Single molecule DNA/RNA sequencing technology based on a parallel Raman scattering readout in a coupled nanochannel/nanopore system

基于耦合纳米通道/纳米孔系统中并行拉曼散射读数的单分子 DNA/RNA 测序技术

• 批准号: 10682588
• 负责人: Steven Brueck
• 金额: $ 90.88万
• 依托单位: ARMONICA TECHNOLOGIES, INC.
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-03-17 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10682588
• 关键词: 3-Dimensional; Adenine; Algorithms; Bacteriophage lambda; Behavior; Binding Proteins; Characteristics; Chemicals; Coupled; Cytosine; DNA; DNA Binding; DNA Damage; DNA Modification Process; DNA sequencing; Data Analyses; Data Storage and Retrieval; Deposition; Detection; Development; Dimensions; Electromagnetic Fields; Electromagnetics; Engineering; Ensure; Enzymes; Epigenetic Process; Film; Genomics; Goals; Grant; Hot Spot; Individual; Isotopes; Label; Lasers; Massive Parallel Sequencing; Measurement; Medical; Metals; Methylation; Modification; Molecular; Motion; Movement; Nanochip Analytical Device; Nanoporous; Nanostructures; Oligonucleotides; Optics; Particle Size; Pattern; Phase; Porosity; Preparation; Process; Protocols documentation; RNA; RNA Transport; Sampling; Signal Transduction; Silicon Dioxide; Single-Stranded DNA; Small Business Innovation Research Grant; Source; Speed; Stretching; Structure; Surface; System; Techniques; Technology; Time; Variant; atomic layer deposition; base; benzonitrile; colloidal nanoparticle; design; ds-DNA; electric field; fabrication; lithography; nanochannel; nanoparticle; nanopore; nanoscale; novel; operation; plasmonics; response; self assembly; sequencing platform; single molecule; transcriptome sequencing; vapor

Single molecule DNA/RNA transport and Raman sequencing technology based on parallel Raman scattering readout in a coupled nanochannel/nanopore system. Abstract Armonica Technologies, Inc. is proposing to develop a novel, high-throughput, label-free, highly accurate, long-read DNA sequencing platform based on inexpensive nanoscale patterning and self- assembly. The platform consists of nanochannels (cross section dimensions of ~ 100nm); tortuous (convoluted 3D) nanopores formed by self-assembly of colloidal nanoparticles; nanoparticle barriers placed across the nanochannels; and a metal-insulator-metal (MIM) field enhancement structure atop the nanochannel roof. In operation, single- stranded- or double-stranded-DNA is partially stretched into a linear configuration in the nanochannels, is blocked at barriers incorporated into the channels and forced (by electric field) to translocate through the tortuous nanopores in the roof. The MIM structure on the roof locally enhances the electromagnetic fields of applied laser sources allowing surface enhanced coherent anti-Stokes Raman scattering (SECARS) detection of individual bases as they pass through the electromagnetic hot spots, thus providing single base sensitivity and spatial localization. The distinct Raman spectra of the individual bases and epigenetic variants allow label-free sequencing. Optical detection allows massively parallel operation since the only requirement is separation of the pores by more than an optical wavelength, easily accomplished in the fabrication. An important feature of the platform is that the porous roofs allow introduction of oligonucleotides, small proteins, and DNA- binding/DNA-processing enzymes, permitting optional manipulation and modification of the DNA in the nanochannels. The goals of this Phase II project are: to develop a protocol for electrophoretic control of the ssDNA translocation; to engineer the structure to ensurethat the ssDNA passes the MIM hot spot; to sequence a short section of ssDNA and evaluate error rates; and to develop parallel optical readout, AI analysis of the data and appropriate storage protocol.

基于并行拉曼的单分子DNA/RNA转运和拉曼测序技术 耦合纳米通道/纳米孔系统中的散射读数。 抽象的 Armonica Technologies, Inc. 提议开发一种新型、高通量、无标签、高度 准确、长读长的 DNA 测序平台，基于廉价的纳米级图案和自 集会。该平台由纳米通道组成（横截面尺寸约为100纳米）；曲折 （卷积3D）由胶体纳米颗粒自组装形成的纳米孔；纳米粒子屏障 放置在纳米通道上；以及顶部的金属-绝缘体-金属（MIM）场增强结构 纳米通道屋顶。在操作中，单链或双链 DNA 被部分拉伸成 纳米通道中的线性构型被纳入通道中的屏障阻挡并被迫 （通过电场）通过屋顶上曲折的纳米孔进行移位。屋顶MIM结构 局部增强所应用激光源的电磁场，允许表面增强相干 当单个碱基通过时，进行反斯托克斯拉曼散射 (SECARS) 检测 电磁热点，从而提供单碱基灵敏度和空间定位。独特的 单个碱基和表观遗传变异的拉曼光谱允许无标记测序。光学的 检测允许大规模并行操作，因为唯一的要求是通过 超过一个光波长，在制造中很容易完成。该系统的一个重要特点是 平台的特点是多孔屋顶允许引入寡核苷酸、小蛋白质和DNA- 结合/DNA 加工酶，允许对 DNA 进行选择性操作和修饰 纳米通道。该第二阶段项目的目标是：开发一种电泳控制协议 单链DNA易位；设计结构以确保ssDNA通过MIM热点；到 对一小段 ssDNA 进行测序并评估错误率；并开发并行光学读出、人工智能 数据分析和适当的存储协议。