DNA sequencing by nanopore-induced photon emission.
DNA sequencing by nanopore-induced photon emission.
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DOI：
10.1007/978-1-61779-773-6_6
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发表时间：
2012-01-01
期刊：
影响因子：
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通讯作者：
Meller, Amit
中科院分区：
文献类型：
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作者：
Singer, Alon;McNally, Ben;Meller, Amit
基于纳米孔的 DNA 分析是一个极具吸引力的研究领域，因为该方法简单，不仅能够探测单个分子，而且能够检测极少量的基因组物质。在这里，我们描述了一种利用光学检测的新型、基于纳米孔的单分子 DNA 测序系统的材料和方法。我们根据二进制代码转换目标 DNA，该代码可被带有两种荧光团的分子信标识别。然后使用固态纳米孔依次剥离信标，从而产生一系列可以用定制显微镜检测到的光子爆发。我们在读出阶段不使用任何酶；因此，我们的系统不受单个酶的高度可变的持续性、寿命和不准确性的限制，这些因素可能会阻碍通量和可靠性。此外，由于我们的系统使用纯光学读出，因此我们可以利用高端、宽视场成像设备同时记录多个纳米孔。这使得我们的系统能够非常简单地并行化到纳米孔阵列。
Nanopore-based DNA analysis is an extremely attractive area of research due to the simplicity of the method, and the ability to not only probe individual molecules, but also to detect very small amounts of genomic material. Here, we describe the materials and methods of a novel, nanopore-based, single-molecule DNA sequencing system that utilizes optical detection. We convert target DNA according to a binary code, which is recognized by molecular beacons with two types of fluorophores. Solid-state nanopores are then used to sequentially strip off the beacons, leading to a series of photon bursts that can be detected with a custom-made microscope. We do not use any enzymes in the readout stage; thus, our system is not limited by the highly variable processivity, lifetime, and inaccuracy of individual enzymes that can hinder throughput and reliability. Furthermore, because our system uses purely optical readout, we can take advantage of high-end, wide-field imaging devices to record from multiple nanopores simultaneously. This allows an extremely straightforward parallelization of our system to nanopore arrays.