Open AccessSingle-Stranded DNA Catalyzes Hybridization of PCR-Products to Microarray Capture Probes
Author(s) -
Simon Dally,
Steffen Rupp,
Karin Lemuth,
Stefan Hartmann,
Ekkehard Hiller,
Susanne M. Bailer,
Cornelius Knabbe,
Jan Weile
Publication year - 2014
Publication title -
plos one
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.99
H-Index - 332
ISSN - 1932-6203
DOI - 10.1371/journal.pone.0102338
Subject(s) - dna microarray , in situ hybridization , dna–dna hybridization , nucleic acid thermodynamics , microarray , hybridization probe , dna , biology , computational biology , microbiology and biotechnology , chemistry , genetics , gene , gene expression , base sequence
Since its development, microarray technology has evolved to a standard method in the biotechnological and medical field with a broad range of applications. Nevertheless, the underlying mechanism of the hybridization process of PCR-products to microarray capture probes is still not completely understood, and several observed phenomena cannot be explained with current models. We investigated the influence of several parameters on the hybridization reaction and identified ssDNA to play a major role in the process. An increase of the ssDNA content in a hybridization reaction strongly enhanced resulting signal intensities. A strong influence could also be observed when unlabeled ssDNA was added to the hybridization reaction. A reduction of the ssDNA content resulted in a massive decrease of the hybridization efficiency. According to these data, we developed a novel model for the hybridization mechanism. This model is based on the assumption that single stranded DNA is necessary as catalyst to induce the hybridization of dsDNA. The developed hybridization model is capable of giving explanations for several yet unresolved questions regarding the functionality of microarrays. Our findings not only deepen the understanding of the hybridization process, but also have immediate practical use in data interpretation and the development of new microarrays.