DNA and RNA structures are defined by Watson-Crick rules of hybridization. However, a variety of DNA and RNA structures have been defined which do not rely on simple A-T/U and G-C binding. Since these structures disobey the Watson-Crick canon, they are described as non-canonical. Non-canonical DNA and RNA segments are formed as a result of secondary structures. These include G-quadruplexes, triplex forming oligos, hairpins, cruciforms, and i-Motif structures.
Oligonucleotide structural analysis has demonstrated that DNA and RNA nucleic acid sequences containing G-tracts separated by other bases spontaneously fold into G-quadruplex structures. G-quadruplexes are formed when four adjacent guanine residues stack in a cyclic Hoogsteen hydrogen-bonding arrangement leading to four-stranded helical structures. The study of G-quadruplexes in basic genetic processes is an active area of research in telomerase activity, gene regulation, and functional genomics. Guanine analogues that have different hydrogen bonding characteristics - 7-deaza-8-aza-dG and 7-deaza-dG - have proved useful in analyzing G-quadruplex structures. Similarly, common DNA lesions - 8-oxo-dG and abasic sites - have been used to investigate their effect on G-quadruplex structure and activity.
Intercalated Motif (i-Motif) DNA structures may be formed in regions rich in 2’-deoxyCytidine. Especially at acidic pH, these structures could be described as C-Quadruplexes with two parallel stranded sequences also held together in an antiparallel orientation by cytosine-cytosine base pairs. Since these structures are stable at acidic pH, they can act as nanoswitches by change in pH. As they were not considered to be stable at physiological pH, they were not initially considered to be relevant to biological systems. However, the stability of the cytosine-cytosine base pair is enhanced by intercallating ligands and so a variety of i-Motif structures are now considered to be biologically significant. Since i-Motif structures have now been observed forming and dissolving in living cells, these structures are now the subject of active investigation of the meaning of their activity in human cells. Research is also being directed to the effect of common DNA lesions, like depurinated sites, 8-oxo-dG and 5-hydroxymethyl-dC, on these transient structures.
Triplex-forming oligonucleotides (TFO) bind in the major groove of duplex DNA in a sequence-specific manner through the formation of non Watson-Crick (Hoogsteen) hydrogen bonds. The formation of a triplex along the major groove competes with the binding of transcription factors and other proteins that are necessary for transcription, thereby inhibiting the expression of particular genes. A variety of nucleoside analogues have been used in TFO - 8-amino-dG, 8-amino-dA, 6-thio-dG and deoxypseudouridine.