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研究生:周楷倫
研究生(外文):CHOU, KAI-LUN
論文名稱:大棘大眼鯛育苗技術與精液凍結保存技術開發
論文名稱(外文):The study of larval rearing and sperm cryopreservation conditions of Priacanthus macracanthus
指導教授:侯清賢侯清賢引用關係李彥宏李彥宏引用關係
指導教授(外文):HOU, CHING-HSIENLEE, YAN-HORN
口試委員:李彥宏侯清賢張錦宜吳豐成翁進興
口試委員(外文):LEE, YAN-HORNHOU, CHING-HSIENCHANG, CHIN-IWU, FENG-CHENGWENG, JINN-SUING
口試日期:2022-07-21
學位類別:碩士
校院名稱:國立高雄科技大學
系所名稱:漁業生產與管理系
學門:農業科學學門
學類:漁業學類
論文種類:學術論文
論文出版年:2022
畢業學年度:110
語文別:中文
論文頁數:61
中文關鍵詞:大棘大眼鯛仔魚胚胎發育海洋資源復育精子凍結保存
外文關鍵詞:Embryonic developmentLarvaeMarine resource restorationPriacanthus macracanthusSperm cryopreservation
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大棘大眼鯛(Priacanthus macracanthus)漁獲量於1996年達最高峰的7,386噸後,即開始逐年減少,2020年時漁獲量僅餘約600噸,為需進行資源復育的目標魚種對象之一。本研究針對大棘大眼鯛育苗技術進行研究,嘗試攻克其初期開口餌料生物問題,並試驗其精子凍結保存條件與可行性實驗,提供未來大棘大眼鯛栽培漁業與資源增裕之魚苗需求。本研究樣本以東港拖網採集,經鏡檢卵徑大於500μm 之雌魚,給予魚用絨毛膜激性腺激素200 IU/kg 1劑後,記錄與量測受精卵與仔魚。精子凍結保存實驗部份,測定精液滲透壓,以4 ℃冷藏條件篩選出適當稀釋液後,再進行冷凍保存試驗。結果顯示,大棘大眼鯛受精卵於水溫28 ℃ 條件下,歷時18小時30分鐘後成功孵化;於水溫26 ℃條件下,歷時20小時58分鐘後成功孵化;而水溫20 ℃ 條件下,歷時30小時30分鐘後成功孵化,水溫愈高仔魚發育速度愈快,且內源性營養利用亦加快,於三個測試水溫(28 ℃、26 ℃、20 ℃)下皆觀察到攝餌仔魚,且28 ℃組於孵化後第5天死亡、26 ℃於孵化後第10天死亡、20 ℃於孵化後第6天死亡。精子凍結保存實驗部份,精液的等滲透壓範圍約283-404 mOsm/kg,稀釋液則以Dulbecco’s Modified Eagle Medium/Nutrient Mixture F-12與368 mOsm/kg NaCl Solution擁有較佳精子存活率,並以低倍率稀釋與低抗凍劑濃度為佳。
In 1996, the 6,093t Priacanthus macracanthus (Red bigeye) was caught and reached the highest catch. Soon after, the catch decreased annually, and the catch reduced to 600 t in 2020. It shows that red bigeye is the one of species need to conduct resource restoration. Therefore, in the present study, we focus on the larval rearing of red bigeye, and attempt to overcome the diet for mouth opening. In addition, we tested the sperm cryopreservation condition and the feasibility of red bigeye to benefit cultivating fishery and resource restoration. The present study sample was caught by trawl net (Donggang) and examined by microscope. If the eggs diameter > 500 μm, one dose human chorionic gonadotropin 200 IU/kg would be injected for fish. In addition, the fertilized eggs and the larvae would be measured and recorded by microscope. In the sperm cryopreservation, the sperm osmolality would be measured in advance. The extender was selected by the ability of stored in 4 ℃ used in the experiment of sperm cryopreservation. The results of study showed that the fertilized egg of bigeye took 18 hr 30 min to hatch at 28 ℃. However, it took 30 hr 30 min to hatch at 20 ℃. The higher the water temperature, the faster of larval development and endogenous nutrition utilization, and the feed intake larvae were observed at three water temperature (28 ℃、26 ℃、20 ℃). Also, the larvae dead at 5 days post hatching (dph) in 28 ℃, and the larvae dead at 10 dph in 26 ℃, and the larvae dead at 6 dph in 20 ℃. Result of sperm cryopreservation experiment, the sperm osmolality ranged from 283 - 404 mOsm/kg. Two extender that Dulbecco‘s Modified Eagle Medium/Nutrient Mixture F-12 and 368 mOsm/kg NaCl Solution had good sperm viability. The low dilution and antifreeze concentration had better performance.
摘要 I
ABSTRACT II
謝辭 III
目錄 IV
圖目錄 VII
表目錄 VIII

