High‐Throughput Block Optical DNA Sequence Identification

Optical techniques for molecular diagnostics or DNA sequencing generally rely on small molecule fluorescent labels, which utilize light with a wavelength of several hundred nanometers for detection. Developing a label‐free optical DNA sequencing technique will require nanoscale focusing of light, a high‐throughput and multiplexed identification method, and a data compression technique to rapidly identify sequences and analyze genomic heterogeneity for big datasets. Such a method should identify characteristic molecular vibrations using optical spectroscopy, especially in the “fingerprinting region” from ≈400–1400 cm^?1. Here, surface‐enhanced Raman spectroscopy is used to demonstrate label‐free identification of DNA nucleobases with multiplexed 3D plasmonic nanofocusing. While nanometer‐scale mode volumes prevent identification of single nucleobases within a DNA sequence, the block optical technique can identify A, T, G, and C content in DNA k‐mers. The content of each nucleotide in a DNA block can be a unique and high‐throughput method for identifying sequences, genes, and other biomarkers as an alternative to single‐letter sequencing. Additionally, coupling two complementary vibrational spectroscopy techniques (infrared and Raman) can improve block characterization. These results pave the way for developing a novel, high‐throughput block optical sequencing method with lossy genomic data compression using k‐mer identification from multiplexed optical data acquisition.