本研究成功開發出具有穩定識別度及低成本恆溫式環形核酸增幅法聚合酶鏈鎖反應檢測設備，利用穩定溫度控制器將反應槽恆溫於設定之溫度區間及設定的時間內，配合螢光讀取系統量測螢光濃度訊號值，最後傳送至開發板分析訊號並顯示於螢幕。本研究所開發之檢測設備包含硬體及軟體兩部分，硬體方面設計出穩定加熱時間控制系統與螢光讀取系統，裝載Raspberry Pi開發板、穩定溫控加熱/降溫模組、螢光光學讀取模組及光學檢測暗箱；軟體部分使用Java程式語言撰寫，包含了主程式碼、溫度時間控制程式碼、UV LED激發程式碼、螢光判讀程式碼、訊號分析及螢幕顯示程式碼。由實驗結果可以得知，本研究所開發之檢測設備加熱效率能於4分鐘內達到65℃且控制於±1℃內，且成功以光敏電阻讀取由UV LED激發試劑所產生之螢光訊號，經重複實驗下仍可穩定判斷正負控制檢體訊號，在製作成本方面也比市售價格降低了50%以上。期望本研究開發之檢測設備於未來能應用於臨床檢驗，提高疾病篩檢之效率，即時對感染者施予治療，降低疾病致死率。

This thesis presents a piece of detection equipment for polymerase chain reaction by loop-mediated isothermal amplification (LAMP PCR) with stable identity and low cost. We designed the case of internal structure by SolidWorks and AutoCAD, and programmed the main program, temperature control, and florescent signal processing by Java. We applied precise temperature control technology, used reagents and fluorescent detection to significantly shorten the experiment time of LAMP PCR. The results reveal that LAMP PCR temperature can be accurately controlled at a margin of ±1℃, which can be completed within between 15 minutes and one hour (according to specimens of every variety), and that the fluorescent signals are read correctly with the positive or negative reaction. Thus, the proposed LAMP PCR detection equipment is more sensitive, stable and cost-effective. With such equipment, a lot of clinical applications lie before us.

[1]Tsugunori Notomi, Hiroto Okayama, Harumi Masubuchi, Toshihiro Yonekawa, Keiko Watanabe, Nobuyuki Amino and Tetsu Hase, “Loop-mediated isothermal amplification of DNA,” Nucleic Acids Research, 28, 2000.
[2]H. A. Erlich, “Polymerase chain reaction,” Journal of Clinical Immunology, 9, pp. 437-447, 1989.
[3]S. H. Zarkesh-Esfahani, M. T. Kardi, and M. Edalati, “Hepatitis C virus genotype frequency in Isfahan province of Iran: a descriptive cross-sectional study,” Virology journal, 1, pp. 7-69, 2010.
[4]K. J. Livak and T. D. Schmittgen, “Analysis of relative gene expression data using real-time quantitative PCR and the 2− ΔΔCT method,” Methods, 25, pp. 402-408, 2001.
[5]https://en.wikipedia.org/wiki/SYBR_Green_I
[6]林于勤，李淑英，陳豪勇，林鼎翔(衛生署疾病管制局研究檢驗組)，新型恆溫式圈環形核酸增幅法簡介與應用，疫情報導，第20卷第6期，2004年6月。
[7]Fukuta S, Iida T, Mizukami Y et al, “Detection of Japanese yam mosaic virus by RT-LAMP,” Archives of Virology, 148, pp. 1713-1720, 2003.
[8]Fukuta S, Kato S, Yoshida K et al, ”Detection of tomato yellow leaf curl virus by loop-mediated isothermal amplification reaction,” Journal of Virological Methods, 112, pp. 35-40, 2003.
