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博碩士論文 etd-0813102-102827 詳細資訊
Title page for etd-0813102-102827
論文名稱
Title
石斑神經壞死病毒結合蛋白及免疫法檢測
Immunological detection and the binding protein of Nervous Necrosis Virus infecting Epinephelus malabaricus
系所名稱
Department
畢業學年期
Year, semester
語文別
Language
學位類別
Degree
頁數
Number of pages
100
研究生
Author
指導教授
Advisor
召集委員
Convenor
口試委員
Advisory Committee
口試日期
Date of Exam
2002-06-27
繳交日期
Date of Submission
2002-08-13
關鍵字
Keywords
石斑神經壞死病毒
Nervous Necrosis Virus, Epinephelus malabaricus
統計
Statistics
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中文摘要
石斑腦神經壞死病毒(MGNNV)是屬於Betanodavirus屬的魚類病毒,感染此科的病毒會引起病毒性神經壞死症或病毒性腦及視網膜病變症。本研究所建立的純化MGNNV實驗方法,可得到相當穩定的病毒濃縮液,以方便將來對MGNNV的研究。Betanodavirus可感染SSN-1細胞,判知在細胞中應有能與之結合的病毒接受子。酵素與VBP之關係的實驗中發現,以proteinase K與SSN-1細胞抽出物作用所得到的結合蛋白最少。接受子拮抗劑與VBP之關係的實驗中顯示,利用拮抗劑與接受子結合,可導致病毒無法經由接受子進入細胞進行感染,判斷serotonin 5-HT1A或α2 adrenergic的接受子可作為MGNNV的接受子。
Abstract
Nervous necrosis virus of Epinephelus malabaricus (MGNNV) belongs to the genus of Betanodavirus that causes vacuolating encephalopathy and retinopathy and viral nervous necrosis. A method to purify MGNNV was proposed. The cellular components of SSN-1 reprent few viral receptors for Betanodavirus. Several cellular virus binding proteins (VBPs) were detected by employing the technique of immobilizing virus on nitrocellulose. The least VBPs were found in SSN-1 cell lysate that was treated with proteinase K. The approaches used receptor antagonists to identify the cell receptor. The antagonists are able to block the viral binding and thus potentially directly against the receptor. The results implied that the receptor of serotonin 5-HT1A or α2 adrenergic may act as the receptor of MGNNV.
目次 Table of Contents
目 錄
謝誌 --------------------------------------------------------------------------- Ⅰ
中文摘要 --------------------------------------------------------------------- Ⅱ
Abstract ---------------------------------------------------------------------- Ⅲ
目錄 --------------------------------------------------------------------------- Ⅳ
表目錄 ------------------------------------------------------------------------ Ⅵ
圖目錄 ------------------------------------------------------------------------ Ⅶ
壹、前言
一、 石斑魚 ---------------------------------------------------------------------- 2
二、 台灣養殖石斑魚之現況 ------------------------------------------------- 3
三、 魚類病毒 ------------------------------------------------------------------- 4
四、 Betanodavirus ------------------------------------------------------------- 5
五、 溫度對Betanodavirus感染力的影響 --------------------------------- 7
六、 Nodavirus的基因體 ------------------------------------------------------ 8
七、 Nodavirus複製機制 ------------------------------------------------------ 9
八、 cDNA clones複製產生之病毒亦具有感染力 ----------------------- 10
九、 建立增殖Betanodavirus之細胞株 ------------------------------------ 11
十、 魚類的免疫系統 ---------------------------------------------------------- 14
十一、 MGNNV的結合蛋白 ----------------------------------------------------- 15
十二、 魚類病毒接受子 ---------------------------------------------------------- 16
十三、 關於血清素的接受子 ---------------------------------------------------- 18
十四、 本研究目的 ---------------------------------------------------------------- 21
貳、材料與方法
一、 Total RNA的抽取 -------------------------------------------------------- 22
二、 RNA電泳 ------------------------------------------------------------------ 23
三、 SSN-1細胞株的培養 ----------------------------------------------------- 23
四、 SSN-1增殖NNV ---------------------------------------------------------- 24
五、 從SSN-1細胞株製備病毒液 -------------------------------------------- 25
六、 SDS-PAGE蛋白質電泳 ------------------------------------------------- 25
七、 銀染 ------------------------------------------------------------------------- 26
八、 TCID50定量病毒 ---------------------------------------------------------- 27
九、 超高速離心純化病毒 ---------------------------------------------------- 28
十、 Virus concentrate之蛋白質濃度定量 ---------------------------------- 29
十一、 細胞全液之製備 ---------------------------------------------------------- 29
十二、 Cooking Buffer的製備 --------------------------------------------------- 30
十三、 細胞全液之總蛋白與MGNNV之結合 ------------------------------- 30
十四、 蛋白
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