Rapid, Binary-Mode Nucleic Acid Sequence Detection

技术摘要特定核酸（NA）序列的检测在传染病诊断、遗传性疾病的遗传筛查、个性化医疗、犯罪学、亲子纠纷、动植物育种等许多应用中至关重要。尽管聚合酶链反应（PCR）技术与寡核苷酸阵列芯片相结合，能够在非常低的拷贝数下对样品中存在的多个序列进行特异性鉴定，但这种方法需要添加标记和试剂来检测目标NA。这些标签和检测它们所需的硬件（例如光学）显着增加了检测特定NA序列的设备的分析时间，成本和占地面积。作为现有方法的替代方案，该项目将开发能够以单分子分辨率检测致病性核酸的新型纳米孔。这些纳米孔将在硅中制造，并将被设计成含有带有互补NA序列标记的珠粒。目标寡核苷酸与微球结合将导致电泳性能的改变，并导致纳米孔被微球遮挡。该事件将通过电化学检测目标NA的二进制yes/no读数。摘要为了医疗或食品安全的应用，需要开发简单、廉价的检测病原体的方法。这一提议将开发一种基于含珠小孔隙的新方法。这些珠子会与病原体生物标志物发生反应，堵塞毛孔，并导致电流的变化。设想这种方法比目前的方法具有更高的灵敏度和更简单的方法。

Technical abstractDetection of specific nucleic acid (NA) sequences is crucial to infectious diseasediagnostics, genetic screening for hereditary diseases, personalized medicine, criminology,paternity disputes, and animal and plant breeding, among many other applications. Althoughpolymerase chain reaction (PCR) technology, coupled with oligonucleotide array chips, hasenabled specific identification of multiple sequences present in a sample at very low copynumber, this method requires added labels and reagents to detect the target NA. These labelsand the hardware (e.g., optics) required to sense them significantly increase the analysis time,cost, and footprint of devices for detecting specific NA sequences. As an alternative to current approaches, this project will develop novel nanopores capable of detecting pathogenic nucleic acids with single molecule resolution. These nanopores will be fabricated in silicon and will be engineered to contain beads labeled with complementary NA sequences. Binding of target oligonucleotides to the beads will result in the change of electrophoretic properties and will lead to nanopores becoming occluded by the beads. This event will be detected electrochemically for binary yes/no readout of the target NA. Non-technical abstractThere is a need to develop simple and inexpensive methods for detecting pathogens for medical or food safety applications. This proposal will develop a new approach based on small pores containing beads. The beads will react to pathogen biomarkers, will clog the pores and will result in the change of electrical current. This method is envisioned to have improved sensitivity and be simpler than the current approaches.