IDBR: Development of a Universal Spectroscopic Nanosensing System for Multiplexed Genomic and Proteomic Analysis

该奖项支持开发一种超灵敏的光谱传感系统，该系统有望在不需要扩增的情况下检测低丰度（zeptomoles或更少）的生物分子目标。 该系统将能够在单一平台上对DNA、RNA和蛋白质进行多重分析。 使用PCR的靶扩增是目前用于检测低丰度核酸的最有效的方法;然而，PCR倾向于诱导背景扩增，这常常破坏其准确性。 由于蛋白质的扩增技术不可用，因此难以检测低丰度。整合的基因组和蛋白质组学分析的重要性日益增加，例如mRNA和蛋白质表达的相关测量，需要能够在单个平台上进行基因组和蛋白质组学分析的通用生物分析工具。该传感系统将基于单分子检测（SMD）、微/纳米流体和分子生物学的创新集成。 该系统将包括一个通用的纳米传感器，能够检测核酸和蛋白质的目标和一个4色微流体SMD分光镜能够多重荧光检测的单分子。 纳米传感器将由功能化的量子点（QD）组成，量子点将给出结合诱导的荧光共振能量转移（FRET）信号。 当与4色微流控SMD光谱仪相结合时，低丰度目标的高通量检测将成为可能，而无需扩增。 该装置的能力将通过在单细胞模型基因表达系统中同时测量mRNA和蛋白质表达，以及通过对未扩增基因组DNA中的单核苷酸多态性（SNP）进行基因分型来证明。 这项工作的跨学科性质将为研究生和本科生的跨学科培训提供极好的机会。 成功开发微流体设备后，学生将参与BioMEMS，微/纳米流体和纳米生物传感器系统领域的一系列独立研究项目。 高中生也有望参与该项目的某些方面。

This award supports development of an ultrasensitive spectroscopic sensing system expected to be able to detect low-abundance (zeptomoles or less) biomolecular targets without the need for amplification. This system will be capable of multiplexed analysis of DNA, RNA, and protein on a single platform. Target amplification using PCR is currently the most effective approach for detection of low-abundance nucleic acids; however, PCR tends to induce background amplification which often undermines its accuracy. Because an amplification technique for proteins is not available, detection of low-abundance is difficult. The increasing importance of integrated genomic and proteomic analysis, e.g. correlated measurements of mRNA and protein expression, calls for a universal bioanalytical tool capable of both genomic and proteomic analysis on a single platform. The sensing system will be based on innovative integration of single-molecule detection (SMD), micro/nanofluidics, and molecular biology. The system will be comprised of a generic nanosensor capable of detecting both nucleic acids and protein targets and a 4-color microfluidic SMD spectroscope capable of multiplexed fluorescence detection of single molecules. The nanosensor will consist of functionalized quantum dots (QDs) that will give a binding-induced fluorescence resonance energy transfer (FRET) signal. When combined with the 4-color microfluidic SMD spectroscope, high-throughput detection of low-abundance targets will be possible without amplification. The capability of the device will be demonstrated through simultaneous measurements of mRNA and protein expression in a model gene expression system in single cells, and through genotyping single-nucleotide polymorphisms (SNPs) in unamplified genomic DNA. The interdisciplinary nature of the effort will provide excellent opportunities for cross-disciplinary training at the graduate and undergraduate level. After successful development of the microfluidic devices, students will be involved in a series of independent research projects in the areas of BioMEMS, micro/nanofluidics, and nanobiosensor systems. High school students are also expected to participate in some aspects of the project.