Application of MicroEmusification：Emulsion Polymerase Chain Reaction

• Main Authors: Ying-Tu Su, 蘇英圖
• Other Authors: Jyh-Jong Sheen
• Format: Others
• Language: zh-TW
• Published: 2009
• Online Access: http://ndltd.ncl.edu.tw/handle/51940901593932821213

碩士 === 國立臺灣海洋大學 === 機械與機電工程學系 === 97 === In this thesis, soft lithography technique was used to produce a micro emulsion chip. This chip can handle tiny volume of DNA solution for polymerase chain reation (ePCR). The DNA solution is dispersed in the emulsified droplets in a continuous oil phase, and the droplets become reaction space for PCR. As a result, the consistency of the multiple types of DNA templates existing in the original reaction space can be improved by emulsification process. Also, it will reduce competitive sequence interference and improve DNA magnification. Differing from usual micro emulsification chips, this study focuses on the technical problems involved in dealing with tiny and expensive DNA solution in order to avoid non-uniform droplets in the transient of droplet generation. The main ideas are as follows: (1) Let water first go through transient process and produce water droplets; (2) Use water and bubble to place DNA solution in between, and pass it through emulsification hydrodynamic focusing channel; (3) Use pneumatic membrane valves control the stop and flow of water and DNAsolution. Characterization of the emulsification chips includes droplet sizes, uniformity of droplets and hot-resistance of water-in-oil droplets. The smallest droplet diameter is less than 10 μm and the coefficient of variation is about 2%. The resulting electrophoretograms of both single or multiple templates’ PCR demonstrate that ePCR can improve the precision of amplified DNA segments. It is suggested from the experiment results that ePCR was proven to be efficient and could be used in further genetic research Key word: ePCR, DNA template, transient process, tiny-volume micro-emulsification