Fig. 6: Meiotic phenotype of Brca2 exon 12 deletion mouse model.

a Schematic depiction of the domain composition of the BRCA2 protein and the exons 11-15 encoding HSF2BP-binding and DMC1-binding domains. Introns are not drawn to scale; different exon phases are indicated by the shape of the boundary. Location of Cas9 cut sites for exon 12 and exons 12–14 excision is shown. b RT-PCR on cDNA from mouse testis with indicated genotypes confirming the loss of exon 12; primer locations are shown on the exon scheme in a. The PCR was performed twice with the same results. c Immunoblot analysis of proteins precipitated from Brca2^+/+ and Brca2^∆12/∆12 mouse testes using anti-HSF2BP antibody and, as control, with anti-RAD51 antibodies; and the input samples; performed as described in “Methods” section. The experiment was performed four times with similar results. d testis weight, e bodyweight, f sperm count, and g epididymis weight in Brca2^∆12/∆12 and control mice. n = 5 animals for Brca2^+/+, n = 6 for Brca2^∆12/∆12, n = 3 (testis weight and bodyweight), and n = 2 (epididymis weight and sperm count) for Brca2^∆12/+. Mean, s.e.m, and p values from one-way ANOVA with Tukey test are indicated. h–n Immunofluorescent analysis of meiotic protein localization on spermatocyte spreads from Brca2^∆12/∆12 and control mice. Representative images (h, l; scale bars = 5 µm) and quantification of RAD51 (i), DMC1 (j), MLH1 (k), HSF2BP (m), and BRME1 (n) foci are shown with mean and s.e.m. indicated with error bars. Symbol shapes designate individual animals: n = 3 animals for Brca2^+/+ and Brca2^∆12/∆12, n = 2 for Brca2^∆12/+, n = 2 (BRME1), and n = 1 (HSF2BP) for Hsf2bp^−/−. Mean, s.e.m, and p values from two-tailed unpaired t-test for selected pairwise comparisons within prophase stages are indicated. The number of analyzed nuclei is indicated for each genotype and stage.