https://scholars.lib.ntu.edu.tw/handle/123456789/86178
標題: | 行政院國家科學委員會專題研究計畫期中進度報告:有機污染物綠色螢光蛋白生物偵測器之研發及應用(1/2) | 作者: | 廖秀娟 | 關鍵字: | 生物偵測器(biosensor);綠色螢光蛋白(GFP);reporter protein;有機污染物;biosensors;green fluorescence protein (GFP);organic pollutants | 公開日期: | 2004 | 出版社: | 臺北市:國立臺灣大學生物環境系統工程學系暨研究所 | 摘要: | 所謂生物偵測器(biosensors)是指sensing component (生化或生理的變化) 和transducing element (當環境改變時,會產生可測量的訊號) 之結合。以細胞為主的生物偵測器,可根據它 們的sensing element 的反應來做分類,例如:細胞代謝的改變、基改生物基因表達的改變等。 對於基因改造(genetically engineered)細胞的生物偵測器,可基因重組sensing element 和 reporter gene,此重組的質體(recombinant plasmid),會對外來污染物反應產生一種可量測的訊 號。Sensing element 通常是由regulatory protein 和promoter 所組成,這些sensing element 對 污染物的專一性決定此生物偵測器的選擇性,而reporter protein 則是決定此生物偵測器的敏 感性及偵測極限。 本研究研發以綠色螢光蛋白(green fluorescent protein,GFP)作為reporter protein 的專一 性有機污染物的生物偵測系統。針對環境中重要的有機污染物,例如:benzene 及其衍生物, naphthalene 及salicylate,PCBs,及chlorocatechols 發展具專一性有機污染物的生物偵測系統。 此種以GFP 作為reporter protein 的生物偵測器, 其優點是它具有自動發出螢光 (auto-fluorescence)的特性,其發光反應不需要添加任何cofactors 或受質,因此,此種生物偵 測器可提供迅速甚至即時(real time)環境中有機污染物資訊,此外,其優點尚包括spectral variants 以及在biological 之pH 下具有高穩定性、操作簡單、快速省時、以及在大部分的生物 中沒有類似的蛋白質等。此種以GFP 作為reporter protein 生物偵測系統,將可克服luc,lux, 及lacZ 作為reporter gene 的缺點。 本研究計畫分為二年執行,第一年的研究為以gfp 作為reporter gene 之生物偵測系統的 建立,其中包括生物偵測系統敏感度(sensitivity)及穩定度分析,以及建立benzene 及其衍 生物的生物偵測器,並探討此生物偵測器和benzene 劑量與反應關係(Dose-response relationship),以及時間與反應關係,以及應用於環境污染實場的測試(environmental sample field testing)。目前我們已成功的建構生物偵測器重組質體,相關的分析正進行中。 A biosensor is composed of a biological sensing component, which recognizes chemical or physical changes, couple to a transducing element that produces a measurable signal in response to the environmental insult. Cell-based biosensors can be classified according to the response of their sensing element, such as changes in cellular metabolism, pH, altered gene expression in genetically modified organisms. Genetically engineered cell-based biosensors can elicit a response in the presence of an environmental insult by coupling the sensing element to a reporter gene fusion, which upon expression produces a readily measurable signal. The sensing element is often composed of regulatory proteins and promoter. The specificity of these sensing elements for the environmental insult confers selectivity to the system, while the reporter protein determines the system’s sensitivity and detection limits. We proposed the construction, laboratory characterization, and environmental testing of genetically engineered GFP-based bacterial biosensors that detect specific organic pollutants (e.g., benzene, naphthalene, salicylate, PCBs, and chlorocatechols). The reporter plasmid in which expression of gfp gene for jellyfish Aequorea victoria green fluorescence protein (GFP) will be placed directly under the control of the specific promoter. The primary advantage of GFP as a reporter protein is its autofluorescence, and therefore, its use does not require the addition of cofactors or exogenous substracts to produce fluorescence. Additionally advantages of GFP included high stability at biological pH, assay simplicity, and lack of endogenous homologues in most target organisms. Therefore, such GFP-based biosensors might provide even real time monitoring environmental pollutants. This is a two-year project. In the first year, the main task is to establish a sensitive and stable biosensing system using GFP as reporter protein. In addition, the benzene and related compound biosensor will be developed. The dose-dependent and time-dependent relationship of the biosensor and organic compound will be investigated. Furthermore, the application of biosensor in detecting environmental will be explored. Currently, we have successfully developed the recombinant biosensor plasmids. Other investigations are in progress. |
URI: | http://ntur.lib.ntu.edu.tw//handle/246246/10764 | 其他識別: | 922211E002054 | Rights: | 國立臺灣大學生物環境系統工程學系暨研究所 |
顯示於: | 生物環境系統工程學系 |
檔案 | 描述 | 大小 | 格式 | |
---|---|---|---|---|
922211E002054.pdf | 127.11 kB | Adobe PDF | 檢視/開啟 |
在 IR 系統中的文件,除了特別指名其著作權條款之外,均受到著作權保護,並且保留所有的權利。