Resolving the Sequence of RNA Strands by Tip-Enhanced Raman Spectroscopy

RNA plays critical roles in guiding protein expression and catalyzing biological reactions. The gold standard RNA sequencing method requires converting RNA to complementary DNA (cDNA). This is followed by DNA amplification via polymerase chain reaction (PCR) and sequencing, making RNA sequencing indirect, complicated, and susceptible to sequence data bias. This paper demonstrates RNA imaging at the single-base level while illustrating a direct method to read RNA sequences by tip-enhanced Raman scattering (TERS) technique. To resolve nucleotides within an RNA strand, we adopted gap-mode TERS involving a gold tip and a gold substrate. After analyzing TERS measurements based on the reference sequence, we identified RNA sequences with 90% accuracy. This proof-of-principle RNA imaging method significantly advances a direct RNA sequencing technique without RNA labeling or reverse transcriptase RT-PCR amplification.