第壹章 前言 1
第一節 漁業資源與栽培漁業概述 1
第二節 種魚培育 3
第三節 繁殖及育苗 4
第四節 魚類精液凍結保存技術 5
第五節 餌料生物 6
第六節 大棘大眼鯛生態與資源量簡介 7
第七節 研究動機 8
第八節 研究目的與目標 8

第貳章 研究材料與方法 9
第一節 生物數據 9
一 樣本獲取 9
二 生殖腺發育變化 9
第二節 種魚培育 10
一 養殖環境條件準備 10
二 馴化 10
第三節 人工催熟與授精 11
一 卵徑與精子存活率測定 11
二 種魚催熟 11
三 人工授精 11
四 受精卵發育觀察與紀錄 12
第四節 仔魚培育 12
一 育苗環境 12
二 仔稚魚餌料 12
三 餌料培育與滋養 12
四 仔魚觀察 13
第五節 精液冷藏與凍結保存 13
一 樣本取得 13
二 滲透壓與等滲透壓條件測試 13
三 精液冷藏保存試驗 13
四 精液凍結保存試驗 14
五 長期凍結保存試驗 14
六 精子螢光染色判讀 14
第六節 統計分析 15

第參章 結果 16
第一節 東港海域大眼鯛生殖腺年週期性發育變化 16
一 雄、雌魚之GSI變化 16
二 生殖腺發育變化 16
第二節 種魚培育 17
一 野外撈捕 17
二 運輸 17
三 馴化 18
四 生殖腺成熟度檢測 18
第三節 人工催熟與授精 19
一 荷爾蒙刺激與人工授精 19
二 受精卵發育 19
第四節 仔魚培育 21
一 水質變化 21
二 仔魚發育 21
三 仔稚魚餌料 22
第五節 精液凍結保存 23
一 滲透壓與等滲透壓條件 23
二 冷藏保存條件測試 23
三 凍結保存與抗凍劑濃度 24
四 長期凍結保存 24
五 精子螢光染色判讀 25

第肆章 討論 26
第一節 大眼鯛生殖與種魚培育 26
一 繁殖期與生殖 26
二 種魚培育 27
第二節 受精卵與仔魚發育 28
一 受精卵發育 28
第三節 精液凍結保存 30
第伍章 結論與建議 31
參考文獻 32

圖目錄
圖1 1993年至2020年臺灣大眼鯛漁獲量與產值變化 39
圖2 2021年後壁湖每月平均海水表層水溫變化 40
圖3 2021年3月至2022年2月大眼鯛生殖腺指數變化 41
圖4 2021年3月至2022年2月大眼鯛生殖腺外觀變化照片 42
圖5 2021年3月至2022年2月大眼鯛鏡檢卵細胞發育變化照片 43
圖6 2021年3月至2022年2月間大眼鯛之不同大小卵細胞佔比頻度 44
圖7 2021年3月至2022年2月大眼鯛精巢組織切片變化照片 45
圖8 2021年3月至2022年2月大眼鯛卵巢組織切片變化照片 46
圖9 大眼鯛運輸流程照片 47
圖10 大眼鯛生殖腺檢測照片 48
圖11 大眼鯛注射激素催熟照片 49
圖12 大眼鯛人工授精流程照片 50
圖13 大眼鯛受精卵於不同水溫發育過程照片 51
圖14 大眼鯛仔魚在不同水溫發育過程照片 52
圖15 大眼鯛仔魚在不同水溫成長變化 53
圖16 大眼鯛仔魚在不同水溫攝餌照片 54
圖17 大眼鯛精液分別添加3種稀釋液與使用4種稀釋比例之精子存活率變化 55
圖18 大眼鯛稀釋精液以4種濃度分別添加3種抗凍劑之精子存活率變化 56
圖19 大眼鯛稀釋精液於長期凍結保存之精子存活率變化 57
圖20 精子螢光染色照片 58
圖21 螢光染色判讀新鮮與凍結24小時精液之精子存活率變化 59

表目錄
表1 受精卵發育特徵 60
表2 仔魚養殖期間水質變化 61

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