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Please use this identifier to cite or link to this item: http://ntour.ntou.edu.tw:8080/ir/handle/987654321/9875

Title: 凋亡相關基因操控誘發不孕基因改造魚技術之研發(I)
Technology Development of Inducible Sterility in Genetically Modified Fish by Control of Apoptotic Gene Expression (I)
Authors: 何國牟
Contributors: NTOU:Institute of Bioscience and Biotechnology
Keywords: 基因改造的生物;不孕;斑馬魚;原始生殖細胞基因;凋亡基因;綠螢光蛋白;生物環境安全
Genetically modified organisms (GMOs);Sterility;Zebrafish;Primordial germ cell (PGC);Apoptoic gene;Green fluorescent protein (GFP);Biological environment safely
Date: 2005-08
Issue Date: 2011-06-28T07:17:47Z
Publisher: 行政院國家科學委員會
Abstract: 摘要:基因改造的生物(GMOs)現存於許多物種,已成為農業生產的重大利基和生物技術學方面的一個主要討論領域。但是,如果這些基因改造的生物有一些適應優點,它們也可能成一個環境危害因子,當它們被釋放出來。GMOs 被釋放出來以後對生態環境主要害處在其轉殖基因的傳播。另一風險,入侵,介入生態系阻斷以基因改造生物優勢個體替換它們的相對的野生物種。雖然以基因轉殖技術為主開發之螢光觀賞魚品種,或其它高經濟價值性(生殖發育,免疫與抗環境逆境能力等其它生理功能)基因改造之魚類之遺傳育種技術以趨成熟。為使台灣基因轉殖觀賞魚及經濟價值性基因改造魚類符合國際生物安全議定之範圍(GMO 生物環境安全評估),及控制高價值基因改造魚類外流,應儘速建立不孕之遺傳育種技術。雖然目前三倍體或賀爾蒙控制生殖器發育等遺傳育種技術能使大部分的基因改造螢光魚不具生殖能力,但卻無法完全達到GMO 生物環境安全評估的標準。這裡, 我們提出一個新穎策略以受控制凋亡(apoptosis)原始生殖細胞(PGC)在發育中的生殖器內,可誘發不育之斑馬魚。為建立斑馬魚具有雄性和雌性退化生殖器,我們能創造基因轉殖斑馬魚藉由原始生殖細胞(PGC)基因起動子如vasa,nanos,dead-end,H1M 和Ziwi 結合Tet-on系統在一個短的時期裡表現凋亡基因譬如bad、bax和bcl-2在發育中的生殖器內。我們能使這個基因轉殖斑馬魚與野生斑馬或其它基因改造的斑馬魚(突變種)在正常情況(Tet off)交配產下小魚,小魚在引發生殖器凋亡之後養成成魚隨後設計做一系列的育種的試驗。在這個研究計劃,我們計劃使用斑馬魚作為一個動物模式來建立預防基因改造生物逃脫至自然環境的一個新穎技術。我們將採取斑馬魚對許多技術的可行性的好處譬如基因轉殖表現上的獲增功能,並且斑馬魚資料庫和知識被積累在相關網站亦成為有用的來源使我們執行這個提出的研究計劃。終於,此基因轉殖斑馬魚藉的創立與可誘導在胚胎階段之原始生殖細胞凋亡或成魚之退化生殖器的發生而使其不孕。在第一這個提出的研究計劃裡,我們已計劃完成下列實驗:1.分析凋亡基因作用在班馬魚生殖器之發育階段之損傷程度第一年的目標是找出凋亡基因作用在班馬魚生殖器發育階段之損傷程度。首先分離原始生殖細胞基因(vasa, nanos, dead-end、H1M和Ziwi等)起動子並測定其活性受。其後構築表現載體,用適當之原始生殖細胞基因起動子來表現凋亡基因(bad,bax and bcl-2等)。損傷班馬魚生殖器的形態改變將由組織學分析及原位處雜交作進一步簡查。2.建立誘發不孕之斑馬魚技術開發之基因轉殖斑馬魚在確定凋亡基作用在班馬魚生殖器之發育階段後,第二年開始建立斑馬魚細統具有雄性和雌性退化生殖器,我們能創造基因轉殖斑馬魚藉由原始生殖細胞(PGC)基因起動子如vasa, nanos, dead-end、H1M 和Ziwi 與Tet-on 系統在一個短的時期裡表現凋亡基因譬如 bad、bax 和bcl-2 在發育中的生殖器內。我們能使這個基因轉殖斑馬魚與野生斑馬或其它基因改造的斑馬魚(突變種)在正常情況 (Tet off)交配產下小魚,小魚在引發生殖器凋亡之後養成成魚隨後設計做一系列的育種的試驗。
Abstract:Genetically Modified Organisms (GMOs) now exist for many organisms, producing significant promise for agricultural production and a major field of discussion in biotechnology. However, if these organisms have some fitness advantage, they may also pose an environmental harm when released. The major risk of ecological harm is associated with the spread of transgenes after a genetically modified organism (GMO) release. The other risk, invasion, involves ecosystem disruption as GM individuals replace their WT counterparts. Now the transgenic technology for generating fluorescent pet aquarium fish or other high economic worth (gonad development, immunity and ability to resist environmental adverse circumstance etc.) GMO fish is very easy. To reach the standard of international biological safety (GMO biological environment is assessed safely) for pet aquarium fish or other high economic worth fish in Taiwan, and control the high value gene and GMO fish's outflow , it is necessary to built up " infertile" breeding technology " as quickly as possible. The " triploid " or "hormone" technology may control GMO fish breeding but those strategies are still unable to totally reach the standard of GMO biological environment safety. Here, we provide an novel strategy of inducible sterility in transgenic zebrafish with controlled apoptosis within developing gonad. To zebrafish lines with a degenerative gonad of both male and female , we can create a transgenic line of zebrafish (Danio rerio ) by expressing a apoptoic genes such as bad, bax and bcl2 driven by a promoter of primordial germ cell (PGC) specific genes such as vasa, nanos-like, dead-end, H1M and Ziwi combined with Tet-on system into developing gonad at a short period of time. We can cross this GM with WT or other transgenic fish under normal conditions (Tet off) in the laboratory and conducted induction of apoptosis in the developing gonad of their progeny and subsequently designed to make a series of breeding trials. In this project, we plan to use zebarfish as a model organism to establish an novel technique for preventin g GMOs from escaping into nature. We will take the advantage that zebrafish is amenable to many techniques such as transgenic expression for "gain of function", and data bases and the knowledge accumulated on the web sites have become useful sources for us to conduct this proposed project. Finally, the establishment of transgenic zebrafish with inducible apoptoic PGCs in embryonic stage or degenerative gonad in adult will be used by Tet-On system and vasa, nanos-like, dead-end, H1M and Ziwi promoter driving bax or bcl-2 gene expression. In this proposed project we had performed the following experiments: 1: Target overexpression of proapoptic genes in the developing gonad of larval zebrafish 2: To generate those transgenic lines for inducible sterility zebrafish models in which the developing gonad is totally or partially damaged and led to asexualize or sterilize adult zebrafishs
Relation: NSC94-2317-B019-005
URI: http://ntour.ntou.edu.tw/ir/handle/987654321/9875
Appears in Collections:[生命科學暨生物科技學系] 研究計畫

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