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Updated: May 13, 2026
13:42
RNA Secondary Structure Prediction Using High-throughput SHAPE
Published on: May 31, 2013
The effect of hybridization-induced secondary structure alterations on RNA detection using backscattering
Nicholas M Adams1, Ian R Olmsted, Frederick R Haselton
1Department of Chemistry, Vanderbilt University, Nashville, TN 37235, USA.
Nucleic Acids Research
|March 23, 2013
Summary
Backscattering interferometry (BSI) can detect viral RNA biomarkers without labels. Optimizing nucleic acid probes, particularly locked nucleic acid (LNA) probes, significantly enhances BSI sensitivity for infection detection.
Area of Science:
- Biochemistry
- Molecular Biology
- Medical Diagnostics
Background:
- Backscattering interferometry (BSI) is a label-free, high-sensitivity technique for monitoring molecular interactions.
- Its properties suggest potential for detecting infection biomarkers.
- Optimizing nucleic acid probe characteristics is crucial for maximizing BSI signal upon target binding.
Purpose of the Study:
- To identify interactions and characteristics of nucleic acid probes that maximize BSI signal for detecting respiratory syncytial virus (RSV) nucleocapsid gene RNA.
- To evaluate the impact of probe sequence, predicted RNA folding, and probe type (DNA vs. locked nucleic acid) on BSI sensitivity.
Main Methods:
- Investigated the correlation between the number of base pairs formed and BSI signal magnitude.
- Utilized RNA folding software (mfold) to predict unpaired nucleotides in target regions.
- Compared the sensitivity of locked nucleic acid (LNA) probes against DNA probes of identical sequences.
- Determined the limit of detection using multiple DNA probes targeting distributed regions of the viral RNA.
Main Results:
- BSI signal magnitude correlated with the number of base pairs formed between probes and the viral RNA target.
- Predicted unpaired nucleotides in targeted RNA regions generally correlated with BSI sensitivity.
- Locked nucleic acid (LNA) probes demonstrated a 4-fold improvement in sensitivity compared to DNA probes.
- Achieved a limit of detection of 624 pM (approximately 10^5 target molecules) using optimized DNA probes.
Conclusions:
- BSI is a promising tool for sensitive RNA detection, particularly for viral biomarkers.
- Optimizing probe design, including the use of LNA, significantly enhances BSI performance.
- Further improvements in BSI detection limits are feasible through strategic probe selection and design.
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