Solving the riddle of the bright mismatches: hybridization in oligonucleotide arrays

Abstract

HDONA technology is predicated on two ideas. First, the differential between high-affinity (perfect match, PM) and lower-affinity (mismatch, MM) probes is used to minimize cross-hybridization. Second, several short probes along the transcript are combined, introducing redundancy. Both ideas have shown problems in practice: MMs are often brighter than PMs, and it is hard to combine the pairs because their brightness often spans decades. Previous analysis suggested these problems were sequence-related; publication of the probe sequences has permitted us an in-depth study of this issue. Our results suggest that fluorescently labeling the nucleotides interferes with mRNA binding, causing a catch-22 since, to be detected, the target mRNA must both glow and stick to its probe: without labels it cannot be seen even if bound, while with too many it won't bind. We show that this conflict causes much of the complexity of HDONA raw data, suggesting that an accurate physical understanding of hybridization by incorporating sequence information is necessary to perfect microarray analysis.

4 figures

References in corpus (2)

• DNA hybridization to mismatched templates: a chip study
• Effect of defects on thermal denaturation of DNA Oligomers

Cited by in corpus (14)

• Brush Effects on DNA Chips: Thermodynamics, Kinetics and Design Guidlines
• The effects of mismatches on hybridization in DNA microarrays: determination of nearest neighbor parameters
• Specific and non specific hybridization of oligonucleotide probes on microarrays
• Thermodynamics of RNA/DNA hybridization in high density oligonucleotide microarrays
• A statistical framework for the analysis of microarray probe-level data
• Breakdown of thermodynamic equilibrium for DNA hybridization in microarrays
• Understanding the physics of oligonucleotide microarrays: the Affymetrix spike-in data reanalysed
• Physics-based analysis of Affymetrix microarray data
• Comment on: ``Solving the riddle of the bright mismatches: Labeling and effective binding in oligonucleotide arrays"
• Modelling background intensity in Affymetrix Genechips
• Effective affinities in microarray data
• Probabilistic analysis of the human transcriptome with side information
• Linear model for fast background subtraction in oligonucleotide microarrays
• High-Throughput SNP Genotyping by SBE/SBH