title: Illumina NovaSeq 6000 paired end sequencing; ChIP-seq of CSF-1R in THP-1 cell lines deleted for EGR1 gene by CRISPR-Cas9 and ChIP-seq of CSF-1R in 2 CMML patients
design: ChIP-seq of CSF-1R in THP-1 cell lines deleted for EGR1 gene by CRISPR-Cas9 and ChIP-seq of CSF-1R in 2 CMML patients
library_construction_protocol: Cells were cross-linked with addition of 1% formaldehyde directly to the culture medium for 10 min at RT with agitation. Fixation was stopped by addition of 125mM glycin during 5 min at RT with agitation and samples were then washed 2 times in ice-cold PBS before addition of SDS lysis buffer (Millipore, 10uL per 1.106 cells) supplemented with 1% protease inhibitor cocktail (Active Motif).  Samples were vortexed and incubated 15 min on ice before 10 min sonication at 40W (Covaris S220, Woodingdean, UK). The chromatin immunoprecipitation was carried out using ChIP-it express kit according to manufacturer's instruction (Active Motif) with a monoclonal anti-CSF-1R antibody (sc-46662, santa cruz bioechnology). Enriched DNA from ChIP and Input DNA fragments were end-repaired, extended with an 'A' base on the 3′end, ligated with indexed paired-end adaptors (NEXTflex, Bioo Scientific, Proteigene, Saint Marcel, France) using the Bravo Platform (Agilent, Les Ulis, France), size-selected after 4 cycles of PCR with AMPure XP beads (Beckman Coulter, Villepinte, France) and amplified by PCR for 10 cycles more.
attributes.tag: Experimental Factor: genotype
attributes.value: wild type genotype
attributes.tag: Experimental Factor: disease
attributes.value: chronic myelomonocytic leukemia
attributes.tag: Experimental Factor: immunoprecipitate
attributes.value: anti-CSF-1R (Santa Cruz Biotechnology)