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Related Experiment Video

Updated: Apr 13, 2026

RNA Pull-down Procedure to Identify RNA Targets of a Long Non-coding RNA

RNA Pull-down Procedure to Identify RNA Targets of a Long Non-coding RNA

Published on: April 10, 2018

Alternative probe chemistries for single-molecule analysis of long noncoding RNA.

Kalika R Pai¹, Aimee M Martin², Madison Kadrmas³

¹Department of Chemistry and Biochemistry, University of Oregon, Eugene, Oregon, USA.

The Journal of Biological Chemistry

|April 12, 2026

Summary

Optimizing probe chemistry in single-molecule kinetic analysis of RNA transient structure (SiM-KARTS) improves long non-coding RNA (lncRNA) structural analysis. Locked nucleic acid (LNA) probes offer enhanced sensitivity for classifying complex RNA structures.

Keywords:

RNA structure fluorescence long noncoding RNA microscopy nucleic acid single-molecule biophysics

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Area of Science:

Molecular Biology
Biophysics
Genomics

Background:

Single-molecule microscopy is vital for RNA studies.
Analyzing complex structures like long non-coding RNA (lncRNA) using methods like single-molecule kinetic analysis of RNA transient structure (SiM-KARTS) presents challenges.
Optimization of experimental variables for SiM-KARTS on lncRNA is underexplored.

Purpose of the Study:

To investigate the impact of alternative probe backbone chemistries on SiM-KARTS analysis of lncRNA.
To establish design principles for applying SiM-KARTS to complex RNA targets.

Main Methods:

Utilized SiM-KARTS, thermal denaturation, and circular dichroism spectroscopy.
Analyzed binding behaviors of DNA probes with locked nucleic acid (LNA) and morpholino backbones.
Employed a model lncRNA segment to control target sequence accessibility.

Main Results:

Optimized probe backbone chemistry enables precise distinction between RNA structures and fine-tunes binding stability.
Locked nucleic acid (LNA) probes demonstrated high sensitivity to RNA structure in binding and unbinding kinetics.
Holistic analysis of binding and unbinding rates using LNA probes allowed accurate classification of different RNA structures.

Conclusions:

Probe backbone chemistry is a critical variable for optimizing SiM-KARTS analysis of lncRNA.
LNA probes provide a robust tool for high-accuracy structural classification of complex RNAs.
This study offers design guidelines for extending SiM-KARTS to challenging RNA targets like lncRNA.