Microfluidic DNA Sequencing

微流控 DNA 测序

基本信息

批准号：
7725510
负责人：
TAL RAZ
金额：
$ 24万
依托单位：
GNUBIO, 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 2011-08-31
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): Most next-generation DNA sequencing methods have focused on either 1) template amplification followed by massively-parallel sequencing-by-synthesis, or 2) single molecule detection. The first method is now commercially available but suffers from relatively large volumes of expensive reagent usage. The latter method, although not yet commercially available, will have a disadvantage in signal-to-noise and therefore require more sensitive and expensive instrumentation. To avoid these disadvantages, we will use droplet-based microfluidics to sequence DNA. By using microfluidics we limit the amount of reagent required to sequence DNA to less than several milliliters, while still retaining the ability to amplify the template that thereby enables us to use relatively inexpensive and robust detection. Hybridization of short probes will be detected in microfluidic droplets by a shift in fluorescence polarization that distinguishes between bound and free oligo. This removes the requirement for a separation phase to detect hybridization. The method is simple and does not require enzymes. In Phase I we will describe a simple platform and resequencing method that will be scaled in a future Phase II project to enable human genome sequencing for under $1000. Public Health Relevance: The proposed Phase I droplet-based microfluidics instrument will generate, selectively merge and analyze over 10,000 aqueous droplets per second. We want to leverage this capability to implement a DNA sequencing method on a microfluidics platform. We have broken the project up into two phases. Phase I is proof of principle. In Phase I we demonstrate that the biochemical assay works in drops and also that our detector works. In Phase II we will scale the process to build a machine that will be able to sequence a human genome for less than $1000.

描述（由申请人提供）：大多数下一代DNA测序方法都集中在1）模板放大，然后是大规模平行的测序，或2）单分子检测。现在，第一种方法可以在市售中获得，但遭受了相对较大的昂贵试剂使用量。后一种方法虽然尚未在市售中可用，但在信噪比方面将有缺点，因此需要更敏感和昂贵的仪器。为了避免这些缺点，我们将使用基于液滴的微流体来序列DNA。通过使用微流体，我们将序列DNA序列DNA所需的试剂量限制为小于几毫升，同时仍保持放大模板的能力，从而使我们能够使用相对便宜且可靠的检测。在微流体液滴中，将通过区分结合寡核能的荧光极化，将在微流体液滴中检测到短探针的杂交。这消除了分离阶段检测杂交的要求。该方法很简单，不需要酶。在第一阶段，我们将描述一种简单的平台和重新方便的方法，该方法将在未来的II阶段项目中进行缩放，以使人类基因组测序以低于$ 1000的价格进行。公共卫生相关性：拟议的基于I液滴的微流体仪器将产生，选择性合并并分析每秒10,000多个水滴。我们希望利用这种能力在微流体平台上实现DNA测序方法。我们已经将项目分为两个阶段。第一阶段是原则的证明。在第一阶段，我们证明了生化测定在滴剂中起作用，并且我们的检测器有效。在第二阶段，我们将扩展过程以构建能够以少于1000美元的价格对人类基因组进行测序的机器。