99.99% Accuracy Direct DNA Sequencing via the Protein Nanopore Method

99.99%%20准确度%20直接%20DNA%20测序%20via%20the%20蛋白质%20纳米孔%20方法

• 批准号: 8318888
• 负责人: Geoffrey Barrall
• 金额: $ 47.24万
• 依托单位: ELECTRONIC BIOSCIENCES, INC.
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-01 至 2013-12-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8318888
• 关键词: Algorithms; Amino Acid Sequence; Award; Back; Base Sequence; Basic Science; Biological Process; Code; Complex; Custom; DNA; DNA Modification Process; DNA Sequence; Data; Data Analyses; Devices; Diagnostic; Diffusion; Electrolytes; Electronics; Elements; Error Sources; Forensic Medicine; Genome; Grant; Hemolysin; Integral Membrane Protein; Link; Lipid Bilayers; Location; Measurement; Measures; Medical; Methods; Modeling; Modification; Motion; Mutation; Noise; Peptide Sequence Determination; Performance; Polymers; Pore Proteins; Process; Proteins; Protocols documentation; Relative (related person); Research; Sampling; Scheme; Signal Transduction; Single-Stranded DNA; Site; Site-Directed Mutagenesis; Structure; System; Technology; Testing; Variant; base; biological research; cost; insight; instrumentation; mathematical model; nanopore; operation; programs; reconstruction; research study; response; simulation; statistics; voltage

Project Summary Our aim is to develop a DNA sequencing device based upon the blockade of ionic current in the transmembrane protein pore ¿-hemolysin (¿HL). Compared to other proposed nanopore-based approaches, the protein pore current blockade (PPCB) method is arguably the simplest, both conceptually and technologically, but recently has been passed over in favor of more complex methods. Recent electronic readout and bilayer lipid membrane advances by the proposers have greatly alleviated prior signal-to-noise ratio and robustness issues. New experimental data and an accurate Stochastic Model for DNA Motion (SMDM) within ¿HL now indicate threshold feasibility for sequencing by the PPCB method. Inspection of calculated SMDM responses to known input DNA sequences shows that the principal issue for sequencing via the PPCB method is random variance in the order the bases pass through the pore, leading to three quantifiable sources of error. This program will make specific structural and measurement parameter modifications to the present apparatus to reduce each type of error and to produce a minimal overall sequencing error. The final apparatus will be evaluated by sequencing up to 30 Mbases of natural DNA.

项目概要 我们的目标是开发一种基于离子电流阻断的 DNA 测序装置 跨膜蛋白孔 ¿-溶血素 (¿HL)。与其他提出的基于纳米孔的 方法中，蛋白质孔电流阻断 (PPCB) 方法可以说是最简单的，既 在概念和技术上，但最近已被忽略，取而代之的是更复杂的 方法。提案者最近在电子读出和双层脂质膜方面取得的进展 极大地缓解了先前的信噪比和鲁棒性问题。新的实验数据 以及 ¿HL 内准确的 DNA 运动随机模型 (SMDM) 现在指示阈值 PPCB方法测序的可行性。 检查计算出的 SMDM 对已知输入 DNA 序列的响应表明 通过 PPCB 方法进行测序的主要问题是碱基顺序的随机方差 通过孔隙，导致三个可量化的误差源。这个程序将使 对本装置的具体结构和测量参数进行修改 减少每种类型的错误并产生最小的总体测序错误。决赛 设备将通过对多达 30 Mbase 的天然 DNA 进行测序来进行评估。