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Nucleic Acids Research 2006 34(8):e60; doi:10.1093/nar/gkl175
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Published online 2 May 2006

© The Author 2006. Published by Oxford University Press. All rights reserved
The online version of this article has been published under an open access model. Users are entitled to use, reproduce, disseminate, or display the open access version of this article for non-commercial purposes provided that: the original authorship is properly and fully attributed; the Journal and Oxford University Press are attributed as the original place of publication with the correct citation details given; if an article is subsequently reproduced or disseminated not in its entirety but only in part or as a derivative work this must be clearly indicated. For commercial re-use, please contact journals.permissions@oxfordjournals.org

Methods Online

Design of LNA probes that improve mismatch discrimination

Yong You, Bernardo G. Moreira, Mark A. Behlke and Richard Owczarzy*

Integrated DNA Technologies 1710 Commercial Park, Coralville, IA 52241, USA

*To whom correspondence should be addressed. Tel: +1 319 626 8459; Fax: +1 319 626 8444; Email: science{at}owczarzy.net

Received December 17, 2005. Revised March 21, 2006. Accepted March 21, 2006.

Locked nucleic acids (LNA) show remarkable affinity and specificity against native DNA targets. Effects of LNA modifications on mismatch discrimination were studied as a function of sequence context and identity of the mismatch using ultraviolet (UV) melting experiments. A triplet of LNA residues centered on the mismatch was generally found to have the largest discriminatory power. An exception was observed for G–T mismatches, where discrimination decreased when the guanine nucleotide at the mismatch site or even the flanking nucleotides were modified. Fluorescence experiments using 2-aminopurine suggest that LNA modifications enhance base stacking of perfectly matched base pairs and decrease stabilizing stacking interactions of mismatched base pairs. LNAs do not change the amount of counterions (Na+) that are released when duplexes denature. New guidelines are suggested for design of LNA probes, which significantly improve mismatch discrimination in comparison with unmodified DNA probes.


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