Fig. 5. Pho binds to the eve locus. (A) Map of the eve locus showing the location of primer sets (numbered black bars) used to analyze Pho binding. The location of the 3' PRE, PRE300, is shown as a yellow box. (B-G) Analysis of Pho binding in wild type (yw) 2-6 hours AED (after egg deposition, B-D) and 7-13 hours AED (E-G) embryos. For 2-6 hours AED embryos, eggs were collected for 4 hours and developed for 2 hours at room temperature; these embryos spanned stages 4-11. For 7-13 hours AED embryos, eggs were collected for 6 hours at room temperature and developed overnight at 17°C; these embryos spanned stages late 11 to early 16. (B,E) ChIP with Pho-specific antiserum (raised in rabbit). (C,F) Negative control: parallel analysis using normal rabbit serum. (D,G) Positive control: parallel PCR reactions using 1/300 of input material. Note that, in addition to positive signals from elsewhere in the locus, some of which change with time, Pho binding is specifically detected at the 3' PRE in both early and later-stage embryos.