Advanced Parallel Readers for DNA Sequencing Through a 2D Nanopore

用于通过 2D 纳米孔进行 DNA 测序的高级并行读取器

• 批准号: 10676761
• 负责人: Marija Drndic
• 金额: $ 15.08万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-08-04 至 2025-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10676761
• 关键词: 3-Dimensional; Advanced Development; Architecture; Biological; Buffers; Calibration; Complex; Custom; DNA; DNA Microarray Chip; DNA Sequence; DNA sequencing; Data Analyses; Detection; Development; Devices; Diagnosis; Diameter; Electrodes; Electrolytes; Entropy; Enzymes; Face; Finite Element Analysis; Freedom; Genetic; Geometry; Glass; Goals; Grant; Height; Image; Ions; Machine Learning; Measurement; Measures; Membrane; Methods; Modeling; Molecular; Motion; Noise; Nucleotides; Optics; Patients; Photons; Polymerase; Procedures; Rationalization; Reader; Research; Resistance; Route; Running; Signal Transduction; Silicon; Single-Stranded DNA; Sodium Chloride; Speed; Stochastic Processes; Symptoms; System; Techniques; Technology; Temperature; Testing; Thick; Thinness; Time; United States National Institutes of Health; base; cost; design; detection platform; disorder prevention; ds-DNA; electronic sensor; experience; fluorophore; improved; nanometer; nanopore; nanoscale; novel strategies; operation; pragmatic implementation; rapid detection; sequencing platform; silicon nitride; single molecule; solid state; temporal measurement; transcriptome sequencing; voltage

Project Summary To improve DNA and RNA sequencing with respect to accuracy, robustness and speed, this NIH R21 project focuses on using a two-layer design with two parallel solid-state SiN-2D nanopores on low-noise all-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 ultrathin SiN membranes, as well as 2D membranes, improve the signal- to-noise ratio for molecular detection and analysis because the resistance to the ionic flow through a nanopore increases linearly with the nanopore 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 important components: we propose to demonstrate a two-layer on- chip solid-state SiN-2D-pore system that limits the range of DNA motion through two parallel proximal pores that are electrically independently addressable. We create devices containing a second layer with one silicon nitride (SiN) pore, parallel to a primary layer containing the atomically-thin 2D pore that confine ssDNA within a device to a restricted geometry, yet allow the free motion of salt ions to maintain a high signal-to-noise ratio. We propose a specific two-layer concept, where the two layers are in close proximity, 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 which we refer to as the main sensing/sequencing 2D nanopore. A secondary layer has a second pore sharing the same electrode pair as the sensing pore, but also having its own independent electrode pair to be probed separately. Although we have two pores, they can operate as a continuous system due to their proximity. We outline the 3D finite element analysis modeling and practical implementation (two versions) of these concepts with Si-based technology, including advantages and challenges involved for DNA (and biomolecule) sequencing (analysis) in solution. Our approach eliminates the need for any enzymes and enables DNA and biomolecules to be guided through robust and long-lasting nanopores, facilitated by the custom- designed chip combining the best of what the SiN and 2D pores can currently offer. Illustration 1: Proposed two-layer device concept for this NIH R21 proposal, relying on minimization of DNA entropic motion using two proximal, parallel SiN-2D pores that are electrically independently contacted: a guiding SiN pore of variable diameter and an optimized sensing 2D materials pore. The spacing between the two layers is adjustable down to a few nm (facilitated by the single nm control of RIE or TEM etching). This Si platform is versatile and compatible with any 2D materials (shown as green triangle). 1

项目摘要 为了改善准确性，鲁棒性和速度的DNA和RNA测序，该NIH R21项目专注于使用两个平行固态SIN-2D纳米孔的两层设计 在低噪声全玻璃芯片上，朝着DNA测序和直接的RNA测序。基本 纳米孔背后的概念涉及使用施加的电压驱动单链DNA 分子通过狭窄的纳米孔，该纳米孔分离了电解质溶液的腔室。这 电压还驱动电解质离子流过孔，以电流测量。 当分子穿过纳米孔时，它们会修改离子的流动和结构 可以通过分析所得电流的持续时间和幅度来提取信息 减少。超薄SIN膜中的纳米孔以及2D膜，改善了信号 - 用于分子检测和分析的待命比率，因为对离子流的抗性 纳米孔随纳米孔厚度线性增加，因此两个离子的幅度 电流和阻塞电流随易位分子的增加而随着纳米孔的降低而增加 高度。具体而言，我们试图制作固态基于离子电流的纳米孔测序 通过结合几个重要组成部分：我们建议证明两层的on- 芯片固态SIN-2D孔系统，该系统限制了DNA运动的范围通过两个平行 近端毛孔是可独立寻址的。我们创建包含一个的设备 第二层含有氮化硅（SIN）孔，平行于一层包含的主要层 将ssDNA限制在设备内的原子薄2D孔与受限的几何形状，但允许 盐离子的自由运动以保持高信噪比。我们提出了一个特定的两层 概念，两个层紧邻，具有两个独立的电气连接， 以及相应的芯片设备体系结构以实现此目标。在这种方法中，有一个中心 高度敏感的2D孔，我们称为主要感应/测序2D纳米孔。一个 次级层具有与传感孔相同的电极对的第二个孔，但也 具有自己的独立电极对，要分别探测。尽管我们有两个毛孔，但 由于其接近性，它们可以作为连续系统运行。我们概述了3D有限元 这些概念的分析建模和实际实施（两个版本）具有基于SI的概念 技术，包括DNA（和生物分子）测序涉及的优势和挑战 （分析）解决方案。我们的方法消除了对任何酶的需求，并启用DNA和 生物分子可以通过强大而持久的纳米孔来指导，并由定制促进 设计的芯片结合了目前可以提供的罪恶和2D毛孔。 插图1：提议的两层 NIH R21的设备概念 提案，依靠最小化 DNA熵运动使用两个 近端，平行的SIN-2D孔 是独立的 联系：一个指导罪的孔 可变直径和优化 感应2D材料孔。这 两层之间的间距是 可调至几nm （通过单个NM控制的促进 rie或tem蚀刻）。这个SI平台 用途广泛，并且与任何 2D材料（如绿色显示 三角形）。 1