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Updated: May 26, 2026

Sequence-specific and Selective Recognition of Double-stranded RNAs over Single-stranded RNAs by Chemically Modified Peptide Nucleic Acids
Published on: September 21, 2017
Secondary binding sites for heavily modified triplex forming oligonucleotides
Antonia S Cardew1, Tom Brown, Keith R Fox
1Centre for Biological Sciences, Life Sciences Building, University of Southampton, Southampton SO17 1BJ, UK.
Synthetic oligonucleotides with specific amino modifications, like bis-amino-U, significantly enhance DNA triple helix stability and binding affinity. Researchers identified sequences with A(n) tracts bound by these modified oligonucleotides, confirming their effectiveness.
Area of Science:
- Molecular Biology
- Biochemistry
- Synthetic Chemistry
Background:
- Synthetic oligonucleotides are engineered to improve DNA triple helix stability.
- Bis-amino-U (B) is a highly effective modified nucleotide with 5-propargylamino and 2'-aminoethoxy groups.
- These modifications enhance both triplex stability and affinity for target sequences.
Purpose of the Study:
- To isolate and characterize DNA sequences bound by a modified triplex-forming oligonucleotide (TFO).
- To investigate the role of specific modifications in TFO binding and triplex formation.
- To identify secondary targets for heavily modified TFOs.
Main Methods:
- Utilized a restriction enzyme protection, selection, and amplification assay (REPSA).
- Employed DNase I footprinting to confirm triple helix formation.
- Analyzed TFO binding to sequences with varying modifications.
Main Results:
- REPSA identified sequences containing A(n) tracts (n=6) as targets for the modified TFO B(6)CBT.
- DNase I footprinting confirmed triple helix formation at these A(n) tracts.
- The modified TFO showed no interaction with oligonucleotides containing thymine (T) or 5-propargylamino-dU.
Conclusions:
- The 5'-end of the TFO, particularly the bis-amino-U modification, is crucial for binding to A(n) tracts and forming stable triple helices.
- Engineered oligonucleotides with specific amino modifications offer enhanced DNA targeting capabilities.
- This study provides insights into the sequence specificity and mechanism of modified TFOs in forming DNA triplexes.
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