3-Nitropyrrole and 5-nitroindole as universal bases in primers for DNA sequencing and PCR

Abstract

3-Nitropyrrole and 5-nitroindole have been assessed as universal bases in primers for dideoxy DNA sequencing and in the polymerase chain reaction (PCR). In contrast to a previous report, we have found that the introduction of more than one 3-nitropyrrole residue at dispersed positions into primers significantly reduced their efficiency in PCR and sequencing reactions. Primers containing 5-nitroindole at multiple dispersed positions were similarly affected; for both bases only a small number of substitutions were tolerated. In PCR experiments neither base, when incorporated into primers in codon third positions, was as effective as hypoxanthine, which was incorporated in six codon third positions in a 20mer oligomer. However, primers containing up to four consecutive 5-nitroindole substitutions performed well in both PCR and sequencing reactions. Consecutive 3-nitropyrrole substitutions were tolerated, but less well in comparable reactions.

Recognition of nonhybridizing base pairs during nucleotide excision repair of DNA

“…In the case of XPA protein, we observed preferential binding not only to carcinogen-damaged or mismatched substrates but, with higher efficiency, also to duplexes containing 5-nitroindole and 3-nitropyrrole residues. These aromatic base analogs lack hydrogen acceptors and donors for Watson-Crick strand pairing and, therefore, generate nonhybridizing sites characterized by increased hydrophobicity (26). Thus, the affinity of XPA for duplexes containing such aromatic analogs suggests that its strong bias for displaced pairing conformations is mediated by hydrophobic interactions with aromatic base components that are abnormally exposed to the helical surface.…”

“…4A). However, their aromatic ring structures have a similar electron distribution as natural bases and, as a consequence, retain at least in part the potential for hydrophobic interactions (26). When three consecutive 5-nitroindole (Fig.…”

Synthesis of Acyclic Nitroazole Nucleosides and Their Incorporation into Oligonucleotides, and Their Duplex and Triplex Formation

“…Similar behavior exhibit 2'-deoxyinosine analogues: 7-deaza-2'-deoxyinosine [5] and 2-aza-2'-deoxyinosine [6]. Melting studies of oligonucleotides containing the deoxyribonucleotides of 3-nitro-1H-pyrrole [7 ± 9], 4-, 5-, or 6-nitro-1H-indole [10,11] or 4-nitro-1H-imidazole [12] have demonstrated that the nitroazoles show less discrimination in their base pairs with the four natural bases than other types of analogues investigated as universal base candidates. To study base-pairing properties azolecarboxamides were also investigated [13 ± 16].…”

Stability and structure of DNA oligonucleotides containing non-specific base analogues

“…We believe, however, that they also stabilise duplexes predominantly by stacking interactions, and are therefore also valuable for studying the nature of non-covalent interactions within the double helix. Some of these analogues have been shown to be effective in some molecular biology applications, notably in PCR and sequencing (Bergstrom et al, 1995;Loakes et al, 1995;Parinov et al, 1996). Thus far, none of the`nonhydrogen bonding' bases has been shown to be an enzyme substrate, nor have their properties in templates been de®ned.…”