Validation of gene expression changes by quantitative PCR. (A) Marker expression in the learning cohort. Fifteen differentially expressed genes identified by microarray analysis were re-examined by real-time PCR in 4 healthy controls (C) and 7 patients with polycythemia vera (P). Boxes represent the interquartile range that contains 50% of the values; the horizontal line in the box marks the median and bars indicate the range of values. Relative gene expression is shown on a logarithmic scale (log₁₀) and the values represent the mRNA expression relative to a healthy control calibrator sample, calculated by the ΔΔC_T method. All 15 genes examined showed a statistically significant difference (P < .05; Mann-Whitney U test) between controls and PV. (B) Marker expression in the test cohort, consisting of 28 healthy controls (C), 47 patients with polycythemia vera (P), 33 patients with essential thrombocythemia (E), 11 patients with idiopathic myelofibrosis (I), 4 patients with secondary erythrocytosis (S), and 4 patients with secondary neutrophilia (N). Annotation as indicated in panel A.