Direct Sensing of Single Native RNA with a Single-Biomolecule Interface of Aerolysin Nanopore

RNA sensing is of vital significance to advance our comprehension of gene expression and to further benefit medical diagnostics. Taking advantage of the excellent sensing capability of the aerolysin nanopore as a single-biomolecule interface, we for the first time achieved the direct characterization of single native RNA of Poly(A)₄ and Poly(U)₄. Poly(A)₄ induces ∼10% larger blockade current amplitude than Poly(U)₄. The statistical duration of Poly(A)₄ is 18.83 ± 1.08 ms, which is 100 times longer than that of Poly(U)₄. Our results demonstrated that the capture of RNA homopolymers is restricted by the biased diffusion. The translocation of RNA needs to overcome a lower free-energy barrier than that of DNA. Moreover, the strong RNA-aerolysin interaction is attributed to the hydroxyl in pentose, which prolongs the translocation time. This study opens an avenue for aerolysin nanopores to directly achieve RNA sensing, including discrimination of RNA epigenetic modification and selective detection of miRNA.