Fig. 2.

NGS of GC B-cell V_H genes from B6 WT versus mA3 KO mice. (A) Sorting strategy. RNA extracted from CD19⁺GL7⁺ cells were used for cDNA synthesis followed by IgG V_H PCR with Illumina primers. (B) Sequence read counts from individual WT and KO mice. To minimize selection bias, identical sequences were collapsed into a single unique sequence. FV⁺ V_H corresponds to the 16 V_H genes in the FV-specific mAbs in Fig. 1G. (C) Validation of sorted GC B cells. The average mutation frequencies of paired GL7⁺ and GL7⁻ populations from the three WT mice were computed for each of the 16 V_H genes. Differences were evaluated using a two-tailed paired Student t test. (D) Total mutation frequency in WT (n = 3) and KO (n = 4) mice for V_H1–19 and V_H1–26 sequences. Differences were evaluated using a two-tailed unpaired Student t test with P < 0.05 considered as significant.