MICROFLUIDS, MICROREACTIONS, AND MICROSEPARATIONS

DNA基因分型和测序的分析方法与从DNA源材料开始提取遗传信息的多步骤处理系统相关联。在传统的方法中，处理系统对微升体积的反应样品进行处理，并配备相应的液体处理装置和电泳分析设备。项目2将提供组件级设计和测试我们改进的纳升体积处理组件，用于DNA测序和基因分型。为了实现在单个设备上执行复杂的多步骤任务的目标，需要离散滴样处理和信息控制。新的或改进的制造技术也将根据需要进行检查。项目中开发的所有组件都保留了最终集成到项目1的复杂系统中的目标。项目2有三个具体目标：1)开发纳升注射和移动系统。我们将构建和测试坚固且灵活的注射和移动系统的组件，该系统可以处理体积范围为10 nl至500 nl的多种液体样品。我们成功的单滴和双滴注射和移动系统将被修改并组装成多个样品液体处理程序。每个离散的滴将被处理和操作独立。2)开发多个独立的反应室。我们将构建和测试反应室，以获得最佳的表面相互作用，温度分布和反应室组件之间的热串扰。一个重要的努力将包括开发控制软件和可重复的反应条件，以实现高质量的DNA测序。3)展示高分辨率的分离。我们将改进现有的凝胶电泳系统（大约10-50 bp分辨率，500 bp长度），以接近传统的大规模DNA测序（1 bp分辨率，1000 bp长度）的质量。优化现有的荧光标记和检测方法将被强调。额外的努力将检查凝胶电泳基质修饰和替代处理生物化学的影响。

Analysis methods for DNA genotyping and sequencing are linked into multiple-step processing systems that begin with DNA source material and extract genetic information. In conventional methods, the processing system operates on batches of microliter-volume reaction samples, together with matched liquid handling devices and electrophoretic analysis equipment. Project 2 will provide component-level design and testing our improved nanoliter-volume processing components for DNA sequencing and genotyping. To fulfill the goal of performing complex multi-step tasks on a single device, discrete drop sample handling and informational control will be required. New or improved fabrication technologies will also be examined as needed. All of the components developed in the Project retain the target of eventual integration in the complex systems of Project 1. Project 2 has three Specific Aims. 1) Develop a nanoliter injection and movement system. We will construct and test components for a robust and flexible injection and movement system that can handle multiple liquid samples in the volume range of 10 nl to 500 nl. Our successful injection and movement systems for single and binary drop systems will be modified and assembled into multiple- sample liquid handling procedures. Each discrete drop will be addressed and manipulated independently. 2) Develop multiple independent reaction chambers. We will construct and test reaction chambers for optimal surface interactions, temperature profiles, and thermal cross talk between reaction chamber components. A significant effort will involve developing control software and reproducible reaction conditions for high quality DNA sequencing. 3) Demonstrate high-resolution separations. We will improve the existing gel electrophoresis system (approximately 10-50 bp resolution, 500 bp length), to approach the quality of conventional large-scale DNA sequencing (1 bp resolution, 1000 bp length). Optimization of the existing fluorescent labeling and detection methods will be emphasized. Additional effort will examine gel electrophoresis matrix modifications and the effect of alternative processing biochemistries.