RNA-Seq and real-time PCR validation in Tcl-1 tg IRF4 WT and deficient CLL cells. (A) Number of significantly downregulated (negative y-axis, green bars) and upregulated genes (positive y-axis, red bars) corresponding to enriched GO terms (x-axis) in Tcl-1 tg IRF4 ΔB/ΔB (N = 3) as compared with Tcl-1 tg IRF4^+/+ mice (N = 4). GO term descriptions and P values are indicated in the table. (B) Heatmap of differentially regulated genes within the GO term 0042110 T-cell activation. The counts per million were used for heatmap generation; unsupervised clustering (Euclidean) was used. (C) Validation of RNA-Seq data by real-time PCR in purified CLL cells derived from the spleen of Tcl-1 tg IRF4^+/+ (red) and Tcl-1 tg IRF4 ΔB/ΔB (blue) mice. PCR was performed with TaqMan primers for the MHC1 genes H2-K1 and D1, (D) the MHC2 genes H2-Dmb and H2-Aa, (E) the MHC2 transactivator CIITA, and (F) the costimulatory molecules CD80 and CD86. Real-time PCR data were normalized to 18S and the mRNA relative expression ratio calculated according to the delta CT method.