High Accuracy Nanopore Sequencing

首席调查员/项目主任(最后、第一、中间)：Gundlach、Jens、H. 高精度纳米孔测序 项目总结 这项提议的目标是使纳米孔测序技术达到最大限度 潜力。在NHGRI的资助下，我们的团队在开发纳米孔方面发挥了关键作用 测序。我们首先设计了高度敏感的孔，为纳米孔测序铺平了道路。 MSPA提供单核苷酸拆分，然后通过MSPA控制DNA速度 用一种酶给出了纳米孔测序概念的第一个证据。在此应用程序中，我们 概述将克服纳米孔测序的最后障碍的技术，低碱基调用 精确度。 我们通过此应用程序申请资金，以提高单通道基本呼叫的准确性 纳米孔测序要好于99%以上一个数量级。这个目标是一个 纳米孔测序向前迈出了革命性的一步，因为它开启了无处不在的应用 这项技术在医疗保健和其他领域都是如此。此外，我们还将探索其他改进措施和 使纳米孔测序成本更低、更有用的变化。 我们的具体目标是：(1)实现DNA的混合电压酶控制，使DNA抖动 从而允许识别和纠正酶错误模式，同时 最大化从离子电流读数中获得的信息，(2)改善基本 通过(I)改善用于测序的孔和(Ii) 与解旋酶专家合作开发DNA控制酶，专门设计成 优化纳米孔测序质量，(3)开发允许高集成密度的技术 使用纳米孔来增加测序吞吐量，并将DNA输入量降至最低。 我们团队到目前为止的成功使我们能够与著名的合作者和 得到学术界和工业界众多优秀实验室的支持，他们的专业知识帮助我们 将纳米孔测序带入医疗保健领域。我们将与合作伙伴实验室合作完成 本提案中概述的目标。 我们的目标是通过提供革命性的测序技术来提高人类的生活质量 这是准确、快速和廉价的。 项目摘要

Principal Investigator/Program director (Last, first, middle): Gundlach, Jens, H. High Accuracy Nanopore Sequencing PROJECT SUMMARY The objective of this proposal is to enable nanopore sequencing technology to reach its fullest potential. With NHGRI funding, our group has played a pivotal role in developing nanopore sequencing. We paved the way to nanopore sequencing by first engineering the highly sensitive pore MspA to provide single-nucleotide resolution and then by controlling the DNA speed through MspA with an enzyme to give the first proof of concept of nanopore sequencing. In this application we outline techniques that will overcome the last hurdle to nanopore sequencing, low base calling accuracy. We request funding through this application to increase the single-passage base calling accuracy of nanopore sequencing by more than an order of magnitude to better than 99%. This goal is a transformative step forward for nanopore sequencing since it opens up ubiquitous applications of this technology in health care and beyond. Additionally, we will explore other improvements and variations that make nanopore sequencing less expensive and more useful. Our specific aims are: (1) implementing a hybrid-voltage-enzyme control of DNA that dithers DNA back and forth, thereby allowing identification and correction of enzyme error modes, while maximizing the information obtainable from the ion current readings, (2) improving the fundamental components of the nanopore sequencer by (i) improving the pore used for sequencing and (ii) teaming up with helicase experts to develop DNA-controlling enzymes specifically engineered to optimize nanopore sequencing quality, (3) developing techniques that permit high integration density of nanopores to increase sequencing throughput and to minimize DNA input quantity. Our team's success to date has enabled us to form partnerships with prominent collaborators and to gain support from many excellent labs in academia and industry, whose expertise assists us in bringing nanopore sequencing into health care. We will work with our partner labs to complete the aims outlined in this proposal. It is our goal to improve the quality of human life by delivering revolutionary sequencing technology that is accurate, fast and inexpensive. Project Summary