Embryo halves were then homogenized in homogenization buffer containing 15mM NaCl, 15mM TrisHCl pH 7.5, 60mM KCl, 1mM EDTA, and 0.1% Triton-X100, with 1mM DTT, 0.1 mM PMSF, and protease inhibit cocktail (Roche) added before use. After homogenization, 0.5% NP40 was added, and following a 5 minute incubation, samples were spun down at a low speed. Nuclei in the pellet were then washed once with the homogenization buffer containing 0.2M NaCl. The low speed centrifugation was repeated, and the recovered nuclei pellet was then re-suspended in nuclear lysis buffer (10 mM TrisCl, pH 7.9, 100 mM NaCl, 1 mM EDTA, 0.5 % Sarkosyl) +1%SDS, +1.5% sarkosyl. The chromatin was recovered by spinning the sample at full speed in a micro-centrifuge at 4oC for 1 hour and was re-suspended in a small volume of nuclear lysis buffer. Chromatin was sheared to an average size of 300bp using a Covaris sonicator (peak power= 140, duty factor = 10, cycle burst = 200, time = 3 minutes). Chromatin immunoprecipitation was performed using 72ng of chromatin and 1.5ug of an anti-Bicoid polyclonal antibody described previously (Li et al., 2008). The Bicoid antibody was coupled to Dynabead M-280 sheep anti-rabbit magnetic beads and the immunoprecipitation was conducted with the standard protocol from the manufacturer. DNA libraries for the chromatin immunoprecitation samples were prepared using the Rubicon genomics thru-plex DNA-seq kit using 16 PCR cycles for posterior and whole samples and 22 cycles for anterior, middle, and input samples.