2009-01-012024-05-17https://scholars.lib.ntu.edu.tw/handle/123456789/679015摘要：本研究計畫將以新的DNA 檢測技術，避開目前PCR 加熱循環(thermal cycling)的增生 專利，進而開發檢測DNA 的微流體晶片。新的檢測技術可完全替代PCR 之酵素化學方法 達到節省昂貴試劑與儀器之花費，也可避免PCR 專利。本實驗室先期的研究成果顯示， 可以將1:100 比例的探針DNA(probe DNA)與條碼DNA(bar-code DNA)，同時接合在奈米 金珠(AuNP)上。如果重複此放大過程五次，就可輕易放大原始目標DNA 訊號1010 倍，遠 大於PCR 五次熱循環的25 倍。利用此一成果，可以設計不需要熱循環作增生原始目標 DNA 的檢測方法，在每個循環檢測過程中，將目標DNA(target DNA)設定為上個循環過 程的條碼DNA，經過數次的轉換目標DNA 的過程，可以將DNA 訊號放大，最後再以螢光 檢測儀定量，達到檢測原始目標DNA 的目的。再者，一般DNA 接種在奈米顆粒的方法都 是將DNA 滴入奈米流體後靜置，非常耗時。本研究計劃則提出利用微流道設計，讓流體 在微流道內產生渦流的方式，或利用奈米磁流體，在加上外加磁場後，可加速DNA 與奈 米顆粒的接合。整個DNA 檢測方法， 皆可整合在一微流體晶片內進行， 實現 lab-on-a-chip，本研究計畫即要證明此方法之可行性。第一年要確認本方法可放大的 倍率與重複性，第二年要確認可在微流道內將DNA 接在奈米金與奈米磁球上，並了解奈 米磁球在微流道內可增加的混合效率，第三年要整合各微流道內的功能在一晶片上，完 成一個完整的目標DNA 的信號放大。<br> Abstract: This project proposes a novel DNA detection technique to avoid some patents of thermal cycling applied in real-time PCR systems. Furthermore, the project will integrate necessary processes into a bio-chip, namely, to develop Lab-on-a-Chip. The new technique not only avoids violating the patents but also saves the cost of equipments and reagents. The preliminary study showed that target DNA and bar-code DNA can be immobilized on gold nanoparticles (AuNPs) with an amount ratio 1:100. Based on this fact, one can design a new DNA detection principle to amplify the signal 1010 times by repeating the process five times.This amplification is much larger than 25 times by PCR under five thermal cycling processes. In stead of duplicating the target DNA with thermal cycling process, for each cycle, set the target DNA as the bar-code DNA from the previous cycle. So after several such process of switching target DNA, one can amplify the signal and determine the amount of the original target DNA by the fluorimeter. In addition, to speed up the immobilization process of DNA on AuNPs in microchannels, magnetic nanoparticles (MNPs) will be used. With existence of the external magnetic fields, MNPs can help the mixing of DNA and nanoparticles and filter magnetic particles from non-magnetic ones. All the detection processes can be integrated into a microchannel chip. The aim of the project is to demonstrate the feasibility of the new detection technique. In the first year, the project would verify the optimal amplification and repeatability by using bar-code DNA. In the second year, design and work necessary detection processes in microchannels. In the third year, integrate all parts on a chip to achieve the goal of lab-on-a-chip for DNA detection.即時偵測同步定量聚合脢連鎖反應溫控機構螢光檢測儀晶片實驗室奈real-time PCRthermal cyclingfluorimeterlab-on-a-chipgold