Detecting diverse nucleic acid biomarkers of cancer with solid-state nanopores

项目摘要 核酸生物标记物对患者的健康具有巨大的潜在价值，但由于它们的性质，往往是困难的。 去探查。固态(SS-)纳米孔具有独特的定位，有助于为这一挑战提供解决方案。这个 平台使单个分子能够在它们通过合成的、 纳米级光圈。固有的灵敏度、紧凑性和电子输出使系统成为一种 作为平移诊断设备的极具吸引力的候选者。此R33项目的中心目标是使用 SS纳米孔技术用于评估核酸生物标记物及其在癌症中的应用 我们实验室开发的选择性分析方法。利用我们的新方法，可以检测到目标核酸片段 并且仅当与蛋白质伴侣结合时才被量化，从而产生二进制输出。测量速度很快， 灵敏，并产生明确的电信号以供分析。我们建议将这一将军 能够评估两个广泛重要的核酸家族的测量方案 生物标志物。在目标1中，我们将评估表观遗传修饰和单碱基损伤。修饰碱基调节 多种细胞功能，但用传统技术进行探索是具有挑战性的。我们将首先使用我们的 研究模型人类癌细胞系中辐射诱导的碱基修饰的全球丰度的方法 然后制定一种策略来确定特定于基因的修饰位置。在目标2中，我们将探索 核酸序列基序，重点是microRNA。MicroRNAs与细胞之间的联系 功能/故障是公认的，但如此简短的主题可能会对研究构成挑战。我们将首先进行优化 我们对探测特定序列的测量，然后将其应用于肺癌相关的评估 未确认身份的患者血液样本中的microRNA。

Project Summary Nucleic acid biomarkers have tremendous potential value to patient health, but by their nature are often difficult to probe. Solid-state (SS-) nanopores are uniquely positioned to contribute to a solution for this challenge. The platform enables individual molecules to be probed as they translocate electrically through a synthetic, nanometer-scale aperture. The intrinsic sensitivity, compact nature, and electronic output make the system an attractive candidate for use as a translational diagnostic device. The central goal of this R33 project is to use SS-nanopore technology to assess nucleic acid biomarkers with applications to cancer through a highly selective assay developed in our lab. With our novel approach, target nucleic acid fragments can be detected and quantified only when bound to a protein chaperone, yielding a binary output. The measurement is rapid, sensitive, and yields an unambiguous electrical signal for analysis. We propose to apply this general measurement scheme in ways that enable the assessment of two broadly significant families of nucleic acid biomarkers. In Aim 1, we will assess epigenetic modifications and single base lesions. Modified bases regulate a variety of cellular functions but are challenging to probe with conventional technology. We will first use our approach to study global abundance of radiation-induced base modifications in model human cancer cell lines and then develop a strategy for determining the gene-specific location of modifications. In Aim 2, we will probe nucleic acid sequence motifs, focusing on microRNA. The link between microRNAs and cell function/malfunction is well established, but such short motifs can be challenging to study. We will first optimize our measurement for probing specific sequences, and then apply it to the assessment of lung cancer-relevant microRNA in de-identified patient blood samples.