TRIPLER FORMATION AT SINGLE-STRANDED NUCLEIC-ACID TARGET SITES OF UNRESTRICTED SEQUENCE BY 2 ADDED STRANDS OF OLIGONUCLEOTIDES - A PROPOSEDMODEL

By using the standard purine nucleosides, guanosine and adenosine, and the pyrimidine C-nucleosides, pseudoisocytidine and pseudouridine, as complements on a probe strand, it is possible to construct a regular Watson-Crick helix with a single-stranded target sequence having any a rrangement of the four naturally-occurring bases found in nucleic acid s. The major groove of this helix will have a unique configuration of hydrogen-bonding sites on the probe strand for each of these four base pairs. By using this duplex as a framework, an ensemble of recognitio n patterns composed of base triads may be constructed. In these patter ns, either a homopyrimidine or homopurine third strand binds in the ma jor groove of the duplex formed by the target and probe strands. Ten d istinct geometries, or motifs, are shown, each one consisting of four isomorphic base triads built upon recognition of C, G, A, or U(T) resi dues in the target strand. Four motifs contain pyrimidines as residues on the third strand which base pair to the second strand through spec ific hydrogen-bonding interactions, four motifs involve purines which base-pair to the second strand through donor-acceptor sites located on their six-membered ring, and two motifs utilize purines binding to th e second strand at sites located on both their five- and six-membered rings. For base triads or for base-pairing interactions which involve the common bases found in nucleic acids, most of the hydrogen-bonding patterns have been previously recognized. In order to maintain specifi c hydrogen bonding and to construct isomorphous triads, the use of sev eral nonstandard bases is proposed. A subset of the base triads may al so be used to design oligonucleotides which may bind as a third strand to naturally-occurring homopyrimidine-homopurine double-stranded targ et sites such as those found in DNA. In addition, another set of four bases which have Watson-Crick complementarity to the target-strand bas es and which provide alternative patterns of donor-acceptor pairs for third-strand interactions can be proposed for use on the second strand . With the palette of eight second-strand bases, four possible permuta tions of (target strand)-(second strand) interactions are shown. For e ach of the four residues on the target strand, any single permutation presents unique hydrogen-bonding patterns for third-strand binding whi ch may occur according to one of the ten triad motifs.