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研究生:李彥君
研究生(外文):Yann-Jium Lee
論文名稱:日本鯷的微隨體基因座及系群判別
論文名稱(外文):Microsatellite DNA and stock identification in Japanese anchovy (Engrualis japonica)
指導教授:丘臺生丘臺生引用關係于宏燦于宏燦引用關係
指導教授(外文):Tai-Sheng ChiuAlex Hon-Tsen Yu
學位類別:碩士
校院名稱:國立臺灣大學
系所名稱:動物學研究所
學門:生命科學學門
學類:生物學類
論文種類:學術論文
論文出版年:2000
畢業學年度:88
語文別:中文
論文頁數:58
中文關鍵詞:日本鯷微隨體
外文關鍵詞:Engraulis japonicamicrosatellite
相關次數:
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摘要
日本鯷(Engraulis japonica)為一經濟性魚種,廣泛分佈於北太平洋西岸的溫帶至熱帶間。成魚於春季時會洄游至台灣東北海域,以及台灣海峽大部份海域。本研究即以日本鯷為對象,進行微隨體基因座的選殖工作,並以所選殖出的7個具高度多型性的微隨體基因座作為遺傳標記,針對3個不同時空的日本鯷樣本進行系群判別。
以(AC)10、(TC)10、(CAC)5CA、CT(ATCT)6、(TGTA)6TG、CT(CCT)5等6種探針,針對2400個菌株進行雜合反應,一共篩選出31個重複單元次數大於5的微隨體基因座其中包括三種額外的基因座,和一些複合式的基因座。其中成功設計引子者有26個。經測試後發現此26個基因座皆呈多型性,其中有7對具有高度的多型性,而且解析效果良好。
在族群狀態方面,日本鯷其3組樣本皆顯著地偏離哈溫平衡,並且有異型合子不足的現象。固定指數和遺傳距離兩個參數皆顯示宜蘭和澎湖的兩個族群的差別,在統計上並不顯著。於相關性的檢測上,對偶基因共享度MXY值的敏感度高於親緣關係度RXY值,MXY值為基礎所畫出的樹狀圖中有較高比例的宜蘭灣族群被包含於一群中。可能暗示了宜蘭灣日本鯷秋季系群的存在。而兩種微隨體基因座演化模式的套用,具現有的數據,並不能明確的指出何種模式適用於日本鯷,但是基因座EJ41.1同時棄卻兩種突變模式(IAM, TPM),可能是由於此基因座本身結構上的因素,不能以TPM解釋,但是不符合IAM的原因仍有待商榷。
日本鯷是否有先混群再分別洄游至台灣東、西北海域;宜蘭灣族群是否有秋季群的存在,則仍須進一步的實驗及分析。
目錄
致謝 I
摘要 II
目錄 III
壹、前言 1
1 研究動物簡介 1
2 微隨體基因座 4
2.1 微隨體的發現 4
2.2 微隨體基因座和其類型 5
2.3 微隨體基因座的特點 6
2.3.1 豐富性與分佈 6
2.3.2 多型性(Polymorphism) 6
2.3.3 共顯性遺傳(Codominance) 7
2.3.4 序列長度及偵測 7
2.4 微隨體基因座的演化 7
貳、實驗材料與方法 9
1 標本取得 9
2 選殖微隨體基因座 9
2.1 萃取高分子量基因組DNA 9
2.2 製備接合體DNA 9
2.3 接合反應(Ligation) 10
2.4 製備勝任細胞(Competent cell) 10
2.5 轉型(Transformation)和藍白篩選 10
2.6 聚合連鎖反應檢查接合作用 11
2.7 製備探針 11
2.8 轉漬(Blotting) 12
2.9 雜合(Hybridization)、引子設計 12
3 篩選多型性的微隨體基因座 13
3.1 萃取標本DNA 13
3.2 聚合連鎖反應 13
4 判讀對偶基因 14
4.1 製備放射性引子 14
4.2 放射性聚合連鎖反應 14
4.3 定序膠體電泳(Sequencing gel electriophoresis) 14
5 資料分析 15
5.1 遺傳變異(Genetic variation) 15
5.2 檢測哈溫平衡(Hardy-Weinberg equilibrium) 15
5.3 共祖係數(Coancestry, θ)遺傳距離(Genetic distance) 16
5.4 以階梯式突變模式(Step-wise mutation model,簡稱SMM)為依據的系群距離指數 17
5.5 測驗突變模式 18
5.6 估計相關性(Relatedness) 18
5.6.1 親緣關係度(Rxy) 18
5.6.1 對偶基因共享度(Mxy) 19
參、結果 20
1 日本鯷的微隨體基因座 20
1.1 微隨體基因座的特性 20
1.2 多型性的微隨體基因座 20
1.3 多型性微隨體基因座對偶基因的特性 20
1.4 遺傳變異 21
2 日本鯷的族群狀態 21
2.1 哈溫平衡 21
2.2 對偶基因分佈 22
2.3 系群分化指標-固定指數 22
2.4 遺傳距離 22
2.5 突變模式的套用 23
2.6 相關性 23
肆、討論 24
1 多型性的日本鯷基因座 24
2 日本鯷的族群狀態 25
2.1 哈溫平衡 25
2.2 固定指數和遺傳距離 26
2.3 對偶基因分佈 26
2.4 突變模式的套用 27
2.5 相關性 27
伍、參考文獻 29
表….……………………………………………………………………...……35
圖……………………………………………………………………..…….….42
附錄……………………………………………………………………..……..54
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