碩士 / 國立成功大學 / 奈米科技暨微系統工程研究所 / 96 / This study presents an integrated microfluidic system for fast diagnosis of agricultural diseases with the ability to perform 4 reverse-transcription polymerase chain reaction (RT-PCR) processes at the same time in an automatic format. This system integrates a micro temperature control module and a microfluidic control module. The micro temperature control module comprising micro temperature sensors and array-type microheaters provides precise, fast, and uniform temperature conditions for bio-reaction. The microfluidic control module made of biocompatible materials can transport samples and reagents by using pneumatic micropumps, microvavles and microchannels. Moreover, by using random primers into the reverse-transcription process, the chip design can be simplified and the consumption of reverse-transcription (RT) products in the following multiple polymerase chain reactions (PCR) is also minimized. Since there is only one reverse-transcription reaction involved, the following steps for distributing reverse-transcription products to four PCR chambers can be done simultaneously. When the transportation process between the RT chamber and the PCR chambers was finished, PCR process was then performed to amplify detection genes for each RNA-based virus. This microfluidic chip system fabricated by using micro-electro-mechanical-system (MEMS) technology may have the following advantages, including high sensitivity, fast diagnosis, disposability, low reagent and sample consumption, portability, low power consumption and low cost. The rapid heating/ cooling of this thermocycling system is 20℃/ 10℃. Experimental data showed that the developed system can successfully detect 4 kinds of purified RNA samples, including Nervous Necrosis Virus, Irido virus, Vibrio anguillarum, and grouper Mx protein gene. The detection limit of this system was found to be 101 copies/ ml while the conventional method was 103 copies/ ml. The total reaction time of this developed method was about 2.5 hours. An end-point detection module was also fabricated in this study. The detection limit of the end-point detection module was 103 copies/ ml. The end-point detection module could be integrated into the microcluidic system for its portability. The developed microfluidic system may provide a powerful tool for fast disease diagnosis of RNA-based virus.
| Identifer | oai:union.ndltd.org:TW/096NCKU5795018 |
| Date | January 2008 |
| Creators | Szu-hsien Lee, 李思賢 |
| Contributors | Gwo-Bin Lee, 李國賓 |
| Source Sets | National Digital Library of Theses and Dissertations in Taiwan |
| Language | zh-TW |
| Detected Language | English |
| Type | 學位論文 ; thesis |
| Format | 120 |
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