Development of Q-PCR on a Chip System for DNA Detection and Quantification

• Main Authors: Huang-Jen Chen, 陳煌仁
• Other Authors: Da-Sheng Li
• Format: Others
• Language: zh-TW
• Published: 2008
• Online Access: http://ndltd.ncl.edu.tw/handle/hz54af

碩士 === 國立臺北科技大學 === 能源與冷凍空調工程系碩士班 === 96 === Real-Time Polymerase Chain Reaction (PCR) technique is one of the most important technologies for detecting biomolecμles in recent years. The technique is requires a thermal cycling process to amplify target DNA strand in vitro, intercalation of labeling into the amplified DNA strand, and detection of the variation of fluorescence intensity of labeling dye during the amplification of DNA strand a Fluorimeter. Then, the concentration of target DNA strand can be determined from the variation of fluorescence intensity. An instrument that performs such a technique is called a real-time PCR machine. This project aims to develop a real-time PCR machine that can amplify DNA strand in a microfluidic chip and simμltaneously detect the variation of fluorescence intensity of sample solution in chip for quantitatively determining the concentration of target DNA strand. Since the amplification of DNA strand is conducted in a microfluidic chip, it coμld improve the sensitivity of quantitative determination of target DNA concentration and reduce the duration for performing such a real-time PCR process. The PCR chips of this study is to develop micro temperature sensors and heaters, which are combined with a suitable control circuit to be used in micro PCR system, based on MEMS(Micro-electro-mechanical system) technologies. The integrated system including the PCR chip by MEMS and the control circuit applies for amplification and detection of DNA. Subsequently, we analyzed the temperature field of the reaction chamber by numerical analyses software, ANSYS. Two of advantages in this technique are displayed. One is the analysis of heating transference in different volume and the other is increasing the speed of cooling. Taken together, our study can significantly improve the efficiency and the success of PCR. Otherwise, this reaserch exploit the flurimeter which a commercial spectrometer with a confocal design for optic path. After performing a calibration test, the detection sensitivity and range of spectrometer outperforms the Fluometer used in the real-time PCR machine provided by major pharmaceutical companies. The proposed spectrometer can perform the continuous detection of fluorescence of which the wavelength ranges from 400 to 800 nm. And focus on the improvement of detection sensitivity of platform by modifying the positioning mechanism of chip and by reducing the background optical noise through a 700 alignment between light source and the light receiving fiber. the design of Compound Parabolic Concentator coμld enhance the flourencence detecting value more efficacious.Here a new technique was established in this study.The outcome of this study has constructed a real time pcr machine for DNA amplification and quantification.In the future, this instrument can be commercialezed for a low cost and high accuracy DNA quantification solution.