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Millikan Sequencing by Label-Free Detection of Nucleotide Incorporation

通过无标记检测核苷酸掺入进行密立根测序

基本信息

批准号：
8546475
负责人：
Javier Farinas
金额：
$ 10.73万
依托单位：
CAERUS MOLECULAR DIAGNOSTICS, INC.
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-09-01 至 2013-06-30
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): The feasibility of a label-free technology, Millikan Sequencing, will be evaluated for de novo sequencing of mammalian genomes for under $1,000. This novel sequencing-by-synthesis approach measures the increased charge as nucleotides are added to DNA templates attached to a tethered bead. Opposing electrical, hydrodynamic and entropic forces will be used to measure the bead displacement, which is a function of the length of DNA attached to the bead. Simultaneous detection of an array of millions of beads undergoing chain elongation will allow high-throughput sequencing. Model calculations and preliminary results indicate that this method should enable accurate, long read length and label-free DNA sequencing. The lack of labels leads to negligible reagent costs while the relatively simple optics leads to a low-cost instrument. Long read lengths will result in low genome assembly cost. The much lower per-bead copy number required compared to the 454 system should enable amplification options other than emulsion PCR, such as bridge PCR, making initial sample preparation easier and cheaper. Ultimately, the method could be used on single molecules thereby further reducing sample preparation costs. The aims of the proposed two-year exploratory project are: (1) to demonstrate the ability to sequence DNA using a single tethered bead, and (2) to develop a scheme that would allow simultaneous detection of large bead arrays for high throughput analysis. PUBLIC HEALTH RELEVANCE: Nucleic acid sequence information is critical to medical research and to basic biological studies. The goal of this project is to develop a DNA sequencing system based on a label-free detection approach, Millikan Sequencing, which is capable of de novo sequencing of mammalian genomes for under $1,000. Such a platform will allow researchers and clinicians to perform the translational research ultimately required to improve human health.

描述（由申请人提供）：将评估无标记技术密立根测序的可行性，用于以低于 1,000 美元的价格对哺乳动物基因组进行从头测序。这种新颖的合成测序方法可测量当核苷酸添加到附着在系留珠子上的 DNA 模板时增加的电荷。相反的电力、流体动力和熵力将用于测量珠子位移，这是附着在珠子上的 DNA 长度的函数。同时检测数百万个正在进行链延长的珠子阵列将允许高通量测序。模型计算和初步结果表明，该方法应该能够实现准确、长读长和无标记的 DNA 测序。缺少标签导致试剂成本可以忽略不计，而相对简单的光学器件则导致仪器成本低廉。长读长将导致较低的基因组组装成本。与 454 系统相比，所需的每珠拷贝数要低得多，因此可以实现乳液 PCR 以外的扩增选项，例如桥式 PCR，从而使初始样品制备变得更容易、更便宜。最终，该方法可以用于单分子，从而进一步降低样品制备成本。拟议的为期两年的探索性项目的目标是：(1) 展示使用单个系留珠子对 DNA 进行测序的能力，以及 (2) 开发一种方案，允许同时检测大型珠子阵列以进行高通量分析。公共卫生相关性：核酸序列信息对于医学研究和基础生物学研究至关重要。该项目的目标是开发一种基于无标记检测方法密立根测序的 DNA 测序系统，该系统能够以不到 1,000 美元的价格对哺乳动物基因组进行从头测序。这样的平台将使研究人员和临床医生能够进行改善人类健康最终所需的转化研究。