Strong position-dependent effects of sequence mismatches on signal ratios measured using long oligonucleotide microarrays.
BACKGROUND
Microarrays are an important and widely used tool. Applications include capturing genomic DNA for high-throughput sequencing in addition to the traditional monitoring of gene expression and identifying DNA copy number variations. Sequence mismatches between probe and target strands are known to affect the stability of the probe-target duplex, and hence the strength of the observed signals from microarrays.


RESULTS
We describe a large-scale investigation of microarray hybridisations to murine probes with known sequence mismatches, demonstrating that the effect of mismatches is strongly position-dependent and for small numbers of sequence mismatches is correlated with the maximum length of perfectly matched probe-target duplex. Length of perfect match explained 43% of the variance in log2 signal ratios between probes with one and two mismatches. The correlation with maximum length of perfect match does not conform to expectations based on considering the effect of mismatches purely in terms of reducing the binding energy. However, it can be explained qualitatively by considering the entropic contribution to duplex stability from configurations of differing perfect match length.


CONCLUSIONS
The results of this study have implications in terms of array design and analysis. They highlight the significant effect that short sequence mismatches can have upon microarray hybridisation intensities even for long oligonucleotide probes. All microarray data presented in this study are available from the GEO database 1, under accession number [GEO: GSE9669]
Rennie
Catriona
C
Biosciences Building, School of Biological Sciences, University of Liverpool, Crown Street, Liverpool, L69 7ZB, UK. catriona.rennie@postgrad.manchester.ac.uk
Noyes
Harry A
HA
Kemp
Stephen J
SJ
Hulme
Helen
H
Brass
Andy
A
Hoyle
David C
DC
D000818:Animals; D020137:Base Pair Mismatch; D020029:Base Pairing; D018076:DNA, Complementary / Q000737:chemistry; D005456:Fluorescent Dyes / Q000737:chemistry; D005544:Forecasting; D018628:Gene Dosage; D014644:Genetic Variation; D051379:Mice; D008805:Mice, Inbred A; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D008815:Mice, Inbred Strains; D008962:Models, Theoretical; D009693:Nucleic Acid Hybridization; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D015345:Oligonucleotide Probes / Q000737:chemistry; D020641:Polymorphism, Single Nucleotide; D012680:Sensitivity and Specificity
D018076:DNA, Complementary; D005456:Fluorescent Dyes; D015345:Oligonucleotide Probes