Large-Scale Nanopore Arrays for DNA Sequencing

描述(申请人提供)：今天，合成纳米孔设备是作为对单个生物大分子的高速非酶询问的有趣候选设备而被密集研究的主题。例如，为了对未知成分的单个DNA片段进行高速测序，需要这种功能。虽然目前几个由美国国立卫生研究院资助的小组正在研究有关大分子移位物理和新型读出方案的许多理论方面，但纳米孔在测序系统中的实际应用也需要大量的纳米孔阵列并行工作，每个阵列都连接到高灵敏度的电子放大器/检测器。 这样的并行性和系统集成是必要的，以便与当今最先进的大规模并行合成方案(SBS)(例如454生命科学公司提供的方案，其报告的细菌基因组高保真测序速度为20Mb/天)相比，在吞吐量方面实现显著改进。为了说明对阵列的需求，可以考虑例如，平均纳米孔移位率为100b/ms/孔的100纳米孔阵列原则上可以以10 GB/天的(高保真，100重复)速率对序列片段进行排序，允许在一天内对整个人类基因组进行测序，速度比SBS快500倍。 这项提议的目的是(A)开发高度集成的纳米孔阵列，这种阵列可以通过光刻方法以及放大和隔离其各种电信号的芯片上基于硅的电子电路和电路技术来制造，以及(B)探索原则上可以区分来自每个DNA碱基的信号的偶极传感方法。一些制造出来的纳米孔中将加入一个正交电容传感器。纳米孔阵列将使用自对准成分方法构建在硅衬底上。将构建正交偶极矩检测器，以产生与DNA分子相对于电极的旋转无关的信号。

DESCRIPTION (provided by applicant): Today, synthetic nanopore devices are the subject of intense investigation as intriguing candidate devices for high-speed non-enzymatic interrogation of individual biological macromolecules. Such functionality is required for example in order to sequence individual DNA fragments of unknown composition at high speeds. While many theoretical aspects regarding macromolecule translocation physics and novel readout schemes are presently under investigation by several NIH-funded groups, the practical utilization of nanopores in sequencing systems also requires large arrays of nanopores working in parallel, each of these connected to high-sensitivity electronic amplifiers/detectors. Such degree of parallelism and systems integration is necessary in order to achieve significant improvements in throughput compared with today's state-of-the-art massively-parallel enzymatic sequencing by synthesis schemes (SBS) (such as those offered by 454 Life Sciences which reports high-fidelity sequencing of bacterial genomes at 20 Mb/day). To illustrate the need for arrays one may consider for example, a 100- nanopore array with average nanopore translocation rate of 100b/ms/pore could in principle sequence fragments at (high-fidelity, 100 repeats) rates of 10 Gb/day, allowing an entire human genome to be sequenced in a day, 500 times faster than SBS. The aims of this proposal are (a) the development of highly-integrated arrays of nanopores that can be fabricated by lithographic methods along with on-chip silicon-based electronic circuits and circuit techniques that amplify and isolate their various electrical signals, and (b) the exploration of a dipole sensing methodology which in principle can distinguish signals from each of the DNA bases. A quadrature capacitance sensor will be incorporated in some of the fabricated nanopores. Arrays of nanopores will be constructed on silicon substrates using a self-aligned compositional approach. Quadrature dipole moment detectors will be constructed that yield a signal independent of the rotation of the DNA molecule relative to the electrodes.