Optimization of Nanopore Genomic DNA Sequencing

纳米孔基因组 DNA 测序的优化

• 批准号: 8749195
• 负责人: MARK A AKESON
• 金额: $ 76.06万
• 依托单位: UNIVERSITY OF CALIFORNIA SANTA CRUZ
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-08-01 至 2017-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8749195
• 关键词: Animal Model; Arabidopsis; Basic Science; Biological; Cell Nucleus; Cells; Communities; Computing Methodologies; CpG dinucleotide; Cytosine; DNA; DNA Damage; DNA Resequencing; DNA Sequence; Development; Devices; Enzymes; Epigenetic Process; Escherichia coli; Genomic DNA; Goals; Healthcare; Heterogeneity; Human; Individual; Length; Libraries; Malignant Neoplasms; Measurable; Medical Research; Methods; Methylation; Modification; Motor; Mus; Noise; Nucleotides; Organism; Proteins; Reading; Reference Standards; Regenerative Medicine; Reporting; Research; System; Techniques; Technology; Testing; Time; Touch sensation; base; biophysical properties; design; helicase; improved; motor control; mutant; nanopore; nanoscale; nucleobase; public health relevance; sensor; single molecule

This proposal concerns optimization of enzymes, pores, and computational methods for single molecule sequencing of genomic DNA fragments. It is based on a proven nanopore device implemented by our group at UCSC. This device is comprised of a sensor that touches and examines each nucleotide within a captured DNA strand as a processive enzyme motor advances the strand. Although the overall goal of nanopore sequencing is de novo reads on very long strands, here we will also focus on resequencing of DNA from organisms important in basic research (mouse, E. coli & Arabidopsis) and in healthcare (human). We are focusing on both de novo and resequencing for two reasons: 1) nanopore sequencing of biological DNA has not been documented publicly. Therefore, nanopore resequencing of reference standards is required for community acceptance, and, importantly, to reveal weaknesses in the technology that impact de novo sequencing accuracy; 2) nanopore resequencing in this application means reading genomic DNA directly and therefore will include epigenetic modifications. This would be an immediate, important contribution to the research community.

该提案涉及单分子的酶、孔和计算方法的优化 基因组 DNA 片段的测序。它基于我们实施的经过验证的纳米孔设备 UCSC 的小组。该设备由一个传感器组成，可以接触并检查内的每个核苷酸。 捕获的 DNA 链作为持续酶​​马达推进该链。虽然总体目标是 纳米孔测序是在很长的链上进行从头读取，这里我们还将重点关注 来自对基础研究（小鼠、大肠杆菌和拟南芥）和医疗保健具有重要意义的生物体的 DNA （人类）。我们关注从头测序和重测序有两个原因：1) 纳米孔测序 生物 DNA 的含量尚未公开记录。因此，参考纳米孔重测序 社会接受需要标准，而且更重要的是，要揭示标准中的弱点 影响从头测序准确性的技术； 2)本应用中的纳米孔重测序 意味着直接读取基因组 DNA，因此将包括表观遗传修饰。这将是 对研究界的直接、重要贡献。