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Mangalathu S. Rajeevan, Suzanne D

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1 Validation of Array-Based Gene Expression Profiles by Real-Time (Kinetic) RT-PCR 
Mangalathu S. Rajeevan, Suzanne D. Vernon, Naovarath Taysavang, Elizabeth R. Unger  The Journal of Molecular Diagnostics  Volume 3, Issue 1, Pages (February 2001) DOI: /S (10) Copyright © 2001 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

2 Figure 1 Scatterplot of the log base 2 of the hybridization intensities for each gene in subclones and as determined by HDFA. Perpendicular lines (dashed) indicate the threshold values for each subclone. The two solid parallel lines indicate the position of twofold differences in intensity. Genes on and outside the parallel lines were identified as differentially expressed by HDFA. The Journal of Molecular Diagnostics 2001 3, 26-31DOI: ( /S (10) ) Copyright © 2001 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

3 Figure 2 Fluorescence versus cycle number plot of G3PDH transcript in subclones and by LightCycler RT-PCR assay based on SYBR Green I dye detection. Both subclones were evaluated for relative levels of G3PDH transcript using 1:200, 1:2000, and 1:20,000 dilutions of cDNA which are indicated as A, B, and C, respectively, in the amplification plot. There were four reactions per dilution per sample. At each dilution, all reactions overlapped in the log-linear phase, indicating identical levels of G3PDH transcript in both samples.Closed arrowheads indicate no-RT reactions with both subclones. Open arrowhead shows PCR with no template.Striped arrow indicates the position of noise band. All data points below the noise band were excluded from quantification. Annealing and signal acquisition temperatures for G3PDH are given inTable 1. The Journal of Molecular Diagnostics 2001 3, 26-31DOI: ( /S (10) ) Copyright © 2001 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

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