Chromatin immunoprecipitation (ChIP) analysis of C/EBPα and C/EBPε during myeloid differentiation.

(A) Chromatin immunoprecipitations were performed from uninduced (0) and ATRA-induced (24 h) MPRO cells using antibodies specific for C/EBPα (lanes 2-3) and C/EBPε (lanes 4-5), a no-antibody control (−) (lanes 1,9), and preimmune serum controls (PIS) (lanes 6-7). The precipitated chromatin was analyzed using primers specific for the murine LF (mLF) C/EBP site. Input mLF chromatin (1:10 dilution) is represented in lane 8. (B) ChIP analysis was performed using uninduced (0) and 4-day Epo-induced (4d) 32Dwt18 cells and antibodies specific for C/EBPα (lanes 2-3) and C/EBPε (lanes 5-6), no-antibody controls (lanes 1, 4, and 10), and preimmune serum (PIS) controls (lanes 7-8). Precipitated chromatin was analyzed using primers for the murine LF (mLF) promoter. Input mLF chromatin (1:10 dilution) is represented in lane 9. M indicates molecular weight markers. (C) ChIP analysis of uninduced (0) and ATRA-induced (24 h) MPRO cells using C/EBPα antibodies (lanes 1-2), C/EBPε antibodies (lanes 3-4), and a no-antibody control (lane 5). Precipitated chromatin was analyzed by PCR using primers specific for murine neutrophil elastase (mNE). Each ChIP experiment was performed 2 to 3 times. All PCR products were subcloned and sequenced by dideoxy method to confirm their identities.