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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
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dc.contributor.author | 鍾至青 | zh_TW |
dc.date.accessioned | 2021-07-01T08:20:23Z | - |
dc.date.available | 2021-07-01T08:20:23Z | - |
dc.date.issued | 1997 | |
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dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/76332 | - |
dc.description.abstract | 以水母綠螢光蛋白cDNA (GFP cDNA)為報導基因,具有容易檢測、高敏感度、可在活體直接觀察等優點。為評估其應用於稻田魚基因轉殖模式的可行性,將建構有human cytomegalovirus immediate early enhancer/promoter的GFP cDNA基因片段(pGreen LANTERN),經Sca I線性化後,各以5μg/ml、7.5μg/ml、10μg/ml三種不同DNA濃度,進行稻田魚未授精卵顯微注射實驗。授精後12小時(stage 15)在FITC螢光濾鏡下,3組實驗組皆開始可以偵測到綠色螢光出現,表現率分別是5μg/ml組為50%、7.5μg /ml組為63%、10μg/ml組則為100%;比較表現GFP胚胎授精後24小時(stage 19)存活率,發現10μg/ml組為0,明顯較5μg/ml組(40%)、7.5μg/ml組(50%)低,顯示乙太高濃度強?動子表現GFP易影響稻田魚胚胎的正常發育。觀察7.5μg/ml組表現GFP存活的胚胎,其綠色螢光至少可維持至孵化後10天,且與PBS對照組比較,顯示在稻田魚發育早期無內生性的綠色螢光幹擾,證明應用GFP cDNA在稻田魚轉殖模式研究基因表現是可行的。 為研究鯉魚視紫質基因在胚胎發育早期的調控表現,分別建構以鯉魚視紫質(rhodopsin)基因(第二型)上游3.5kb與0.5kb驅動的GFP cDNA基因片段,觀察其在基因轉殖稻田魚胚胎的表現情形。視紫質上游3.5片段在授精後18小時(stage 16)開始驅動GFP cDNA表現,進入原腸胚初期(授精後48小時,stage 23)曾短暫消失,在授精後72小時(stage 27)又再度可以偵測到GFP表現,且僅限於眼睛部位。眼中的綠色螢光至少可以維持至孵化後一天,顯示鯉魚視紫質基因上游3.5kb對於視紫質的表現具有組織專一性。但是在轉殖此基因上游0.5kb片段時發現其對於驅動GFP cDNA表現的能力較弱,自授精後18小時(stage 16)開始可以偵測到微弱GFP表現;原腸胚階段並沒有短暫消失現象,且只在卵黃囊外圍有微量GFP分佈。發育至授精後96小時,GFP都僅在卵黃囊分佈;眼睛部位則在孵化前一天開始有綠色螢光產生,顯示此片段不具組織專一性。因此,與鯉魚視紫質基因組織專一性表現有關的序列,應介於上游0.5-3.5kb間。 另一方面為瞭解由吳郭魚選殖出c-ski基因在魚類胚胎早期表現情形,將此基因上游3kb建構至GFP cDNA表現載體。經基因轉殖後,在稻田魚胚胎發育至授精後12小時(stage 15)開始表現,進入原腸胚期後,則專一在卵黃囊外膜表現,至授精後120小時(stage 34)達到最高,並持續至孵化。孵化後,隨著卵黃囊消失而減弱,至孵化後第5天完全消失。顯示吳郭魚c-ski的表現可能與維持卵黃囊正常生理活動有關。 | zh_TW |
dc.description.provenance | Made available in DSpace on 2021-07-01T08:20:23Z (GMT). No. of bitstreams: 0 Previous issue date: 1997 | en |
dc.description.tableofcontents | 論文摘要. . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 第一章 評估應用水母綠螢光蛋白cDNA在稻田魚轉殖模式做為報導基因的可行性 I、前言. . . . . . . . . . . . . . . . . . . 3 II、材料與方法. . . . . . . . . . . . . . . . . . . . . . . . . . 7 III、結果. . . . . . . . . . . . . . . . . . 13 IV、討論. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14 第二章 鯉魚視紫質基因在基因轉殖魚早期胚胎發育時的表現 I、前言. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 16 II、材料與方法. . . . . . . . . . . . . . . . . . . . . . . . . . . . 19 III、結果. . . . . . . . . . . . . . . . . . . . . . . . . . . . . 23 IV、討論. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 25 第三章 吳郭魚c-ski基因在基因轉殖魚早其胚胎發育時的表現 I、前言. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 28 II、材料與方法. . . . . . . . . . . . . . . . . . . . . . . . . . . . 31 III、結果. . . . . . . . . . . . . . . . . . . . . . . . . . 33 IV、討論. . . . . . . . . . . . . . . . . . . . . . . . . . . . . 34 參考資料. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 36 藥品與配方. . . . . . . . . . . . . . . . . . . . 42 | |
dc.language.iso | zh-TW | |
dc.title | 應用水母綠熒光蛋白研究基因在胚胎發育早期的表現動態 | zh_TW |
dc.title | Application of Aequorea victoria Green Fluorescent Protein to Study the Dynamics of Genes Expression | en |
dc.date.schoolyear | 85-2 | |
dc.description.degree | 碩士 | |
dc.relation.page | 80 | |
dc.rights.note | 未授權 | |
dc.contributor.author-dept | 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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