Anti histone h3 antibody

The following antibodies were used: anti-PCNA (PC10) antibody (Santa Cruz Biotechnology, sc-56, 1:2000); anti-Ubiquityl-PCNA (Lys164) antibody (Cell Signaling, #13439, 1:1000); anti-FLAG antibody (Sigma, F3165, 1:1000); anti-V5 antibody (Sigma, V8137, 1:2000); anti-RFC3 antibody (Bethyl, A300-186A, 1:500); anti-RFC4 antibody (Abcam, ab182145, 1:1000); anti-RFC5 antibody (Abcam, ab79871, 1:500); anti-Lamin B1 antibody (Abcam, ab16048, 1:1000); anti-Histone H3 antibody (Merck-millipore, 07-690, 1:5000); anti-Strep-Tactin HRP Conjugate antibody (IBA-life sciences, 2-1502-001, 1:1000). The anti-human ATAD5 antibody (1:1000) was raised in rabbits using the N-terminal 1–297 amino-acid fragment and then affinity-purified^{3 (link)}.

Western blots were performed by the SDS-PAGE electrophoresis. Total cell extracts were prepared and fractionated by gel electrophoresis and transferred to nitrocellulose membranes. The following primary antibodies were used: Anti-KDM4D (ab93694; Abcam), anti-H3K9me3 (ab8898; Abcam), anti-Sav1 (105105; Abcam), Yap1 (ab81183; Abcam), pYap1 (ab76252; Abcam), Myc (ab32072; Abcam), Anti-Histone H3 Antibody (07–690 from Sigma-Aldrich), GAPDH (ab181603; Abcam). Horseradish peroxidase anti-rabbit (ab205718; Abcam) was used as the secondary antibody. The membranes were initially cut according to the molecular weight size of proteins. The signal was detected using the super-signal-enhanced chemiluminescence system (Pierce).

Aspirin (over 99% purity) and dimethyl sulfoxide (DMSO) were purchased from Sigma-Aldrich (St. Louis, MO, USA). Ultrapure lipopolysaccharide from Porphyromonas gingivalis (LPS) was purchased from InvivoGen (San Diego, CA, USA). Antibodies specific for COX₂, iNOS, p65 NF-κB, IκK, IκB, phosphorylated IκK (p-IκK), and p-IκB were purchased from Cell Signaling Technology (Boston, MA, USA). An anti-β-actin antibody, anti-Heat shock protein 90 (H90) antibody and anti-histone-H3 antibody were purchased from Sigma-Aldrich (St. Louis, MO, USA). The anti-MHC Class II(I-ab)-PE antibody was purchased from eBioscience (San Diego, CA, USA). The anti-F4/80-FITC antibody, FIZZ antibody, ARG1 antibody, YM-1 antibody were purchased from Abcam (Cambridge, Cambs, UK). A prostaglandin E2 receptor (EP₂) antagonist (C23H20FNO5) was purchased from TOCRIS (Bristol, C Cnty, UK). The thioglycollate was purchased from Aobox Biotechnology (Beijing, China).

The cytosolic and nuclear fractions were isolated essentially according to the protocol described previously (Shang et al., 2010 (link)). Ten-day-old Arabidopsis seedlings were ground to fine powder using liquid nitrogen and pre-chilled using a mortar and pestle. Cytosolic protein isolation buffer is composed of 10mM HEPES, pH 8.0, 250mM sucrose, 0.5% (v/v) Triton X-100, 1mM EDTA, 5mM MgCl₂, 50mM NaCl, 1mM phenylmethylsulphonyl fluoride (PMSF), and 1× Roche Cocktail (protease inhibitor cocktail). The buffer was added at 1ml g^–1 to powder to generate the homogenate. After centrifuging at 10 000 g for 15min, the supernatant was mixed with 2× SDS sample buffer and denatured for 10min in boiling water. The isolated cytosolic fraction was examined by immunodetecting the presence of the nuclear marker histone H3 with anti-histone H3 antibody (Sigma-Aldrich) to verify that the cytosolic fraction was not contaminated by the nuclear fraction. The nuclear fraction was isolated according to the protocol of Cold Spring Harbor Laboratory as described on its website, and examined by immunodetecting the presence of the cytosolic marker PEPC (phosphoenolpyruvate carboxylase) with anti-PEPC antibody (Agrisera) to ensure that the nuclear fraction was not contaminated by the cytosolic fraction.