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Evaluation of DNA microarray results with quantitative gene expression platforms


We have evaluated the performance characteristics of three quantitative gene expression technologies and correlated their expression measurements to those of five commercial microarray platforms, based on the MicroArray Quality Control (MAQC) data set. The limit of detection, assay range, precision, accuracy and fold-change correlations were assessed for 997 TaqMan Gene Expression Assays, 205 Standardized RT (Sta)RT-PCR assays and 244 QuantiGene assays. TaqMan is a registered trademark of Roche Molecular Systems, Inc. We observed high correlation between quantitative gene expression values and microarray platform results and found few discordant measurements among all platforms. The main cause of variability was differences in probe sequence and thus target location. A second source of variability was the limited and variable sensitivity of the different microarray platforms for detecting weakly expressed genes, which affected interplatform and intersite reproducibility of differentially expressed genes. From this analysis, we conclude that the MAQC microarray data set has been validated by alternative quantitative gene expression platforms thus supporting the use of microarray platforms for the quantitative characterization of gene expression.

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Figure 1: Effect of the number of transcript molecules on assay precision.
Figure 2: Analysis of assay accuracy.
Figure 3: Correlation of fold change between alternative quantitative platforms.
Figure 4: Performance of microarray platforms relative to alternative quantitative platforms.
Figure 5: Assessment of true positive rates and false discovery rates using TaqMan assays.
Figure 6: Resolution of fold-change discrepancy results.

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We would like to acknowledge the contribution to this manuscript from the following members of the MAQC team: Shawn B. Baker, Anne Bergstrom Lucas, Jim Collins, Eugene Chudin, Stephanie Fulmer-Smentek, Damir Herman, Richard Shippy, Chunlin Xiao and Necip Mehmet.

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Correspondence to Federico M Goodsaid.

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J.C.W. is a consultant for and has significant financial interest in Gene Express, Inc.

Supplementary information

Supplementary Fig. 1

TaqMan® assays SD vs. CT plot. (DOC 31 kb)

Supplementary Fig. 2

TaqMan® assays R2 distribution plot. (DOC 166 kb)

Supplementary Fig. 3

Boxplot of fold change as a function of signal. (DOC 43 kb)

Supplementary Fig. 4

Expression characteristics of EPHA7. (DOC 91 kb)

Supplementary Fig. 5

Endogenous control expression. (DOC 204 kb)

Supplementary Table 1

Gene lists used for the analysis of performance metrics of alternative quantitative platforms and for comparison with microarrays. (DOC 51 kb)

Supplementary Table 2

Inter-site discordance in detection among genes expressed at low level. (DOC 156 kb)

Supplementary Table 3

TPR, FDR tables. (DOC 100 kb)

Supplementary Table 4

Discordant probes among the 997 genes. (DOC 36 kb)

Supplementary Table 5

Discordant gene expression values in alternative quantitative platforms FC expression values for all discordant genes. (DOC 44 kb)

Supplementary Methods (DOC 48 kb)

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Canales, R., Luo, Y., Willey, J. et al. Evaluation of DNA microarray results with quantitative gene expression platforms. Nat Biotechnol 24, 1115–1122 (2006).

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