Reaction, Separation and Dtection of Biomacromolecules using Disposable Microchips

利用一次性微芯片进行生物大分子的反应、分离和检测

基本信息

批准号：
11450318
负责人：
FUJII Teruo
金额：
$ 3.52万
依托单位：
The University of Tokyo
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1999
资助国家：
日本
起止时间：
1999 至 2001
项目状态：
已结题

项目摘要

In this research project, we have been trying to realize low-cost microchips for biochemical reaction and analysis, which can be fabricated through micro-molding process of the silicone rubber material using silicon wafer as a mold master. Based on the research progress achieved in the previous years, we investigated the performance of the microchip for free-flow Electrophoretic Separation of DNA fragments in detail. And on-chip biochemical reactions such as PCR (polymerase chain reaction) and cell-free protein synthesis were also conducted to examine the potentials of the developed microchip.1) Free-flow Electrophoresis: Cr-Au electrodes are patterned on a glass substrate for application of high voltage electric field required for the electrophoretic separation of DNA fragment. By pasting the silicone rubber microchip, which has microfluidic structure on it, on the glass substrate, we could carry out the experiments on electrophoresis of DNA fragments. 100bp to several hundreds by DNA fragments can be successfully separated and detected within just two or three minutes. This is almost ten to fifteen times faster than the conventional slab-gel based method.2) Biochemical Reactions: with the same hybrid structure with a glass substrate and a silicone rubber microchip, we have conducted two kinds of biochemical reactions with temperature control through the integrated electrodes; PCR and cell-free protein synthesis. For the PCR reaction , it is confirmed that 700bp DNA fragment can be amplified. For cell-free protein synthesis, it is proved that Green Fluorescent Protein can be synthesized by detecting the emitted fluorescence from the target molecule.

在这个研究项目中，我们一直在尝试实现低成本的微芯片，以进行生化反应和分析，可以通过使用硅晶体制成的硅橡胶材料的微型焊接工艺来制造，硅橡胶材料作为霉菌。根据前几年取得的研究进展，我们研究了微芯片详细详细介绍DNA片段的自由流动电泳分离的性能。还进行了芯片生化反应，例如PCR（聚合酶链反应）和无细胞蛋白合成，以检查已发达的微芯片的电势。1）自由流量电泳：CR-AU电极在玻璃底物上用于应用高压电场的玻璃基材料，用于用于用于电动电场的高压电场。通过粘贴有硅橡胶微芯片，该微芯片在其上具有微流体结构，在玻璃基板上，我们可以对DNA片段电泳进行实验。仅在两到三分钟内可以成功分离和检测到100bp至数百个DNA片段。这比基于平板基凝胶的传统方法快十到十五倍。2）生化反应：具有带有玻璃基板和硅胶橡胶微芯片的相同杂种结构，我们已经通过集成电极进行了两种具有温度控制的生化反应； PCR和无细胞蛋白质合成。对于PCR反应，可以证实可以扩增700bp DNA片段。对于无细胞蛋白的合成，可以通过检测靶分子从靶分子中检测发射的荧光来合成绿色荧光蛋白。