Short oligonucleotide probes containing G-stacks display abnormal binding affinity on Affymetrix microarrays.
BACKGROUND
In microarray experiments, probe design is critical to the specific and accurate measurement of target concentrations. Current designs select suitable probes through in silico scanning of transcriptome/genome based on first principles. However, due to lack of tools, the observed microarray data have not been used to assess the performance of individual probes to provide feedback to improve future designs.


RESULTS
In this study, we describe a probe performance assessment method based on the concordance of the observed signals from probes that share common targets. Using this method, we found that probes containing multiple guanines in a row (G-stacks) have abnormal binding behavior compared with other probes, both in gene expression assays and genotyping assays using Affymetrix microarrays. These probes are less likely to covary with other probes that interrogate the same genes. Moreover, we found that these probes are much more likely to produce outliers when fitting the observed signals according to the positional dependent nearest neighbor model, which gives reasonable estimates of binding affinity for most other probes. These results suggest that probes containing G-stacks tend to have increased cross hybridization signals and reduced target-specific hybridization signals, presumably due to multiplex binding forming G-quartet structures. Our findings are expected to be useful in microarray design and data analysis.
Wu
Chunlei
C
Genomic Institute of Novartis Research Foundation, 10675 John Jay Hopkins Dr, San Diego, CA 92121, USA.
Zhao
Haitao
H
Baggerly
Keith
K
Carta
Roberto
R
Zhang
Li
L
D000345:Affinity Labels; D000465:Algorithms; D001483:Base Sequence; D018390:Gene Targeting / Q000379:methods*; D017403:In Situ Hybridization / Q000379:methods*; D008969:Molecular Sequence Data; D020411:Oligonucleotide Array Sequence Analysis / Q000295:instrumentation / Q000379:methods*; D015345:Oligonucleotide Probes / Q000235:genetics*; D011485:Protein Binding; D016415:Sequence Alignment / Q000379:methods; D017422:Sequence Analysis, DNA / Q000379:methods*; D012984:Software
D000345:Affinity Labels; D015345:Oligonucleotide Probes