Design and calibration of Magnetic Tweezers for Investigating RNA Folding Dynamics in SARS-CoV-2

Ribosomal frameshifting is a critical moment in many coronaviruses' life cycles, during which an untranslated mRNA structure called the frameshift element (FSE) stalls the ribosome and allows it to shift 1 nucleotide back, which is crucial for the proper translation of downstream genes. In order to investigate the frameshifting mechanism of the FSE from SARS-CoV-2, we will use a combination of single-molecule FRET and custom built magnetic tweezers designed to interrogate the folding of the FSE at the molecular level.



This poster outlines the design and calibration of our magnetic tweezers and preliminary measurements of the stability of the FSE's two primary structural states. In particular, we compare the frequently used "look-up table" approach to determining a magnetic bead's z position with a more-robust and more-efficient method using the Fast Fourier Transform of the bead's image, and we discuss registration of the magnetic bead tracking data with the single-molecule fluorescence microscopy data using a method previously developed by members of this lab, S-T Polynomial Decomposition. Finally, we present preliminary measurements of transitions between the FSE's unusual "threaded" structural state and unthreaded state as a function of the applied force.