title: Lamin A ChIP-seq in human dermal fibroblasts
design: For LMNA ChIP/seq, cells (10e7 per ChIP) were cross-linked with 1% formaldehyde, lysed in RIPA buffer (140 mM NaCl, 10 mM Tris-HCl, pH 8.0, 1 mM EDTA, 0.5 mM EGTA, 1% Triton X-100, 0.1% SDS, 0.1% Na-deoxycholate) adjusted to 1% SDS, and sonicated into chromatin fragments of 200-400 bp. Chromatin was incubated overnight with 50 microgram anti-lamin A/C antibody (Santa Cruz sc-7292) pre-coupled to magnetic Dynabeads Protein G. ChIP material was collected, washed, crosslink reversed and DNA eluted for 6 h on a shaker in elution buffer (50 mM NaCl, 20 mM Tris-HCl, pH 7.5, 5 mM EDTA, 1% SDS) containing RNase A and Proteinase K). DNA was purified by phenylchloroform isoamylalcohol extractino and ethanol precipitation, processed for library preparation (Illumina) and sequenced on an Illumina HiSeq2500. Input sequences corresponding to each cell type were also sequenced.
identifiers.namespace: University of Oslo
identifiers.id: LDF_LMNA