Triplex formation of an oligonucleotide containing 2′-O-methylpseudoisocytidine with single and double stranded nucleic acids at neutral pH

An oligonucleotide analog containing 2'-O-methylpseudoisocytidine (P) and 2'-O-methyluridine (X) in an alternated homopyrimidine sequence (P-X-)(7)P-T can form triplexes with d-A-(G-A-)(7)G single strand and [d-A-(G-A-)(7)G]:[d-C-(T-C-)(7)T] duplex in neutral conditions. An UV mixing titration showed an end point of two units of (P-X-)(7)P-T to one unit of d-A-(G-A-)(7)G. This indicates that a [(P-X-)(7)P-T].[d-A-(G-A-)(7)G].[(P-X-)(7)P-T] is formed. The [(P-X-)(7)P-T].[d-A-(G-A-)(7)G].[(P-X-)(7)P-T] triplex is stable in a 0.1 M NaCl solution at neutral pH. However, the formation of triplex with [(P-X-)(7)P-T] and a duplex [d-A-(G-A-)(7)G].[d-C-(T-C-)(7)T] can be accomplished in solution also containing 5 mM MgCl2. CD spectra of both triplexes showed large negative bands at wavelength 210-230 nm. Both triplexes can be detected by native gel electrophoresis The thermal dissociation/association results indicate that this tripler dissociates to the single strands directly without going through a stable duplex intermediate.