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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
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dc.contributor.advisor | 蕭信宏 | |
dc.contributor.author | Yi-Shan Wu | en |
dc.contributor.author | 吳依珊 | zh_TW |
dc.date.accessioned | 2021-06-17T08:22:52Z | - |
dc.date.available | 2024-08-27 | |
dc.date.copyright | 2019-08-27 | |
dc.date.issued | 2019 | |
dc.date.submitted | 2019-08-13 | |
dc.identifier.citation | Anders RF, Adda CG, Foley M, and Norton RS. Recombinant protein vaccines against the asexual blood stages of Plasmodium falciparum. Hum Vaccin. 2010; 6:39-53.
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dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/74172 | - |
dc.description.abstract | 蚊子是許多疾病的傳播媒介,例如:瘧疾、登革熱,茲卡、屈躬病、日本腦炎、西尼羅河熱以及黃熱病等。其中,登革熱為目前重要的疾病之一,根據世界衛生組織統計,全球每年大約有3億9千萬的感染病例,且當中約9千萬人具有嚴重臨床症狀。而在2014至2015年間台灣共有超過5萬件的感染案例,但由於目前仍無有效的治療藥物上市,因此,如何預防病媒蚊叮咬以及控制病媒蚊的數量,是目前防治蚊媒傳染疾病最重要的研究課題。根據先前哺乳類研究文獻指出,第二型登革病毒 (dengue virus serotype 2, DENV2)中的非結構性蛋白5 (non-structural protein 5, NS5)在感染宿主細胞的過程中,會進入宿主細胞的細胞核內,且會對病毒的感染力產生影響,因此,本研究以埃及斑蚊為研究模式,探討NS5蛋白入核對登革病毒在埃及斑蚊體內複製之重要性。首先,我們利用免疫螢光染色法分析登革病毒第一至四型的NS5蛋白入核狀況,結果顯示只有第二型登革病毒的NS5蛋白在埃及斑蚊體內會進入細胞核中。接著,我們利用可抑制NS5蛋白入核的抑制劑Ivermectin來處理埃及斑蚊,發現經Ivermectin處理的組別,登革病毒的複製能力及感染力皆有顯著下降。除此之外,先前研究顯示NS5蛋白入核會影響messenger RNA的剪接,而在本研究中也發現NS5蛋白入核會影響免疫相關基因Caspar的剪接。未來我們將更進一步探討NS5蛋白入核對於影響登革病毒複製能力的詳細機制,以及探討與Caspar相關的免疫調控機制。 | zh_TW |
dc.description.abstract | Mosquitoes are one of the fatal animals in the world and they act as vectors for several diseases, including malaria, dengue fever, Zika, japanese encephalitis, West Nile fever, and chikungunya. Among these diseases, dengue fever is currently one of the world’s most important tropical diseases. Previous studies have indicated that the non-structural protein 5 (NS5) was able to translocate into the nucleus and influence the dengue virus serotype 2 (DENV2) infectivity in mammalian cell lines. However, the detail mechanisms of NS5-mediated infectivity remain largely unknown. Therefore, the aim of this study is to investigate the role of the dengue virus NS5 nuclear localization on DENV replication in the mosquito Aedes aegypti. We showed that dengue virus NS5 was localized in the nucleus only in the case of DENV2 (16681 strain) infection in Aedes aegypti. Next, we investigated the effect of NS5 nuclear localization in viral replication by application of Ivermectin, which was demonstrated to be an inhibitor for NS5 protein nuclear localization. We showed that treatment of Ivermectin resulted in the significant reduction of DENV2 replication and infectivity. We also showed that Caspar splicing was inhibited in the case of NS5 nuclear localization, suggesting the important role of NS5 nuclear localization on host mRNA splicing. In the future, we will further investigate the detail mechanisms underlying the regulation of NS5 nuclear localization on DENV2 replication and immune response modulation. | en |
