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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
---|---|---|
dc.contributor.advisor | 蕭信宏 | |
dc.contributor.author | I-Yi Chen | en |
dc.contributor.author | 陳依沂 | zh_TW |
dc.date.accessioned | 2021-06-16T10:50:26Z | - |
dc.date.available | 2018-09-24 | |
dc.date.copyright | 2013-09-24 | |
dc.date.issued | 2013 | |
dc.date.submitted | 2013-08-12 | |
dc.identifier.citation | Anders, R. F., Adda, C. G., Foley, M., and Norton, R. S. (2010). Recombinant protein vaccines against the asexual blood stages of Plasmodium falciparum. Hum Vaccin, 6(1), 39-53.
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Sustained Notch signaling in progenitors is required for sequential emergence of distinct cell lineages during organogenesis. Genes Dev, 20(19), 2739-2753. Zou, Z., Shin, S. W., Alvarez, K. S., Kokoza, V., and Raikhel, A. S. (2010). Distinct melanization pathways in the mosquito Aedes aegypti. Immunity, 32(1), 41-53. | |
dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/61165 | - |
dc.description.abstract | 蚊子是許多重要傳染疾病的媒介,例如瘧疾、登革熱、黃熱病、西尼羅熱、絲蟲病等。其中只有黃熱病具有效疫苗,而現有的抗瘧疾藥物也開始出現抗藥性的問題,因此,控制病媒蚊的數量是抑制疾病傳播的一個替代性策略。我們希望透過研究埃及斑蚊產卵及發育的相關機制來找出控制病媒蚊族群的可能性。在蚊子的生活史中,蚊卵產出後需要經過一段時間卵殼才會黑化硬化,此時的卵才能夠保護其內的胚胎。有趣的是,我們在先前的實驗中發現,當我們以RNAi的方式抑制埃及斑蚊Notch的表現,會造成部分的卵無法黑化,而且不論這些卵有無黑化,絕大部分的卵皆無法順利孵化。過去的研究證實蚊子體內的黑化作用主要參與先天免疫反應、傷口癒合、表皮硬化以及卵殼黑化,而參與黑化反應的酵素包含原酚氧化酵素(prophenoloxidase, PPO)、多巴脫羧酵素(dopa decarboxylase, DDC)、多巴色素轉換酵素(dopachrome conversion enzyme, DCE)等,但關於黑化反應與卵殼黑化的研究並不多。此外在哺乳類動物及果蠅的研究也發現Notch是一個高度保留的訊息傳遞路徑,在胚胎發育與決定細胞分化扮演重要的角色。因此我們在本研究中想深入探討埃及斑蚊卵殼黑化的調控機制,以及卵殼黑化反應與Notch的關係。我們利用RNAi的方式分別抑制了原酚氧化酵素與多巴脫羧酵素,結果顯示當多巴脫羧酵素被抑制後可見卵殼黑化明顯受到抑制,而不論我們抑制單一或多個原酚氧化酵素,結果卻發現對卵殼黑化無明顯影響,但若我們使用抑制劑來抑制酚氧化酵素,結果顯示卵殼黑化也受抑制;當我們抑制Notch時,結果顯示原酚氧化酵素之RNA表現與酵素活性皆下降,但多巴脫羧酵素之RNA與蛋白質表現並不受影響,故我們認為Notch透過調控原酚氧化酵素的表現來調控蚊卵卵殼黑化。本研究結果將有助於未來應用於病媒傳病能力之研究及病媒控制策略之制訂。 | zh_TW |
dc.description.abstract | Melanization is a prominent mechanism for insect physiology, such as innate immune response, cuticular sclerotization, wound healing and egg tanning. Egg tanning, which resulted in the hardening of eggshell, is pivotal for the protection of developing embryos from harsh environmental conditions. It has been demonstrated that prophenoloxidase (PPO) and dopa decarboxylase (DDC), both are important components of PPO cascade, are critical for the regulation of melanization. On the other hand, Notch signaling was shown to be a highly conserved signaling pathway and required for embryonic development. Our previous results revealed that silencing of Aedes aegypti Notch by using RNAi-mediated technique resulted in the significant inhibition of egg tanning. Therefore, we attempt to investigate the potential regulatory machinery between Notch-mediated egg tanning and prophenoloxidase cascade. First, we examine the effect of PPOs and DDC in the egg tanning. Our results showed that silencing of DDC resulted in the inhibition of egg tanning in the mosquito Aedes aegypti, whereas silencing of individual PPO or grouped-PPOs had no effect on egg tanning. Injection