電子顯微鏡觀察纖毛表現 ………………………… 38 4.5.1 觀察克雷伯氏肺炎桿菌纖毛的表現 ………………………… 38 4.5.2 在無纖毛大腸桿菌HB101中表現克雷伯氏肺炎桿菌KP0596- KP0602基因叢集 ……………………………………………………… 38 4.6 病人血清中抗KP0597抗體的偵測 ……………………………… 39 第五章 討論 …………………………………………………………… 40 參考文獻 ……………………………………………………………… 63 | |
| dc.language.iso | zh-TW | |
| dc.subject | 纖毛 | zh_TW |
| dc.subject | 克雷伯氏肺炎桿菌 | zh_TW |
| dc.subject | Klebsiella pneumoniae | en |
| dc.subject | fimbrial | en |
| dc.title | 侵襲性克雷伯氏肺炎桿菌特有纖毛基因叢集之分析 | zh_TW |
| dc.title | Characterization of the fimbrial gene cluster in invasive Klebsiella pneumoniae | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 93-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.oralexamcommittee | 鄧麗珍,賴信志 | |
| dc.subject.keyword | 克雷伯氏肺炎桿菌,纖毛, | zh_TW |
| dc.subject.keyword | Klebsiella pneumoniae,fimbrial, | en |
| dc.relation.page | 69 | |
| dc.rights.note | 有償授權 | |
| dc.date.accepted | 2005-07-22 | |
| dc.contributor.author-college | 醫學院 | zh_TW |
| dc.contributor.author-dept | 微生物學研究所 | zh_TW |
| 顯示於系所單位： | 微生物學科所 | |
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