Nucleobase-Modified Triplex-Forming Peptide Nucleic Acids for Sequence-Specific Recognition of Double-Stranded RNA and DNA.

Non-coding RNAs play important roles in the regulation of gene expression. Sequence-specific recognition of these regulatory RNAs is essential for both fundamental scientific research and the pharmaceutical industry. However, the complex helical structures formed by biologically relevant RNAs present a formidable challenge for molecular recognition. The present protocol describes nucleobase-modified triplex-forming peptide nucleic acids (PNAs) designed for sequence-specific recognition of double-stranded RNA and DNA under physiologically relevant conditions. Protocols for PNA synthesis and purification, as well as RNA binding studies using isothermal titration calorimetry and UV thermal melting, are described in detail. We also describe improved PNA bis-clamps with the ability to invade double-stranded DNA at physiological salt conditions.