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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 張繼堯(Chi-Yao Chang) | |
| dc.contributor.author | Bin-Hong Hsu | en |
| dc.contributor.author | 徐賓鴻 | zh_TW |
| dc.date.accessioned | 2021-06-16T17:15:19Z | - |
| dc.date.available | 2015-08-20 | |
| dc.date.copyright | 2012-08-20 | |
| dc.date.issued | 2012 | |
| dc.date.submitted | 2012-08-17 | |
| dc.identifier.citation | 參考文獻
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(2005) Human ISG15 conjugation targets both IFN-induced and constitutively expressed proteins functioning in diverse cellular pathways. National Academy of Sciences 102: 10200-10205. | |
| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/63631 | - |
| dc.description.abstract | 石斑魚虹彩病毒─可感染石斑魚生活史各個階段，魚隻罹病後容易死亡，常導致石斑魚育成率降低，並造成經濟上的損失。對於虹彩病毒的防範，開發一個有保護力的疫苗，並對疫苗效力的研究與評估是各方所關注的重點。過去，在評估疫苗的效力，通常僅能鎖定專一性抗體 (specific antibody) 的效價高低與持續的時間，並搭配活毒攻毒後的相對存活率 (relative percent survival, RPS) 做為評估依據。但對於病毒與宿主間交互作用的基礎研究仍然缺乏。本實驗室先前已藉由研究微陣列晶片 (microarray) 的結果，篩選出虹彩病毒感染石斑魚後，38個具有兩倍以上正調控，48個兩倍以上負調控的基因群，並進一步以RT-qPCR確認感染虹彩病毒後不同時間點，魚隻的頭腎、脾臟基因表現變化，共分析出30個虹彩病毒反應基因 (virus responsive genes, VRGs) ，參與宿主先天性免疫和後天性免疫反應。
本研究以福馬林不活化 GIV 疫苗，對於點帶石斑魚苗進行腹腔免疫注射後，再進行虹彩病毒活毒攻毒測試。分析相關的15個 VRGs 在病毒攻毒後的表現變化量。其中 DHX58、HECT、Mx、Viperin、ISG15、PLAUR、CD9、VLIG1 等基因表現量在感染過後第一天到第三天的頭腎及脾臟中有上升趨勢，在第七天之後逐漸下降，這些免疫基因的調控與先天性免疫系統中的干擾素傳遞路徑非常符合。此外在本研究所分析的基因之中，攻毒過後大多數的基因表現量在疫苗注射組是低於控制組的，這樣的現象與魚體內 GIV MCP (major capsid protein) 的表現量有正比的關係，顯示經疫苗免疫後的魚隻確實產生專一性抗體，可以中和 (neutralize) 病毒減低病毒量。攻毒後 30 天的觀察結果，在免疫注射組的存活率 100 %，相較於控制組的 0 %，達到 100 % 的相對存活率 (relative percent survival, RPS) ，證實所施打的疫苗確實具有保護效力。 在施打兩種不同劑量的疫苗過後，魚血清內專一性抗體 (specific antibody) 的追蹤實驗，也發現無論在高或低劑量免疫注射組別之中，抗體於一周後就攀升到最高，持續到第八周有些微下降之趨勢，而專一性抗體生成的量對於注射進去魚體內的疫苗有劑量正比 (dose-dependent) 關係。 | zh_TW |
| dc.description.abstract | Grouper iridovirus (GIV) can infect all life stages in grouper and cause serious death. It also make low survival rate in grouper from fry to juvenile stage, and may cause aquaculture economic damage. To develop a high efficacy vaccine is important to resist iridovirus . Calculating the relative percent survival (RPS) rate post challenge and detecting the titer of specific antibody and its maintain efficiency, are used to estimate the efficacy of vaccine in the past. But the interaction between host and iridovirus is not clear completely. According microarray results in GIV infection orange-spotted grouper (Epinephelus coioides) , we selected many candidate gene in our lab previously. Thus, we have confirmed and analyzed 30 virus responsive genes (VRGs) in response to innate immunity and adaptive immunity in grouper’s head kidney and spleen after GIV infection. In this study, we analyzed the expression of 15 VRGs in grouper following vaccination against GIV infection by intraperitoneal injection. We discovered that DHX58, HECT, Mx, Viperin, ISG15, PLAUR, CD9 