[9]鄧明中，林育如，宣詩玲，李敏旭，黃天祥，黃金城，簡茂盛，應用反轉錄環形恆溫核酸增幅法偵測豬瘟病毒，台灣獸醫期刊，第34卷，第97-105頁，2008年。
[10]Ihira M, Yoshikawa T, Enomoto Y et al, “Rapid diagnosis of human herpesvirus 6 infection by a novel DNA amplification method, loop-mediated isothermal amplification,” Journal of Clinical Microbiology, 42, pp. 140-145, 2004.
[11]李敏旭，應用環形恆溫核酸增幅法偵測家禽流行性感冒病毒核酸，家畜衛試所學術研討專訊，第2卷，第5-6頁，2008年。
[12]C. H. Wang, K. Y. Lien , J. J. Wu and G. B. Lee, “A magnetic bead-based assay for the rapid detection of methicillinresistant Staphylococcus aureus by using a microfluidic system with integrated loop-mediated isothermal amplification,” Lab on a Chip, 11, pp. 1521-1531, 2011.
[13]Kuboki N, Inoue N and Sakurai T et al, “Loop-mediated isothermal amplification for detection of African trypanosomes,” Journal of Clinical Microbiology, 41, pp. 5517-5524, 2003.
[14]Poon LL, Leung CS, Tashiro M et al, “Rapid Detection of the Severe Acute Respiratory Syndrome (SARS) Coronavirus by a Loop-Mediated Isothermal Amplification Assay,” Clinical Chemistry, 50, 6, 2004.
[15]K. B. F. F. Mullis, F. Faloona, S. Scharf, R. K. Saiki, G. T. Horn, and H. Erlich, “Specific enzymatic amplification of DNA in vitro: the polymerase chain reaction,” Cold Spring Harbor Symposia on Quantitative Biology, 51, pp. 263-273, 1986.
[16]M. A. Northrup, C. Gonzalez, and D. Hadley, “A MEMS-based miniature DNA analysis system, ”Proceedings of the International Conference on Solid-State Sensor and Actuators, 1995.
[17]M. A. Burns, C. H. Mastrangelo, T. S. Sammarco, F. P. Man, J. R.Webster, B. N. Johnsons, and D. T. Burke, “Microfabricated structures for integrated DNA analysis,” Proceedings of the National Academy of Sciences, 93, pp. 5556-5561, 1996.
[18]A. T. Woolley, D. Hadley, P. Landre, A. J. deMello, R. A. Mathies, and M. A. Northrup, “Functional integration of PCR amplification and capillary electrophoresis in a microfabricated DNA analysis device,” Analytical Chemistry, 68, pp. 4081-4086, 1996.
[19]T. B. Taylor, E. S. Winn-Deen, E. Picozza, T. M. Woudenberg, and M. Albin, “Optimization of the performance of the polymerase chain reaction in silicon-based microstructures, ”Nucleic Acids Research, 25, pp. 3164-3168, 1997.
[20]M. S. Ibrahim, R. S. Lofts, P. B. Jahrling, E. A.Henchal, V. W Weedn, M. A. Northrup, and P. Belgrader, “Real-time microchip PCR for detecting single-base differences in viral and human DNA, ”Analytical Chemistry, 70, pp. 2013-2017, 1998.
[21]N. Zhang, and E. S. Yeung, “On-line coupling of polymerase chain reaction and capillary electrophoresis for automatic DNA typing and HIV-1 diagnosis,” Journal of Chromatography B: Biomedical Sciences and Applications, 714, pp. 3-11, 1998.
[22]R. P. Oda, M. A. Strausbauch, A. F. R. Huhmer, N. Borson, S. R. Jurrens, J. Craighead, and J. P. Landers, “Infrared-mediated thermocycling for ultrafast polymerase chain reaction amplification of DNA,” Analytical Chemistry, 70, pp. 4361-4368, 1998.
[23]M. U. Kopp, A. J. De Mello, and A. Manz, “Chemical amplification: continuous-flow PCR on a chip,” Science, 280, pp. 1046-1048, 1998.
