Real-time DNA Sequencing:Nucleotide Synthesis and Use

实时 DNA 测序：核苷酸合成和使用

• 批准号: 6338382
• 负责人: Susan H Hardin
• 金额: $ 9.69万
• 依托单位: VISIGEN BIOTECHNOLOGIES, INC.
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-06-01 至 2003-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6338382
• 关键词: DNA; adenosine triphosphate; fluorescent dye /probe; method development; nanotechnology; nucleic acid chemical synthesis; nucleic acid sequence; nucleotides; protein engineering; time resolved data

DESCRIPTION (Applicant's abstract): A first step toward the development of a method to sequence a single DNA molecule in real-time is proposed. Engineering a polymerase to act as a direct molecular sensor of DNA base identity, coupled with the synthesis of appropriately modified nucleotides, allows us to create the fastest enzymatic DNA sequencing system possible. Our motivation for developing a single-molecule sequencing approach includes eliminating the intermediate steps currently necessary for determining DNA sequence information, thereby meeting the needs of both the genomics and the medical diagnostics communities. We outline a series of milestones towards our ultimate goal of a single-molecule DNA sequencing system. However, only two of these intermediate goals are the specific aims of the proposed research. In particular, we propose to synthesize an appropriately modified nucleotide and to determine the relative efficiency at which commercially available DNA polymerases use this modified nucleotide to extend a DNA strand. PROPOSED COMMERCIAL APPLICATION: Development of a single-molecule sequencing system is generally applicable for DNA diagnostics. Furthermore, parallel processing of samples will allow a genome to be sequence in a day or less, and will be important for pathogen identification and large-scale, comprehensive genome analysis. This system eliminates sequencing reaction processing, gel or capillary loading, and electrophoresis, promising huge savings in time, labor, and cost per base. This single-molecule DNA sequencing system is expected to replace current DNA sequencing technology.

描述（申请人的摘要）：迈向开发的第一步 提出了实时测序单个DNA分子的方法。工程 作为DNA碱基身份的直接分子传感器的聚合酶，耦合 通过合成适当修饰的核苷酸的合成，使我们能够创建 最快的酶促DNA测序系统可能。我们的动力 开发单分子测序方法包括消除 当前确定DNA序列所需的中间步骤 信息，从而满足基因组学和医学的需求 诊断社区。我们概述了一系列的里程碑，以实现我们的最终 单分子DNA测序系统的目标。但是，其中只有两个 中间目标是拟议研究的具体目标。在 特别是，我们建议合成适当修饰的核苷酸和 确定可商购DNA的相对效率 聚合酶使用这种修饰的核苷酸延伸DNA链。 拟议的商业应用： 单分子测序系统的开发通常适用于DNA 诊断。 此外，样品的平行处理将允许基因组为 序列在一天或更短的时间内，对于病原体鉴定和 大规模，全面的基因组分析。 该系统消除了测序 反应加工，凝胶或毛细管负荷和电泳，有望巨大 节省时间，劳动和每个基础成本。 这种单分子DNA测序 系统有望替代当前的DNA测序技术。