Sequence-Dependent Fluorescence of Cyanine Dyes on Microarrays

Author(s)

Christy Agbavwe, Mark Manuel Somoza

Abstract

Cy3 and Cy5 are among the most commonly used oligonucleotide labeling molecules. Studies of nucleic acid structure and

dynamics use these dyes, and they are ubiquitous in microarray experiments. They are sensitive to their environment and

have higher quantum yield when bound to DNA. The fluorescent intensity of terminal cyanine dyes is also known to be

significantly dependent on the base sequence of the oligonucleotide. We have developed a very precise and highthroughput

method to evaluate the sequence dependence of oligonucleotide labeling dyes using microarrays and have

applied the method to Cy3 and Cy5. We used light-directed in-situ synthesis of terminally-labeled microarrays to determine

the fluorescence intensity of each dye on all 1024 possible 59-labeled 5-mers. Their intensity is sensitive to all five bases.

Their fluorescence is higher with 59 guanines, and adenines in subsequent positions. Cytosine suppresses fluorescence.

Intensity falls by half over the range of all 5-mers for Cy3, and two-thirds for Cy5. Labeling with 59-biotin-streptavidin-Cy3/-

Cy5 gives a completely different sequence dependence and greatly reduces fluorescence compared with direct terminal

labeling.

Organisation(s)

Department of Inorganic Chemistry

Journal

PLoS ONE

Volume

ISSN

1932-6203

Publication date

2011

Peer reviewed

Yes

Austrian Fields of Science 2012

104003 Inorganic chemistry

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