[24]Y. C. Lin, M. Y. Huang, K. C. Young, T. T. Chang, and C. Y. Wu, “A rapid micro-polymerase chain reaction system for hepatitis C virus amplification,” Sensors and Actuators B: Chemical, 71, pp. 2-8, 2000.
[25]T. M. Lee, I. M. Hsing, A. I. Lao, and M. C. Carles, “A miniaturized DNA amplifier: its application in traditional Chinese medicine,” Analytical Chemistry, 72, pp. 4242-4247, 2000.
[26]K. Yasuda, K. Okano, and S. I. Ishiwata, “Focal extraction of surface-bound DNA from a microchip using photo-thermal denaturation,” Biotechniques, 28, pp. 1006-1011, 2000.
[27]G. V. Kaigala, V. N. Hoang, A. Stickel, J. Lauzon, D. Manage, L. M. Pilarski, and C. J. Backhouse, “An inexpensive and portable microchip-based platform for integrated RT–PCR and capillary electrophoresis,” Analyst, 133, pp. 331-338, 2008.
[28]Assawapong Sappat, Suriya Mongpraneet, Tanom Lomas and Adisorn Tuantranont (2009, Oct.), Multi-Channel Turbidity Detection of Shrimp Virus by Loop-Mediated Isothermal Amplification Reaction, IEEE SENSORS 2009 Conference, New Zealand.
[29]廖鴻昀，紀鈞瑋，勞業興，翁瑞鴻，黃振康，莊啟佑與謝書榮，應用於核酸檢測之可攜式螢光偵測模組開發，農業機械學刊，第二十一卷，2012年。
[30]Jianfeng Zhou, Yong Liao, Haitao Li, Xuelian Lu, Xiufeng Han, Yanli Tian, Shanshan Chen and Rongya Yang, “Development of a Loop-Mediated Isothermal Amplification Assay for Rapid Detection of Trichosporon asahii in Experimental and Clinical Samples,” BioMed Research International, 2015, Article ID 732573, 2015.
[31]劉文海，鋁合金潛力產品與前景分析，金屬中心產業報告，2014年10月，取自：https://goo.gl/oBhuUM
[32]Richard Lawler, “Raspberry Pi credit-card sized Linux PCs are on sale now, $25 Model A gets a RAM bump,” Retrieved April 24, 2017, from Aol Tech on the World Wide Web: https://goo.gl/ShvbwN
[33]Debian, “About Debian,” Retrieved April 24, 2017, from Debian on the World Wide Web: https://www.debian.org/
[34]Wikipedia, “Java (programming language),” Retrieved April 24, 2017, from Wikipedia on the World Wide Web: https://goo.gl/QPkij
[35]Wikipedia, “NetBeans,” Retrieved April 24, 2017, from Wikipedia on the World Wide Web: https://goo.gl/neQ1on
[36]Antonio Feteira, “Negative Temperature Coefficient Resistance (NTCR) Ceramic Thermistors: An Industrial Perspective,” Journal of the American Ceramic Society, 92, pp. 967-983, 2009.
[37]http://www.token.com.tw/index.html
[38]C.L. Tsai (2014/04/24), “MCP3008讀取類比訊號測溫度與光度,” Retrieved April 24, 2017, from World Wide Web: https://goo.gl/S4rpwv
[39]江賢龍、周玉崑(2009)，基礎電子實習(下)。台北：台科大。
[40]C. Aaij and P. Borst, “The gel electrophoresis of DNA,” Biochimica et Biophysica Acta, 269, 2, pp. 192-200, 1971.
[41]金龍俊科技股份有限公司，撓性超薄電熱片，Retrieved April 26, 2017, from World Wide Web: https://goo.gl/ERTDsu
[42]金龍俊科技股份有限公司，薄型高溫陶瓷電熱片，Retrieved April 26, 2017, from World Wide Web: https://goo.gl/bH84yr