All cell lines were cultured in DMEM medium with 10% fetal bovine serum, 100 U/ml penicillin and 100 mg/ml streptomycin (Millipore-Sigma) in a humidified incubator with 5% CO2 at 37 °C.
Initial experiments were performed on primary MEFs derived from 13.5 days NM1 WT and KO embryos46 (link). In other experiments were used immortalized MEFs (ATCC® CRL-2752) or cell lines derived from these immortalized cells.
The antibodies against H3K9me3 (ab8898), H3K27ac (ab4729), H3K4me1 (ab8895), H3K4me3 (ab8580), H3K9ac (ab10812), H3 (ab1791), γH2AX (ab2893), p53-K370 (ab183544), PCAF (ab12188), Set1 (ab70378), GAPDH (ab8245), and nonspecific rabbit IgG isotype control (ab37415) were purchased from Abcam (Cambridge, MA, USA). The antibody against NM1 has been previously characterized2 (link). Alexa Fluor 555 Goat Anti-Rabbit (ab150078) and Alexa Fluor 488 Goat Anti-Mouse (ab150117) were used as secondary antibodies for immunofluorescence, and horseradish peroxidase (HRP)-fused Goat Anti-Rabbit (ab6721) and Rabbit Anti-Mouse (ab6728) secondary antibodies for western blottings were purchased from Abcam. Hoechst 43222 (H1399) and ProLong Gold Antifade Mountant with 4′,6-diamidino-2-phenylindole (DAPI; P36931) were purchased from Invitrogen, Waltham, MA, USA. All antibodies have been used according to the manufacturers’ protocols.
Initial experiments were performed on primary MEFs derived from 13.5 days NM1 WT and KO embryos46 (link). In other experiments were used immortalized MEFs (ATCC® CRL-2752) or cell lines derived from these immortalized cells.
The antibodies against H3K9me3 (ab8898), H3K27ac (ab4729), H3K4me1 (ab8895), H3K4me3 (ab8580), H3K9ac (ab10812), H3 (ab1791), γH2AX (ab2893), p53-K370 (ab183544), PCAF (ab12188), Set1 (ab70378), GAPDH (ab8245), and nonspecific rabbit IgG isotype control (ab37415) were purchased from Abcam (Cambridge, MA, USA). The antibody against NM1 has been previously characterized2 (link). Alexa Fluor 555 Goat Anti-Rabbit (ab150078) and Alexa Fluor 488 Goat Anti-Mouse (ab150117) were used as secondary antibodies for immunofluorescence, and horseradish peroxidase (HRP)-fused Goat Anti-Rabbit (ab6721) and Rabbit Anti-Mouse (ab6728) secondary antibodies for western blottings were purchased from Abcam. Hoechst 43222 (H1399) and ProLong Gold Antifade Mountant with 4′,6-diamidino-2-phenylindole (DAPI; P36931) were purchased from Invitrogen, Waltham, MA, USA. All antibodies have been used according to the manufacturers’ protocols.