of a PO inhibitor into female mosquitoes after a blood meal resulted in the inhibition of egg tanning. RT-PCR analysis and PO activity assay both showed that significant decreases in the PPOs expression and the PO activity were exhibited in the absence of Notch. On the contrary, silencing of Notch had no effect on DDC expression by means of RT-PCR and western blot analysis. Therefore these results suggested an important role of PPOs in the Notch-mediated egg tanning. Data revealed by this study will be crucial for future studies on vectors competence and vector control in the field. | en |
dc.description.provenance | Made available in DSpace on 2021-06-16T10:50:26Z (GMT). No. of bitstreams: 1 ntu-102-R00445201-1.pdf: 4700094 bytes, checksum: d2586aa624f579d24ae833b278f07c3c (MD5) Previous issue date: 2013 | en |
dc.description.tableofcontents | 誌謝 i
中文摘要 ii Abstract iii 目錄 iv 圖目錄 vii 表目錄 viii 第一章 緒論 1 1.1. 蚊子傳播的疾病 1 1.1.1. 瘧疾(Malaria) 2 1.1.2. 登革熱(Dengue fever) 3 1.2. 埃及斑蚊之生活史 3 1.3. 埃及斑蚊卵黃生成機制(Vitellogenesis) 4 1.4. 黑化反應(Melanization) 5 1.5. Notch signaling pathway 7 1.6. 實驗動機與假說 8 第二章 實驗材料與方法 9 2.1. 實驗步驟與流程 9 2.1.1. 埃及斑蚊之飼養與餵血 9 2.1.1.1. 飼養與繼代 9 2.1.1.2. 餵血 9 2.1.2. RNA萃取(RNA extraction) 9 2.1.3. 反轉錄作用(Reverse transcription, RT) 10 2.1.4. 聚合酶連鎖反應(Polymerase chain reaction, PCR) 10 2.1.5. 即時定量聚合酶連鎖反應(Real-time PCR, Quantitative PCR) 10 2.1.6. 原位雜交(In situ hybridization) 11 2.1.6.1. 質體建構(Plasmid construction) 11 2.1.6.2. RNA探針(probe)之製備 12 2.1.6.3. Whole mount in situ hybridization 12 2.1.7. Double-stranded RNA (dsRNA)製備 13 2.1.7.1. 質體建構(Plasmid construction) 13 2.1.7.2. dsRNA合成(dsRNA synthesis) 14 2.1.8. RNAi-mediated silencing與knockdown效率測試 14 2.1.9. 抑制劑注射(Inhibitors injection) 15 2.1.10. Phenoloxidase活性試驗(PO activity assay) 15 2.1.11. 西方點墨法(Western blot) 15 2.1.12. 孵化率試驗(Hatching assay) 16 2.1.13. 掃描式電子顯微鏡 17 2.2. 實驗試劑之製備 17 第三章 結果 20 3.1. 埃及斑蚊Notch mRNA於卵泡(follicle)中之分佈情形 20 3.2. 埃及斑蚊PPO胺基酸序列之比對 20 3.3. 埃及斑蚊PPO於各發育時期之mRNA表現情形 20 3.4. RNAi抑制埃及斑蚊PPO對卵殼黑化之影響 21 3.5. PTU對埃及斑蚊卵殼黑化之影響 22 3.6. RNAi抑制埃及斑蚊Notch對PPO mRNA表現量之影響 22 3.7. RNAi抑制埃及斑蚊Notch對PO activity之影響 22 3.8. 埃及斑蚊DDC於各發育時期之mRNA表現情形 23 3.9. 埃及斑蚊DDC於成蚊時期之蛋白質表現情形 23 3.10. RNAi抑制埃及斑蚊DDC對卵殼黑化之影響 24 3.11. RNAi抑制埃及斑蚊DDC對卵殼表面構造之影響 24 3.12. RNAi抑制埃及斑蚊DDC對蚊卵孵化之影響 25 3.13. RNAi抑制埃及斑蚊DDC對蚊卵受精之影響 25 3.14. RNAi抑制埃及斑蚊Notch對DDC mRNA表現量之影響 26 3.15. RNAi抑制埃及斑蚊Notch對DDC蛋白質表現之影響 26 第四章 討論 27 附圖 32 附表 54 參考文獻 58 | |
dc.language.iso | zh-TW | |
dc.title | Notch與原酚氧化酵素連鎖反應調控埃及斑蚊卵殼黑化作用之探討 | zh_TW |
dc.title | Notch-mediated egg tanning is co-operated with prophenoloxidase cascade in the mosquito Aedes aegypti | en |
dc.type | Thesis | |
dc.date.schoolyear | 101-2 | |
dc.description.degree | 碩士 | |
dc.contributor.oralexamcommittee | 吳君泰,詹智強 | |
dc.subject.keyword | 埃及斑蚊,Notch,卵殼黑化,原酚氧化酵素,多巴脫羧酵素, | zh_TW |
dc.subject.keyword | Aedes aegypti,Notch,egg tanning,prophenoloxidase,dopa decarboxylase, | en |
dc.relation.page | 63 | |
dc.rights.note | 有償授權 | |
dc.date.accepted | 2013-08-12 | |
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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