Phase II year
2022
(last award dollars: 2023)
Phase II Amount
$1,852,664
Single molecule DNA/RNA transport and Raman sequencing technology based on parallel Raman scattering readout in a coupled nanochannel/nanopore system. 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.
Public Health Relevance Statement: Single molecule DNA/RNA sequencing technology based on a parallel Raman scattering readout in a coupled nanochannel/nanopore sequencing system.
Project narrative: Rapid, label-free, highly accurate, long-read DNA sequencing has important implications for genomic and medical applications. A novel nanostructured platform consisting of nanochannels, tortuous (convoluted 3D) nanopores, nanopore barriers along the nanochannels and metal-insulator-metal (matamaterial/plasmonic) antennas is proposed. Sequencing of the DNA is by label-free surface- enhanced Raman scattering, an highly specific process that identifies epigenetic chemical modification and does not require labeling or amplification, simplifying the sequencing flow, and, most importantly allowing processing of cellular DNA with minimum pre-preparation. The optical readout enables massively parallel sequencing with dramatically faster throughput than current technologies.
Project Terms: Adenine; 1H-Purin-6-amine; Vitamin B4; Algorithms; Behavior; Cytosine; Data Analyses; Data Analysis; data interpretation; DNA; Deoxyribonucleic Acid; DNA Damage; DNA Injury; Electromagnetic Fields; Electromagnetic Fields Radiation; electromagnetic field; Electromagnetics; Engineering; Enzymes; Enzyme Gene; Goals; Grant; Isotopes; Lasers; Laser Electromagnetic; Laser Radiation; Metals; Methylation; Motion; Movement; body movement; Oligo; oligos; Oligonucleotides; optical; Optics; Particle Size; Coliphage lambda; Enterobacteria phage lambda; Phage lambda; Bacteriophage lambda; Non-Polyadenylated RNA; RNA Gene Products; Ribonucleic Acid; RNA; Cell Communication and Signaling; Cell Signaling; Intracellular Communication and Signaling; Signal Transduction Systems; Signaling; biological signal transduction; Signal Transduction; Cristobalite; Sand; Silica; Tridymite; Silicon Dioxide; Technology; Time; benzonitrile; Film; base; Label; Surface; Phase; Variant; Variation; Medical; Ensure; Chemicals; Individual; Ligand Binding Protein; Ligand Binding Protein Gene; Protein Binding; bound protein; Binding Proteins; Measurement; data retrieval; data storage; Data Storage and Retrieval; Double-Stranded DNA; dsDNA; ds-DNA; Deposit; Deposition; electric field; Dimensions; Protocol; Protocols documentation; Source; Pattern; Techniques; System; 3-D; 3D; three dimensional; 3-Dimensional; vapor; Ribonucleic Acid Transport; RNA Transport; lithography; Stretching; Speed; Structure; novel; Sampling; response; monolayer; single molecule; Genomics; DNA Binding Interaction; DNA bound; DNA Binding; DNA Modification; DNA Modification Process; nano chip; nano chip Analytical Device; nanochip; Nanochip Analytical Device; nano-structures; Nanostructures; Detection; Epigenetic Process; Epigenetic; Epigenetic Change; Epigenetic Mechanism; Small Business Innovation Research Grant; SBIR; Small Business Innovation Research; Preparation; Characteristics; Hot Spot; Molecular; Process; Modification; Development; developmental; self assembly; design; designing; atomic layer deposition; colloidal nanoparticle; colloidal nano particle; nanochannel; nano channel; nanoscale; nano meter scale; nano meter sized; nano scale; nanometer scale; nanometer sized; nanopore; nano pore; nanoparticle; nano particle; nano-sized particle; nanosized particle; Coupled; plasmonics; operation; transcriptome sequencing; RNA Seq; RNA sequencing; RNAseq; Massive Parallel Sequencing; Massively Parallel DNA Sequencing; Massively Parallel Sequencing; sequencing platform; DNA sequencing; DNA seq; DNAseq; Nanoporous
