GSM6944491: B16F02L1, SX682, TFCP2L1; Mus musculus; ChIP-Seq
Next, cells were fixed with fresh formaldehyde at final concentration of 1% for 10 min at room temperature. Glycine was added to final concentration of 0.125 M to quench the reaction. Cells were resuspended at 1x107/ml in sonication buffer (10 mM Tris-HCl pH 8.0, 2 mM EDTA pH 8.0, 1% SDS, with Protease Inhibitor Cocktail (Roche) and PhosSTOP™ Phosphatase Inhibitor Cocktail (Roche) and sonicated for 25-30 sec with Diagenode One sonicator in a 50 ul Bioruptor® One Microfluidic Chip till most DNA fragments were in the range of 200 to 700 bp. After sonication, the lysate was adjusted to RIPA buffer conditions (10 mM Tris-HCl pH 8.0, 1 mM EDTA pH 8.0, 140 mM NaCl, 1% Triton X-100, 0.1% SDS, 0.1% sodium deoxycholate, with Protease Inhibitor Cocktail and PhosSTOP™ Phosphatase Inhibitor Cocktail). For each immunoprecipitation, lysate from 2x106 cells and 6 μg of anti-TFCP2L1 antibody (Boster Bio) were used. 6.0 μg of normal rabbit IgG was used as a control. The final enriched DNA library for each immunoprecipitation was created by PCR with KAPA HiFi HotStart Ready Mix and Nextera custom primers using the optimum number of PCR cycles determined by qPCR.