Tunnel Junction for reading all four bases with high discrimination

描述(申请人提供)：我们已经发现，对于所有四个核苷(和5-甲基脱氧胞苷)，可以使用一对隧道电极产生不同的隧道信号，该电极由含有氢键供体和氢键受体的简单试剂功能化。这项提议的目标是将测量范围扩大到水溶液中的核苷酸，然后再扩展到小分子低聚物。我们将量化单分子读数的比例，并确定控制这一比例的因素，目标是消除一次来自缺口中多个核苷酸的信号。我们将探索控制所有四个碱基(和5-甲基C)电流信号分布宽度的因素，目的是提高单一读数的识别率。我们将测量成功读取的比例，并表征复合体(产生信号)在隧道间隙中形成所需的时间。从这些测量中，我们将确定提高读出效率所需的改进，并为配备隧道电极的纳米孔测序系统的设计制定标准。在这项研究过程中开发的试剂将提供给其他开发使用电子隧道作为读数的纳米孔测序器的研究小组。 与公共健康相关：至少有七个由美国国立卫生研究院支持的组织正在探索测序方法，这些方法提议使用电子隧道作为纳米孔测序仪的读数，这种方法可能会大大降低测序成本。我们已经证明，所有四种核苷和5-甲基胞苷都可以被功能化电极读取，我们将开发适合于在水溶液中进行DNA测序的试剂，并使其得到广泛应用。

DESCRIPTION (provided by applicant): We have discovered that distinct tunneling signals can be generated for all four nucleosides (and 5-methyldeoxycytidine) using one pair of tunneling electrodes functionalized with a simple reagent containing a hydrogen-bond donor and a hydrogen bond acceptor. The goals of this proposal are to extend the measurements to nucleotides in aqueous electrolyte, and then to small oligomers. We will quantify the fraction of single-molecule reads and determine the factors that control this fraction with the goal of eliminating signals that come from more than one nucleotide in the gap at a time. We will explore the factors that control the width of the distribution of current signals for all four bases (and 5-methyl C) with the goal of improving the discrimination of a single read. We will measure the fraction of successful reads and characterize the time required for the complex (that gives rise to the signal) to form in the tunnel gap. From these measurements, we will identify improvements needed to increase the readout efficiency and also develop criteria for design of a nanopore sequencing system equipped with tunneling electrodes. The reagents developed during the course of this research will be made available to other research groups developing nanopore sequencers that use electron tunneling as the readout. PUBLIC HEALTH RELEVANCE: At least seven NIH-supported groups are exploring sequencing methods that propose to use electron tunneling as the readout for a nanopore sequencer, an approach that might greatly reduce the cost of sequencing. We have shown that all four nucleosides and 5-methyl cytidine can be read by functionalized electrodes and we will develop reagents suitable for DNA sequencing in aqueous electrolyte and make these widely available.