(a) Flow cytometry of Bach2-reporter expression in NP-specific IgG1+ LZ GC B cells (NP+B220+IgG1+CD138−CD38−CXCR4loCD86hi) from NP-CGG immunized Bach2-reporter mice at day 10 (upper left). VH186.2 sequences analysis of Bach2-reporter expression low- and high-NP-specific IgG1+ LZ GC B cells (lower left and right). The pie charts and the numbers of mutations are described as in Fig. 1c. Gray histograms represent the non-Bach2-reporter wild-type control. (b) Venn diagrams illustrating the overlap between the high-affinity population (magenta) and Bach2-reporter low-population (green) (left) or between the low-affinity population (orange) and Bach2-reporter high-population (blue) (right). The lists of up-regulated genes for each population were from the differentially expressed genes (FDR <0.05) between the high- and low-affinity populations or between the Bach2-reporter low- and high-populations. In the case of Fig. 2b analysis, we selected up the genes whose difference was more than 2-fold. (c) Heatmaps between Bach2-reporter low- and high-populations are shown as normalized expression (log2) of selected genes which were listed in Fig. 2c. (d) qRT-PCR analysis of the indicated genes in Bach2-reporter low- and high-NP-specific IgG1+ LZ GC B cells.*p < 0.05, **p < 0.01 and ***p < 0.001 (unpaired Student’s t-test). Data are representative of three independent experiments (a; mean), are from one experiment with three biological replicates (b and c; mean in b) and are pooled from three independent experiments (d; mean and s.d. n=3 samples (each prepared from 2 to 3 mice)).