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  Impact of point-mutations on the hybridization affinity of surface-bound DNA/DNA and RNA/DNA oligonucleotide-duplexes: Comparison of single base mismatches and base bulges

Naiser, T., Ehler, O., Kayser, J., Mai, T., Michel, W., & Ott, A. (2008). Impact of point-mutations on the hybridization affinity of surface-bound DNA/DNA and RNA/DNA oligonucleotide-duplexes: Comparison of single base mismatches and base bulges. BMC BIOTECHNOLOGY, 8: 48. doi:10.1186/1472-6750-8-48.

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Item Permalink: https://hdl.handle.net/21.11116/0000-000F-29FD-7 Version Permalink: https://hdl.handle.net/21.11116/0000-000F-2EAB-E
Genre: Journal Article

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This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
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 Creators:
Naiser, Thomas1, Author
Ehler, Oliver1, Author
Kayser, Jona2, Author
Mai, Timo1, Author
Michel, Wolfgang1, Author
Ott, Albrecht1, Author
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1external, ou_persistent22              
2External Organizations, ou_persistent22              

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 Abstract: Background: The high binding specificity of short 10 to 30 mer
oligonucleotide probes enables single base mismatch ( MM) discrimination
and thus provides the basis for genotyping and resequencing microarray
applications. Recent experiments indicate that the underlying principles
governing DNA microarray hybridization - and in particular MM
discrimination - are not completely understood. Microarrays usually
address complex mixtures of DNA targets. In order to reduce the level of
complexity and to study the problem of surface-based hybridization with
point defects in more detail, we performed array based hybridization
experiments in well controlled and simple situations.
Results: We performed microarray hybridization experiments with short 16
to 40 mer target and probe lengths ( in situations without competitive
hybridization) in order to systematically investigate the impact of
point-mutations - varying defect type and position - on the
oligonucleotide duplex binding affinity. The influence of single base
bulges and single base MMs depends predominantly on position - it is
largest in the middle of the strand. The position-dependent influence of
base bulges is very similar to that of single base MMs, however certain
bulges give rise to an unexpectedly high binding affinity. Besides the
defect ( MM or bulge) type, which is the second contribution in
importance to hybridization affinity, there is also a sequence
dependence, which extends beyond the defect next-neighbor and which is
difficult to quantify. Direct comparison between binding affinities of
DNA/DNA and RNA/DNA duplexes shows, that RNA/DNA purine-purine MMs are
more discriminating than corresponding DNA/DNA MMs. In DNA/ DNA MM
discrimination the affected base pair ( C . G vs. A . T) is the
pertinent parameter. We attribute these differences to the different
structures of the duplexes ( A vs. B form).
Conclusion: We have shown that DNA microarrays can resolve even subtle
changes in hybridization affinity for simple target mixtures. We have
further shown that the impact of point defects on oligonucleotide
stability can be broken down to a hierarchy of effects. In order to
explain our observations we propose DNA molecular dynamics - in form of
zipping of the oligonucleotide duplex - to play an important role.

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Language(s): eng - English
 Dates: 2008-05-132008-05-13
 Publication Status: Issued
 Pages: -
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 Table of Contents: -
 Rev. Type: -
 Identifiers: ISI: 000257250600001
DOI: 10.1186/1472-6750-8-48
 Degree: -

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Title: BMC BIOTECHNOLOGY
Source Genre: Journal
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Pages: - Volume / Issue: 8 Sequence Number: 48 Start / End Page: - Identifier: ISSN: 1472-6750