Probing RNA Conformations Using a Polymer-Electrolyte Solid-State Nanopore

Nanopore systems have emerged as a leading platform for the analysis of biomolecular complexes with singlemolecule resolution. The conformation of biomolecules, such as RNA, is highly dependent on the electrolyte composition, but solid-state nanopore systems often require high salt concentration to operate, precluding analysis of macromolecular conformations under physiologically relevant conditions. Here, we report the implementation of a polymer-electrolyte solid-state nanopore system based on alkali metal halide salts dissolved in 50% w/v poly(ethylene) glycol (PEG) to augment the performance of our system. We show that polymer-electrolyte bath governs the translocation dynamics of the analyte which correlates with the physical properties of the salt used in the bath. This allowed us to identify CsBr as the optimal salt to complement PEG to generate the largest signal enhancement. Harnessing the effects of the polymer-electrolyte, we probed the conformations of the Chikungunya virus (CHIKV) RNA genome fragments under physiologically relevant conditions. Our system was able to fingerprint CHIKV RNA fragments ranging from similar to 300 to similar to 2000 nt length and subsequently distinguish conformations between the cotranscriptionally folded and the natively refolded similar to 2000 nt CHIKV RNA. We envision that the polymer-electrolyte solidstate nanopore system will further enable structural and conformational analyses of individual biomolecules under physiologically relevant conditions.

Automated summary

This study presents a polymer-electrolyte solid-state nanopore system for analyzing biomolecular conformation under physiological conditions. By using specific salts, the system's performance is enhanced, allowing for the analysis of Chikungunya virus RNA conformation.