dc.description.provenance | Made available in DSpace on 2021-06-17T08:22:52Z (GMT). No. of bitstreams: 1 ntu-108-R06445202-1.pdf: 2548049 bytes, checksum: 9781c906ff3f1778ecc932d2d48a408b (MD5) Previous issue date: 2019 | en |
dc.description.tableofcontents | 誌謝 i
中文摘要 ii Abstract iii 圖目錄 vii 第一章 緒論 1 1.1. 病媒蚊傳播疾病(mosquito-borne disease) 1 1.1.1. 瘧疾 (malaria) 3 1.1.2. 茲卡病毒感染症 (Zika virus infection) 3 1.2. 登革熱 (Dengue fever) 4 1.2.1. 登革病毒 (Dengue virus;DENV) 4 1.2.2. 臨床症狀 4 1.2.3. 台灣之登革熱流行病學 5 1.3. 埃及斑蚊生活史 6 1.4. 登革病毒生活史 6 1.4.1. 宿主細胞內之複製 6 1.4.2. 病媒蚊體內之複製 7 1.5. 非結構性蛋白5 (Non-structure protein 5, NS5) 7 1.5.1. NS5蛋白之功能 7 1.5.2. NS5蛋白之入核 8 1.6. 實驗動機與假說 9 第二章 實驗材料與方法 10 2.1. 實驗步驟與流程 10 2.1.1. 埃及斑蚊飼養與繼代 10 2.1.2. 細胞培養 (cell ulture) 10 2.1.3. 病毒製備 11 2.1.4. 免疫螢光染色實驗 (Immunofluorescent assay;IFA) 11 2.1.5. 西方點墨法 (western blotting) 13 2.1.6. 雙股RNA (double-stranded RNA,dsRNA)製備 14 2.1.7. RNA萃取 (RNA extraction) 15 2.1.8. 反轉錄作用 (Reverse transcription,RT) 15 2.1.9. 聚合酶連鎖反應 (Polymerase chain reaction,PCR) 16 2.1.10. 即時定量聚合酶連鎖反應 (Real-time PCR,Quantitative PCR) 16 2.1.11. 顯微注射 (microinjection) 17 2.1.12. 溶班試驗 (plaque assay) 17 2.1.13. 埃及斑蚊之病毒血感染 (oral infection) 18 2.2. 實驗試劑製備 18 第三章 結果 22 3.1. 登革病毒之NS5蛋白入核 22 3.2. 16681病毒株與NGC病毒株入核相關之序列差異 22 3.3. 16681病毒株及NGC病毒株在細胞內外複製之比較 23 3.4. 16681病毒株與NGC病毒株之E病毒蛋白胺基酸序列比對 23 3.5. 16681病毒株與NGC病毒株之生長曲線比較 24 3.6. 顯微注射感染之16681病毒株與NGC病毒株在埃及斑蚊體內複製差異 24 3.7. NS5蛋白入核之重要性 25 3.8. 埃及斑蚊感染登革病毒後各組織內入核蛋白β (Importin β)之表現情形 26 3.9. 入核蛋白β為登革病毒在埃及斑蚊體內複製時所需之重要蛋白 27 3.10. NS5蛋白入核與Caspar之切割具相關性 27 3.11. 第二型登革病毒16681病毒株與NGC病毒株影響Caspar切割之差異 28 3.12. 病毒血感染之第二型登革病毒16681病毒株與NGC病毒株在埃及斑蚊體內複製差異與下游基因的表現量 28 3.13. 感染登革病毒之埃及斑蚊體內受Caspar調控之antimicrobiol peptide表現量 29 第四章 討論 30 4.1. 登革病毒之NS5蛋白表現位置 30 4.2. DENV2 16681 病毒株及NGC病毒株之差異 30 4.3. 抑制NS5蛋白入核對DENV2之影響 32 4.4. NS5蛋白入核對下游基因Caspar之影響 33 附圖 35 附表 59 表 一 實驗所使用之PCR引子序列 59 表 二 各實驗所使用之抗體與操作條件 60 附錄 62 參考文獻 65 | |
dc.language.iso | zh-TW | |
dc.title | 探討NS5蛋白入核對登革病毒在埃及斑蚊體內複製之重要性 | zh_TW |
dc.title | Dengue virus NS5 protein nuclear localization is crucial for virus replication in Aedes aegypti | en |
dc.type | Thesis | |
dc.date.schoolyear | 107-2 | |
dc.description.degree | 碩士 | |
dc.contributor.oralexamcommittee | 余明俊,顧家綺 | |
dc.subject.keyword | 埃及斑蚊,登革病毒,入核,免疫調控,病毒複製, | zh_TW |
dc.subject.keyword | Aedes aegypti,dengue virus,nuclear localization,immune regulation,virus replication, | en |
dc.relation.page | 70 | |
dc.identifier.doi | 10.6342/NTU201903186 | |
dc.rights.note | 有償授權 | |
dc.date.accepted | 2019-08-14 | |
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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