Cell suspension was sonicated using Branson 250 Sonifier for 3 × 20 s cycles at 20% amplitude. 10ul of Turbo DNase (Ambion, AM2238) was added to sonicated material, incubated at 37°C for 10 minutes, and spun down at max speed for 10 minutes at 4°C. Protein-G beads were washed and pre-incubated with antibodies for 30 minutes at room temperature. Lysate and beads were incubated at 4°C for 2 hours. Beads were washed 3x using the following wash buffer (1x PBS, 0.1% SDS, 0.5% NP-40) followed by 2x using a high salt wash buffer (5x PBS, 0.1% SDS, 0.5% NP-40) and crosslinks were reversed and proteins were digested with 5ul proteinase-K (NEB, P8102S) at 65° for 2–4 hours. RNA was purified using phenol/chloroform/isoamyl alcohol and RNA was precipitated in isopropanol.
