title: Zfh1ChIP_IP_6-8h_zfh1RL_chrB
organism: Drosophila melanogaster
description: Protocols: Canton S wild-type Drosophila melanogaster were grown in population cages at 25 degr. Staged 2 hr collections of embryos were made on apple-agar plates and aged at 25 degr until the required stage of development. Embryos were dechorionated using 50% bleach and formaldehyde fixed in 10 ml cross-linking solution (50 mM Hepes, 1 mM EDTA, 0.5 mM EGTA, 100 mM NaCl, 1.8 % formaldehyde, pH 8.0) and 30ml n-heptane on a shaker table at RT for 15 minutes. The reaction was terminated with 125 mM glycine, 0.1% Triton X-100 in PBS. After washing out the fix, the embryos were blotted dry, snap frozen in liquid nitrogen, and stored at -80 degr. A small number of embryos from each collection was set aside for staging. Sandmann T, Jakobsen JS, Furlong EEM. ChIP-on-chip protocol for genome-wide analysis of transcription factor binding in Drosophila melanogaster embryos. Nat Protoc. 2006;1(6):2839-55. Libraries were prepared by using Illumina ChIP-Seq sample prep kit (part#11257047). ChIP-DNA was first end-repaired, added 3'-dA overhang and ligated with adapter. Adapter-modified DNA was then size-selected by electrophoresis, purified by gel purification (Qiagen part#28704), and enriched by PCR (and purified by using Qiagen Minielute PCR purification kit, part #28004). Size, purity and concentration of the library samples were examined by Agilent DNA 1000 kit with an Agilent Technologies 2100 Bioanalyzer.