design: ChIP-seq of ERα in MCF7 cells treated with the hypoxia mimetic DMOG versus vehicle control
library_construction_protocol: MCF7 cells (RRID:CVCL_0031) were cultured in DMEM (Gibco) with 10% FBS. Cells were seeded at 2 × 10⁶ per 15 cm dish and grown for 3 days. On day 4, media was replaced and cells were treated with 2 mM DMOG (or vehicle control). After 16 hours, cells were washed with PBS and crosslinked using 1% formaldehyde. Following immunoprecipitation, protein–DNA complexes were eluted, reverse crosslinked, and treated with RNase and proteinase K. DNA was purified using phenol-chloroform extraction and ethanol precipitation. ChIP DNA was used as input for library prep using the NEBNext Ultra II DNA Library Prep Kit, including end repair, adaptor ligation, size selection, and PCR amplification.