NON CONTACT INFRARED MEDIATED THERMOCYCLING FOR CHIP PCR

描述(改编自《调查者摘要》)：映射和 组成DNA的大约3×10exp9碱基对的测序 人类基因组是人类基因组中承担的巨大任务 主动性(HGI)。而分子生物学技术的进步使 这是可能的，测序方案中的方法学“瓶颈” 是因为有必要通过以下方式分离测序片段 常规平板凝胶电泳法。因此，出现了激烈的 有兴趣用新的方法来加强传统的平板凝胶法 可以提高测序吞吐量的技术。两个相对较新的 电泳法、毛细管电泳法和电泳法 微制造的“芯片”已经显示出提供这项技术的希望。 尽管存在一些固有的技术挑战，但超快执行的能力 刻蚀到平板玻璃表面的多个通道中的分离 或塑料基质可显著提高测序速度。然而， 芯片平台的真正力量不仅来自于 快速的电泳分离，但从无缝的能力 将样品制备集成到芯片中。这将允许直接和 在一台设备中实现了聚合酶链式反应和电泳法的自动对接。 这项提议的目的是开发一种新的热循环方法 与其他方法不同，它可以很容易地集成到芯片中 站台。聚合酶链式反应将使用基于红外线的热循环进行 一种利用钨灯作为简单但有效的加热的技术 输送系统，配合溶液的红外探测/控制 温度。结合起来，这将允许在芯片上执行PCR 在不接触芯片的情况下，具有纳升体积。因此，成本- 与传统的聚合酶链式反应相比，时间效率有了显著的提高。 方法论。这一发展将为快速对接奠定基础 循环测序反应与先决条件的样品清理过程和 聚合酶链式反应后的电泳分离，全部放在一个一次性的 微量分析装置。

DESCRIPTION (Adapted from the Investigator's Abstract): The mapping and sequencing of the approximately 3 X 10exp9 base pairs of DNA that constitute the human genome is the colossal task undertaken in the Human Genome Initiative (HGI). While advances in molecular biological techniques make this possible, a methodological "bottleneck" in the sequencing protocol has been created by the necessity to separate sequencing fragments via conventional slab gel electrophoresis. Consequently, there has been intense interest in augmenting the conventional slab gel-based methodology with new technologies that can enhance sequencing throughput. Two relatively new electrophoretic formats, capillary electrophoresis and electrophoresis in microfabricated "chips," have shown promise for providing this technology. Despite some inherent technical challenges, the ability to execute ultrafast separations in multiple channels etched into the surface of a planar glass or plastic substrate could enhance sequencing speed significantly. However, the true power of the chip platform stems, not only from the capability for rapid electrophoretic separations, but from the ability to seamlessly integrate sample preparation into the chip. This would allow for direct and automated interfacing of PCR and electrophoresis in a single device. The object of this proposal is to develop a novel thermocycling approach that, unlike other approaches, can easily be integrated into the chip platform. PCR will be carried out using an infrared-based thermocycling technology that exploits a tungsten lamp as a simple but effective heat delivery system, coupled with infrared detection/control of solution temperature. Combined, this will allow for PCR to be executed on the chip with nanoliter volumes without contacting the chip. Consequently, the cost- and time-efficiency of PCR will be improved dramatically over conventional methodology. This development will set the stage for interfacing rapid cycle sequencing reactions with the prerequisite sample clean-up process and the post-PCR electrophoretic separation, all in a single, disposable microanalysis device.