title: Illumina Genome Analyzer II sequencing; Glucocorticoid receptor profiling in IMR90,K562 and U2OS (ChIP-seq)
design: Glucocorticoid receptor profiling in IMR90,K562 and U2OS (ChIP-seq)
library_construction_protocol: Cells were treated with 0.01% ethanol vehicle or 1 μM dexamethasone for 1 hour. Cells were cross-linked by adding formaldehyde to a final concentration of 1% and subsequent incubation 3 minutes at room temperature before quenching the reaction by adding glycine to a final concentration of 125 mM. Chromatin was sheared with a Bioruptor water bath sonicator (Diagenode) to produce fragments of ∼100-200 bp. Protein G-coupled magnetic beads (dynabeads, Invitrogen) were preincubated for one hour with GR-antibody (N499) before chromatin was added and incubated for an additional 2-4 h while rotating at 4°C. Subsequently, beads were washed 4 times with 10mM Tris-HCl pH 8.0, 1mM EDTA, 500 mM NaCl, 5% Glycerol, 0.1% Sodium deoxycholate, 0.1% SDS, 1% Triton X-100, 0.5 mg/μl BSA followed by 4 additional washes with 20mM Tris, pH 8.0, 1 mM EDTA, 250 mM LiCl, 0.5% NP-40, 0.5% sodium deoxycholate.
attributes.tag: Experimental Factor: immunoprecipitate
attributes.value: input DNA
attributes.tag: LIBRARY_STRAND
attributes.value: not applicable