and VLIG1 expression patterns are very similar, and these genes are considered to involving in interferon regulated pathway in innate immunity. On the other hand, most VRGs expression are lower in vaccination group than in control after GIV challenge. The maximum anti-GIV specific antibody is produced in fish sera post vaccination one week, even in the group of low dose vaccine treatment. The ELISA results showed that host immune response by vaccination exist dose-dependent relationship. These results indicated that the neutralization between ant-GIV specific antibody and GIV caused viral load reducing in the vaccinated group after GIV challenging. | en |
| dc.description.provenance | Made available in DSpace on 2021-06-16T17:15:19Z (GMT). No. of bitstreams: 1 ntu-101-R99b45018-1.pdf: 1452411 bytes, checksum: 64b3be165e63b4dc48cb9454aca2c232 (MD5) Previous issue date: 2012 | en |
| dc.description.tableofcontents | 目錄
中文摘要 I Abstract III 目錄 V 表目錄 VIII 圖目錄 IX 第一章 前言 1 1.1 石斑魚之簡介與台灣養殖現況 1 1.2 魚類虹彩病毒之相關研究 2 1.3 魚類免疫系統 7 1.4 水產疫苗之介紹 8 1.4.1 不活化疫苗 (Inactivated vaccine) 8 1.4.2 活的弱毒化疫苗 (Live-attenuated vaccine) 9 1.4.3 次單位疫苗 (subunit vaccine) 9 1.4.4 DNA疫苗 (DNA vaccine) 10 1.5 即時定量聚合酶連鎖反應 (Quantitative Real-Time Polymerase Reaction) 11 1.6 石斑魚免疫基因 11 1.7 研究動機 16 第二章 材料與方法 18 2.1 細胞培養與繼代 18 2.2 石斑魚虹彩病毒增殖與力價測定 18 2.2.1 石斑魚虹彩病毒增殖 19 2.2.2 石斑魚虹彩病毒力價測定 19 2.3 虹彩病毒不活化疫苗製備 19 2.4 點帶石斑魚苗蓄養條件 20 2.5 點帶石斑魚免疫注射及攻毒實驗 20 2.5.1 實驗魚隻GIV之檢測 21 2.5.2 免疫腹腔注射與攻毒實驗 21 2.6 RNA組織萃取與純化 23 2.7 cDNA 之合成 24 2.8 以即時定量聚合酶連鎖反應分析基因表現量 24 2.8.1 即時定量聚合酶連鎖反應之引子對設計 24 2.8.2 即時定量聚合酶連鎖反應 25 2.8.3 基因表現之分析 25 2.9 疫苗專一性抗體效價分析 26 2.9.1 專一性抗體免疫力分析實驗 26 2.9.2 GIV之純化 27 2.9.3 病毒蛋白質之定量 28 2.9.3 不活化疫苗接種後專一性抗體力價分析 28 第三章 結果 30 3.1 實驗魚隻GIV檢測結果 30 3.2 疫苗安全性評估 30 3.3 攻毒測試後之累計死亡率結果 30 3.4 高劑量免疫組在攻毒後不同時間點病毒表現量分析 31 3.5 高劑量疫苗免疫注射─攻毒後不同時間點基因表現量之分析 31 3.6 不活化疫苗接種後專一性抗體力價分析 32 第四章 討論 33 參考文獻 37 表目錄 表一. 魚苗腹腔注射疫苗後攻毒前安全性評估 45 表二. 引子對序列 46 圖目錄 圖一. 實驗魚隻GIV檢測。 48 圖二. 低劑量不活化疫苗免疫後三周攻毒結果。 49 圖三. 低劑量不活化疫苗追加注射後攻毒結果。 50 圖四. 高劑量不活化疫苗免疫27天後攻毒結果。 51 圖五. 高劑量疫苗免疫27天後攻毒不同時間點病毒表現量。 52 圖六. 高劑量疫苗免疫組攻毒後不同時間點基因表現變化量。 55 圖七. 不活化疫苗接種後專一性抗體力價分析。 56 | |
| dc.language.iso | zh-TW | |
| dc.title | 點帶石斑魚經疫苗接種後其對抗虹彩病毒感染
之基因表現與免疫反應 | zh_TW |
| dc.title | Genes expression and immune responses in orange spotted grouper (Epinephelus coioides) following vaccination against grouper iridovirus (GIV) infection | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 100-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.oralexamcommittee | 林正輝,蘇建國 | |
| dc.subject.keyword | 點帶石斑,虹彩病毒,疫苗, | zh_TW |
| dc.subject.keyword | orange spotted grouper,iridovirus,vaccine, | en |
| dc.relation.page | 56 | |
| dc.rights.note | 有償授權 | |
| dc.date.accepted | 2012-08-20 | |
| dc.contributor.author-college | 生命科學院 | zh_TW |
| dc.contributor.author-dept | 漁業科學研究所 | zh_TW |
| 顯示於系所單位： | 漁業科學研究所 | |
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