Extended Data Fig. 6: Generation and analysis of Wapl mutant mice.
From: Wapl repression by Pax5 promotes V gene recombination by Igh loop extrusion
a, Mutations introduced at the Pax5-binding sites P1 and P2 in the Wapl∆P1,2, Wapl∆P1 and Wapl∆P2 alleles by CRISPR–Cas9-mediated mutagenesis. The consensus Pax5 motif and the extent of the deletions (red) are indicated. b, Loss of Pax5 binding at the mutated P1 site in Wapl∆P1/∆P1 pro-B cells. One of two ChIP-seq experiments is shown. c. Wapl mRNA expression in short-term-cultured Wapl∆P1,2/∆P1,2 and Wapl+/+ pro-B cells is shown as mean TPM value with s.e.m. n, number of RNA-seq experiments. d, Wapl protein expression in short-term-cultured Wapl∆P1,2/∆P1,2Rag2−/− and Wapl+/+Rag2−/− pro-B cells was determined by immunoblotting of twofold serially diluted whole-cell extracts with antibodies detecting Wapl or TBP. One of two experiments is shown with marker proteins (kilodaltons). e, Wapl mRNA expression in ex vivo-sorted Wapl∆P1,2/∆P1,2 (blue), Wapl∆P1,2/+ (light blue) and Wapl+/+ (black) pro-B and pre-B cells, Pax5∆/∆ progenitors (green; Vav-Cre Pax5fl/fl) as well as Wapl∆P1,2/∆P1,2 and Wapl+/+ T cell subsets from the thymus and spleen, as determined by RT–qPCR analysis relative to Tbp expression. The Wapl expression of each cell type is indicated relative to that of the Wapl+/+ cells (set to 1) and is shown as mean value with s.e.m., based on 2–4 independent RT–qPCR experiments for each cell type and genotype. Each dot (c, e) corresponds to one mouse. The different cell types are defined in Methods.
