title: Human_012h Input replicate 2
organism: Homo sapiens
description: Protocols: BLaER1 cells are as described here: https://doi.org/10.1016/j.celrep.2013.03.003 and are derived as a subclone of Seraphina cells CVCL_M646. Cells were collected after 12 hours of transdifferentiation. To track the transdifferentiation process along the 7 days it lasts, fluorescent staining of cell surface markers was done with antibodies against Mac1 (APC) and CD19 (APC-Cy7) (BD Pharmigen). Samples were analyzed on the LSRII flow cytometer (BD Biosciences) and data were analyzed with FlowJo software (Tree Star). Human BLaER1 cells and mouse C10 cells were grown in RPMI (Invitrogen) supplemented with 10% heat-inactivated fetal bovine serum, 2 mM L-glutamine, and 100 U/ml penicillin G sodium. Transdifferentiation was induced in BlaER1 by addition of 100nM of β-estradiol (Calbiochem) and grown with 10nM of IL-3 and CSF-1 (Peprotech). Cells were crosslinked with 1% formaldehyde for 10 minutes at room temperature and sheared with a Covaris sonicator. 5 μg of chromatin and 5 μg of antibody against rat CEBPA (sc-61 X, Santa Cruz) were incubated in RIPA buffer (140 mM NaCl, 10 mM Tris HCl pH 8.0, 1 mM EDTA, 1 % Triton X-100, 0.1 % SDS, 0.1 % Na deoxycholate, protease inhibitors). DNA libraries were prepared with 1 ng of purified ChIP using NEBNext DNA Library kit for Illumina following the manufacturer's protocol.