PNA/DNA duplex

To elaborate upon this scheme, the authors replaced the DNA-based capture strand with one that is based on peptide nucleic acid (PNA) in order to distinguish single nucleotide mismatches [34]. Since PNA does not have a phospho-diester backbone, a PNA-DNA duplex is more thermodynamically stable than DNA-DNA or RNA-RNA helices due to a lack of electrostatic repulsion between the two chains. PNAs have also been shown to be more selective. However, PNAs are more difficult to synthesize and PNA-DNA interactions are not as well understood as DNA-DNA binding. In this study, quaternary ammo-... 

DNA and PNA/DNA duplexes was examined. The 32P-labeled DNA target strand was 5 -TCGCTGGcAAZAAGGbTAGGaA-3 where Z = an abasic site and three different 5 -GG-3 sites were present (GGA-C). The PNA strand s base sequence was complementary to the DNA target strand s sequence with the exception that anthraquinone (AQ) was bound to the PNA strand in place of a normal DNA base opposite the abasic site (Z) in the DNA target strand. 

Irradiation of the above PNA-DNA duplex showed preferential strand cleavage at the 5 G of the GGA and GGB sites, which was much the same result as occurred in DNA/DNA duplexes under similar conditions. Somewhat in contrast to this result, DNA strand cleavage occured equally at each G of the GGC site. The 5 -G of GGA was 6 bases away from the abasic site (ca. 20.4 A), where AQ was intercalated, yet as much damage occured there as at the 5 -G of GGB and each G of GGC. Both of these latter two sites were only two bases away from the AQ site thus distance from the site of photoexcitation was not an imporatant factor in determining strand cleavage. 

Another method for nonfluorescent detection of DNA involves the noncovalent binding of an inexpensive cyanine dye to PNA-DNA duplexes (27). The dye assembles into a helical aggregate in the presence of PNA-DNA, which results in a vivid blue-purple color change. This phenomenon has been used in SNP detection assays. 

When one or several isolated 2PA ligands were introduced in LNA DNA duplexes, addition of 1 equiv of Ni, Zn, or Cu per 2PA ligand significantly increased the duplex stability with respect to that of the metal-free duplexes (121). A similar increase in stability was observed upon addition of 1 equiv of Ni or Cu to 2PA-modified PNA DNA duplexes (137). The effect of Ni was larger than that of Cu for both types of duplexes. In the case of Zn, 2 equiv of metal ions were necessary to measure reversible melting transitions for PNA DNA duplexes, which was attributed to the low-binding constant of 2PA for Zn + (137). 

It is conceivable that an interstrand complex can be created by coordination of a metal ion to the 2PA situated on the PNA or LNA strand and to a nucleobase or phosphodiester group from the opposite, non-modified DNA strand in the nucleic acid duplex. Molecular modeling of LNA DNA duplexes showed that a [3 -I-1] 2PA-M -P04R2 coordination mode can be realized by coordination of a 2PA ligand from one strand and a phosphodiester from either the same strand or from the opposite strand of the LNA DNA duplex (121). This model was supported by data obtained for 2PA-modified PNA DNA duplexes showing that (a) the... 

Stability of a Ni -containing, 2PA-modilied PNA DNA duplex did not depend on the duplex sequence, and (b) the effect of Ni on the duplex stability depended on the ionic strength of the solution (137). In most cases, addition of > 1 equiv of metal ion per pair of adjacent 2PA ligands destabilized the duplex, suggesting that steric and electrostatic repulsion occurs when each of the 2PA ligands binds a metal ion. 

Table 13. PNA duplex characteristics I PNA/RNA and PNA/DNA duplexes show increased stability when compared with natural duplexes (Conditions 10 mM phosphate buffer 0.1 mM EDTA 100 mM NaCl pH 7.0). II Dependence of Tm-values of PNA/DNA and natural duplexes on counterion strength. From [223]...

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