Fig. 6.
γH2AX and RAD51 localization in cisplatin-treated and untreated Mei1m1Jcs/Mei1m1Jcs spermatocytes. RAD51 foci (green) are absent from Mei1m1Jcs/Mei1m1Jcs zygotene spermatocytes (B) but appear after cisplatin treatment (C). A wild-type untreated control is shown in A. Localization of γH2AX antibody (green) in Mei1m1Jcs/Mei1m1Jcs leptotene (E) spermatocytes is greatly reduced compared with wild type (D) and comparable to that of Spo11–/– spermatocytes (F). In all cases, spreads were colabeled with anti-SCP3 (red). To quantitate the data represented in D–F, average fluorescence across each of 15 anti-γH2AX labeled nuclei (n = 15) was calculated for each genotype by using the metamorph imaging system (G). The amount of γH2AX in Mei1m1Jcs/Mei1m1Jcs and Spo11–/– spermatocyte nuclei was reduced 4-fold when compared with that of normal spermatocytes (P < 0.0001). Unpaired Student t tests and ANOVA were used to calculate P values. Fig. 7 shows, for each genotype, three images for each of five nuclei: anti-SCP3 staining alone, anti-γH2AX alone, and merged.