High Throughput Single Molecule DNA Sequencing

描述（由申请人提供）：人类基因组计划花了几年时间才完成，但这只是一个时期的开始，在这个时期，医学诊断、法医学和发育生物学将需要大量的DNA和RNA序列信息。传统的测序技术在成本、速度和灵敏度方面存在局限性，对序列信息的需求远远超出了目前的获取能力。我们最近展示了一项新技术的首次原理实验证明，该技术将为DNA和RNA测序提供快速、低成本和高度并行的技术。该技术在DNA聚合酶将标记的dNTPs整合到固定的单个DNA分子中后，使用荧光标记核苷酸的单分子检测。与目前的测序方法相比，该技术的一个主要优势在于它能够从数百万个独立分子中并行获取序列信息。在这里，我们建议开发具有更长的读取长度和更高精度的单分子测序试剂和方法，最终达到NIH黄金标准的99.99%，同时将哺乳动物基因组测序的成本降低到10万美元以下。

DESCRIPTION (provided by applicant): The Human Genome Project took several years to complete, yet it is only the beginning of a period in which large amounts of DNA and RNA sequence information will be required for medical diagnostics, forensics, and developmental biology. Conventional sequencing technology has limitations in cost, speed, and sensitivity and the demand for sequence information far outstrips the current ability to obtain it. We recently demonstrated the first proof of principle experiments for a new technology that will provide a fast, low cost, and highly parallel technique for DNA and RNA sequencing. This technology uses single molecule detection of fluorescently labelled nucleotides after DNA polymerase incorporates labeled dNTPs into immobilized individual DNA molecules. A major advantage of this technique over current sequencing methods lies in its ability to obtain sequence information from millions of independent molecules in parallel. Here we propose to develop reagents and methods for single molecule sequencing runs with longer read lengths and higher accuracy, ultimately reaching the NIH gold standard of 99.99%, while reducing the cost of sequencing a mammalian genome to below $100,000.