Fig. 3.

The effect of data normalization in differential expression following recovery sleep. a Bar graph displaying the number of up and downregulated probesets detected at each time point relative to time-of-day matched controls following RMA (light grey) or RUV (dark grey) normalization. RUV normalization profoundly affects the detection of differentially expressed genes following various lengths of recovery sleep. Positive controls for genes differentially expressed following 1 h of recovery sleep after were obtained as detailed in Methods. b Heatmap of differentially expressed probesets detected using RUV normalization relative to circadian time-matched controls. In red, upregulated genes. In green, downregulated genes. Clustering based on patterns of gene expression is represented by the dendrogram and color coded. Genes responding within 1–3 h to recovery sleep are indicated by black bars (fast responders), while genes that respond at 6 h are indicated by grey bars (slow responders, grey bar). Genes upregulated by sleep deprivation show two different patterns of response within the first three hours (red and pink clusters, black bar) and two different patterns of recovery at 6 h (green and orange clusters, grey bar). The majority of genes upregulated by sleep deprivation respond slowly with recovery sleep. The majority of genes downregulated are fast responders (mint green, black bar), while a very small proportion recovers within 6 h (lilac, grey bar). SD, sleep deprivation; RS1, sleep deprivation followed by 1 h of recovery sleep; RS2, sleep deprivation followed by 2 h of recovery sleep; RS3, sleep deprivation followed by 3 h of recovery sleep; RS6, sleep deprivation followed by 6 h of recovery sleep