Illumina HiSeq 2000 sequencing; Histone modifications along transdifferentiation from B cell to macrophage
Histone modifications along transdifferentiation from B cell to macrophage
BLaER1 cells at different time points after trasndifferentiation induction were fixed for 10 minutes at room temperature with 1% formaldehyde. Chromatin was sheared by using a Covaris ultrasonicator until fragments ranged from 200 to 1,000 bp. Chromatin immunoprecipitations were performed with 2 ug of chromatin and following standard procedures. BLaER1 cells were grown in RPMI 1640 medium (Gibco) supplemented with 10% inactivated Foetal Bovine Serum (Gibco), 5% Penicillin-Streptomycin (Gibco) and 5% L-Glutamine (Gibco). Cells were maintained at 37ºC in a humid atmosphere containing 5% CO2. Transdifferentiation was induced by addition of 100 nM of beta-estradiol (Calbiochem) and 10 nM of IL-3 and CSF-1 (Peprotech). Immunoprecipitated chromatin was purified with Qiagen PCR purification columns, following the manufacturer's instructions. Sequencing libraries were performed with I ug of ChIP'ped DNA with NebNext ChIP-Seq library kit, following the manufacturer's instructions.
Experimental Factor: stimulus
none
Experimental Factor: time
0
Experimental Factor: immunoprecipitate
anti-H3K4me1