Nanopore-Based Selective Discrimination of MicroRNAs with Single-Nucleotide Difference Using Locked Nucleic Acid-Modified Probes

Abstract

The accurate discrimination of microRNAs (miRNAs) with highly similar sequences would greatly facilitate the screening and early diagnosis of diseases. In the present work, a locked nucleic acid (LNA)-modified probe was designed and used for α-hemolysin (α-HL) nanopore to selectively and specifically identify miRNAs. The hybridization of the LNA probe with the target miRNAs generated unique long-lived signals in the nanopore thus facilitated an accurate discrimination of miRNAs with similar sequences, even a single-nucleotide difference. Furthermore, the developed nanopore-based analysis with LNA probe could selectively detect target miRNAs in a natural serum background. This selective and sensitive approach may be highly valuable in the detection of clinically relevant biomarkers in complex samples.

Nanopore device-based fingerprinting of RNA oligos and microRNAs enhanced with an Osmium tag

“…A limited number of experiments was conducted with the wt a-HL/EBS membrane (EBS) platform (for examples, see Figure S18 in the Supplementary Information). Nanopore experiments were conducted with 10 µM synthetic DNA/RNA oligo in 1.0 M KCl, 10 mM TRIS.HCl buffer at pH 8.0 and at 20 ± 1 °C, as described in detail 61 , and summarized here. wt α-HL was purchased from List Biological Laboratories in the monomer form of lyophilized power and dissolved in water at 1 mg/mL.…”

Membrane protein-based biosensors

“…This large destabilization allowed the detection of the single mismatch on the PNA-RNA hybrid by the current signals and, thus, enabled the discrimination of single nucleotide-difference from the target miRNA. For the discrimination of the single nucleotide mismatch, Wang and colleagues used a locked nucleic acid (LNA) probe [51], which is a nucleic acid analogue bridging between 2 0 oxygen and 4 0 carbon that significantly stabilizes the hybridization. Similar to the PNA-RNA, the LNA probe was able to discriminate a single nucleotide difference on a miRNA from the differences of the blocking dwell times.…”

Enzyme and AIEgens Modulated Solid‐State Nanochannels: In Situ and Noninvasive Monitoring of H2O2 Released from Living Cells

“…The recent decades have seen the great development of sensing field using functionalized solid‐state nanochannels . So far, the functionalized solid‐state nanochannels have revealed tremendous abilities in the sensing of ions, small biomolecules, biological macromolecules, and even cellular targets with the unique merits of rapid response, high sensitivity, miniaturization and low‐cost .…”

“…[24][25][26][27][28][29] The recent decades have seen the great development of sensing field using functionalized solid-state www.advancedsciencenews.com www.small-methods.com nanochannels. [30][31][32][33][34][35] So far, the functionalized solid-state nanochannels have revealed tremendous abilities in the sensing of ions, [36][37][38][39][40] small biomolecules, [41][42][43][44] biological macromolecules, [45][46][47][48][49] and even cellular targets [18] with the unique merits of rapid response, high sensitivity, miniaturization and lowcost. [50][51][52] However, to fulfill cellular target detection, the current test platform requires at least two steps (Scheme 1a).…”