Championchip one day kit

ChIP assay was performed, as described previously14 (link)17 (link), with SABiosciences Corporation’s ChampionChiP One-Day kit (Qiagen, Frederick, MD) following the manufacturer’s protocol, with some modifications. Briefly, pellets of 5 × 10⁶ cells were treated with fresh fixing buffer (1% formaldehyde) for 10 min at 37 °C to crosslink DNA and proteins. The reaction was terminated by the addition of stop buffer and incubated at room temperature for 5 min. After cell lysis, the cross-linked chromatin was sonicated to an average size of ∼500 bp and was immunoprecipitated with antibodies against TET1, GFI1 (Santa Cruz Biotechnology Inc., Santa Cruz, CA), the N′-terminal portion of MLL (MLL-N), the C′-terminal of MLL (MLL-C), H3K27Me3, H3K4Me3, RNA polymerase II, EZH2, SIN3A or IgG (Abcam, Cambridge, MA). Purified ChIP DNA was amplified by real-time qPCR using specific primers targeting the CpG-enriched upstream region of human miR-22: forward: 5′- GTTGTTGGAGTCGTGAGTG -3′; reverse: 5′- CGCTCCACCTTTCCTTAAA -3′; or mouse miR-22: forward: 5′- TGAATGGGCGGGAGTAA -3′; reverse: 5′- CCACGAGCTGCGAATGAA -3′.

ChIP assay was conducted as described previously^{50 (link)}, with SABiosciences Corporation’s ChampionChIP One-Day kit (Qiagen, Frederick, MD) following the manufacturer’s protocol. Chromatin from THP-1 cells were cross-linked, sonicated into an average size of ~500 bp, and then immunoprecipitated with antibodies against the N’-terminal of MLL (MLL-N), the C’-terminal of MLL (MLL-C), EZH2, SIN3A, H3K27Me3 or IgG (Abcam, Cambridge, MA). Purified DNA was amplified by real-time qPCR using primers targeting the promoter of ALOX5 as described before^{32 (link)}.

ChIP assay was conducted as described previously^{50 (link)}, with SABiosciences Corporation’s ChampionChIP One-Day Kit (Qiagen, Frederick, MD) following the manufacturer’s protocol. Chromatin from MONOMAC-6 cells were cross-linked, sonicated into an average size of ~500 bp, and then immunoprecipitated with antibodies against STAT3 (C-20, Santa Cruz Biotechnology, Dallas, TX), STAT5 (610191, BD Biosciences, San Jose, CA), TET1 (Y-14, Santa Cruz, Dallas, TX), or IgG (ab2410, Abcam, Cambridge, MA). Purified DNA was amplified by real-time qPCR using primers targeting the promoter of TET1 as described before^{19 (link)}. Sequences of qPCR primers for the TET1 promoter sites are: Site 1 forward: 5′-ACTTTGACCTCCCAAAGTGCTGGA-3′, reverse: 5′-ACCTGAGTGATGCTGAGACTTCCT-3′; Site 2 forward: 5′-TTTGGGAACCGACTCCTCACCT-3′, reverse: 5′-TCGGGCAAACTTTCCAACTCGC-3′; Site 3 forward: 5′-ACGCTGGGCATTTCTGATCCACTA-3′, reverse: 5′-TATTGTGCAGCTCGTTTAGTGCCC-3′; Site 4 forward: 5′-CCATCTCCCGACACACA-3′; reverse: 5′-TTGGCAGTGACCTTGAGA-3′.