Multilayer Device for Sequencing DNA Through a Solid-State Nanopore

项目摘要 提高DNA测序的准确性、稳健性和速度，并 开发实用的RNA测序方法，NIH R43第一阶段项目重点使用 低噪声玻璃芯片上的多层设计固态孔洞传感器，用于DNA测序 和直接RNA测序。纳米孔背后的基本概念涉及使用应用的 驱动单链DNA分子通过狭窄的纳米孔的电压，该纳米孔分离 电解液的腔室。该电压还驱动电解液离子流过 孔洞，用电流来测量。当分子穿过纳米孔时，它们会被修饰 离子的流动和结构信息可以通过分析持续时间和 由此产生的电流减少的幅度。原子薄2D膜中的纳米孔 提高分子检测和分析的信噪比，因为对 流过孔洞的离子流量随着孔洞厚度线性增加，因此 随着分子移位的增加，离子电流和阻挡电流的增加 纳米孔高度。具体地说，我们试图制造基于离子电流的固态纳米孔 通过组合几个组件来制造模块化多层片上可以进行排序 固态超薄孔系统，限制DNA在感应区的运动范围 毛孔。我们通过制造包含第二层氮化硅空穴的器件来做到这一点，这些空穴平行于 包含传感2D孔的初级层，该孔将设备内的DNA定向到受限的 几何形状，但允许离子的自由运动，以保持高信噪比。我们提出了一个 具有两个独立电气连接的特定多层概念以及相应的芯片 设备体系结构来实现这一目标。在这种方法中，存在一个中心的、高度敏感的2D 毛孔。第二层是共享相同电极对的纳米孔阵列(NPA) 感测2D气孔。这些小孔平行于“感应”小孔构造，起到了 “喂食”元素以伸展DNA并将其送入感应孔。我们概述了实用的 利用硅基技术实现这一概念，包括对DNA(和 生物分子)在溶液中测序(分析)。我们的方法不再需要任何 酶，并使DNA分子能够被引导通过坚固和持久的纳米孔， 由定制设计的“阵列芯片”促成，并有可能以创纪录的高测序速度进行。 图1：建议的多层设备 NIH R43第一阶段提案的概念， 依赖于DNA熵的最小化 运动：一种导向阵列和优化的二维 毛孔。 1

Project Summary To improve DNA sequencing capabilities with respect to accuracy, robustness and speed and to develop practical methods of RNA sequencing, this NIH R43 Phase I project focuses on using multilayer-design solid-state pore sensors on low-noise glass chips, towards DNA sequencing and direct RNA sequencing. The basic concept behind nanopores involves using an applied voltage to drive single-stranded DNA molecules through a narrow nanopore, which separates chambers of electrolyte solution. This voltage also drives a flow of electrolyte ions through the pore, measured as an electric current. When molecules pass through the nanopore they modify the flow of ions, and structural information can be extracted by analysis of the duration and magnitude of the resulting current reductions. Nanopore in atomically-thin 2D membranes improve the signal-to-noise ratio for molecular detection and analysis because the resistance to the ionic flow through a pore increases linearly with the pore thickness, so both the magnitudes of the ionic current and the blocked current with a translocating molecule increase with decreasing nanopore height. Specifically, we seek to make solid-state ionic-current based nanopore sequencing possible by combining several components to make a modular multilayer on-chip solid-state ultrathin-pore system that limits the range of motion for DNA in the sensing region of a pore. We do so by creating devices containing a second layer of silicon nitride holes, parallel to primary layer containing the sensing 2D pore that orient DNA within a device to a restricted geometry, yet allow the free motion of ions to maintain a high signal-to-noise ratio. We propose a specific multilayer concept with two independent electrical connections, and corresponding chip device architecture to achieve this goal. In this method, there is a central, highly sensitive 2D pore. A secondary layer is a nanopore array (NPA) sharing the same electrode pair as the sensing 2D pore. These pores are constructed parallel to the “sensing” pore and serve as “feeding” elements to stretch and feed DNA into the sensing pore. We outline the practical implementation of this concept with Si-based technology, including advantages for DNA (and biomolecule) sequencing (analysis) in solution. Our approach eliminates the need for any enzymes and enables DNA molecules to be guided through robust and long-lasting nanopores, facilitated by the custom-designed “array chip”, and at potentially record high sequencing speeds. Illustration 1: Proposed multilayer device concept for this NIH R43 Phase I proposal, relying on minimization of DNA entropic motion: a guiding array and an optimized 2D pore. 1