Fig. 3.

Changes at the promoter regions of differentiation genes as spermatogonia become spermatocytes. Real-time PCR analysis of the enrichment of three tTAF target genes (Mst87F, dj and fzo) and one non-target gene (CycA) by ChIP using (A) anti-H3K4me3, (B) anti-RNA polymerase II (Pol II) (8WG16) specific for the form of Pol II in the preinitiation complex, which has an unphosphorylated C-terminal domain (CTD), (C) anti-Pol II (4H8) that recognizes both the unphosphorylated and Ser5 phosphorylated CTD of Pol II, (D) anti-H3K27me3, and (E) anti-Polycomb (Pc) from bam, aly, can or wild-type (wt) testes. To compare values across different genotypes, for each data point the level of ChIP DNA (ChIP DNA/input) at the target gene (Mst87F, dj or fzo) was first normalized to the level of ChIP DNA at the CycA gene from the same sample, and then the normalized values from at least three independent ChIP reactions per genotype were averaged. ChIP values for antibodies against modified histone H3 in A (H3K4me3) and D (H3K27me3) were further normalized to values from ChIP with anti-H3 to control for possible variation in nucleosomal distribution. **, P<0.01; *, P<0.05; #, P>0.1 (one sample t-test). Error bars indicate s.d. from three independent biological replicates.