Instrument to Optimize DNA Sequencing by Recognition Tunneling

通过识别隧道优化 DNA 测序的仪器

• 批准号: 8542509
• 负责人: STUART LINDSAY
• 金额: $ 99.27万
• 依托单位: ARIZONA STATE UNIVERSITY-TEMPE CAMPUS
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-08-15 至 2015-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8542509
• 关键词: Automobile Driving; Base Sequence; Basic Science; Binding; Buffers; Carbon Nanotubes; Chemicals; Clinical; Code; Collaborations; DNA; DNA Sequence; Data; Devices; Electrodes; Electrolytes; Electronics; Electrons; Environment; Epigenetic Process; Future Generations; Genome; Goals; Gold; Grant; Individual; Ions; Kinetics; Laboratories; Letters; Libraries; Liquid substance; Maps; Measurement; Measures; Mechanics; Metals; Modeling; Modification; Nanotechnology; Nature; Nucleosides; Nucleotides; Organic solvent product; Positioning Attribute; Preparation; Process; Production; Reader; Reading; Reagent; Running; Scanning Probe Microscopes; Scheme; Signal Transduction; Single-Stranded DNA; Speed; Surface; Techniques; Testing; Time; Tissues; Water; Water Pollution; Work; analytical tool; aqueous; base; computing resources; cost; data acquisition; design; improved; instrument; multi-scale modeling; nanofluidic; nanopore; nanoscale; news; next generation; quantum; residence; simulation; single molecule; single walled carbon nanotube

DESCRIPTION (provided by applicant): Nanopore sequencing is a technique in which DNA is driven electrophoretically through an orifice so small that each base must pass through one at a time. Translocation of thousands of bases of single stranded DNA has been demonstrated. If such long sequence runs could be read rapidly and accurately with no need for chemical reagents or the preparation of elaborate libraries, costs might be reduced to the point where personal genomes would become available for clinical use. Readouts based on the blockading of ion current have been able to resolve individual nucleotides and a single base trapped at a double-single strand junction in a hairpin but have not been able to read along a DNA molecule continuously. Very recently, we have shown that it is possible to identify individual bases and read along a DNA molecule using a technique we call Recognition Tunneling. Recognition molecules, covalently bound to electrodes, are used to transiently trap each base in turn through noncovalent bonds, giving distinct electronic signatures of all four bases and 5-methyl C. The trapping time with no external force applied to the DNA is long (seconds). However, unbinding is readily accelerated to very short times by the application of small forces, so Recognition Tunneling also provides a straightforward approach to translocation control. Here, we propose to combine Recognition Tunneling with nanopore translocation using metal or graphene nanopores, and metal or carbon nanotube reading electrodes, the probes and pores both being functionalized with recognition molecules. We will study translocation-control in functionalized, conducting nanopores, using both the bias across the pore and the surface potential of the conducting pore as control signals. Using a scanning-tunneling microscope (STM) platform, we will make measurements of Recognition Tunneling signals as DNA emerges from the nanopore. Multiscale (quantum to fluid-mechanical) simulations at Oak Ridge National Laboratory will help us to understand and optimize the translocation and readout processes. This understanding will be shared with collaborators who are developing nanopores with fixed (as opposed to STM) reading schemes with the ultimate goal of producing sequencing chips that are cheap and contain many thousands of devices.

描述（由申请人提供）：纳米孔测序是一种通过电泳驱动 DNA 通过小孔的技术，该孔非常小，以至于每个碱基一次必须通过一个孔。已经证明了单链 DNA 的数千个碱基的易位。如果可以快速准确地读取如此长的序列，而不需要化学试剂或准备复杂的文库，那么成本可能会降低到个人基因组可用于临床的程度。基于离子电流阻断的读数已经能够解析发夹中双单链连接处捕获的单个核苷酸和单个碱基，但无法连续读取 DNA 分子。最近，我们已经证明，使用我们称为“识别隧道”的技术可以识别单个碱基并沿着 DNA 分子进行读取。识别分子与电极共价结合，通过非共价键依次瞬时捕获每个碱基，给出所有四个碱基和 5-甲基 C 的独特电子特征。在不向 DNA 施加外力的情况下，捕获时间很长（秒） ）。然而，通过施加较小的力，解绑很容易加速到非常短的时间，因此识别隧道还提供了一种直接的易位控制方法。在这里，我们建议将识别隧道与使用金属或石墨烯纳米孔以及金属或碳纳米管读取电极的纳米孔易位结合起来，探针和孔都用识别分子功能化。我们将研究功能化导电纳米孔中的易位控制，使用跨孔的偏压和导电孔的表面电位作为控制信号。当 DNA 从纳米孔中出现时，我们将使用扫描隧道显微镜 (STM) 平台测量识别隧道信号。橡树岭国家实验室的多尺度（量子到流体力学）模拟将帮助我们理解和优化易位和读出过程。这种理解将与正在开发具有固定（而不是 STM）读取方案的纳米孔的合作者分享，其最终目标是生产廉价且包含数千个设备的测序芯片。