Western blot was performed on whole cell (25 μg, lane 1) and histone extracts (15 μg, lane 2) from HeLa cells, and on 1 μg of recombinant histone H3 (lane 3) using H3K79me1 Polyclonal Antibody (Cat. No. bs-53126R). The antibody was diluted 1:200 in TBS-Tween containing 5% skimmed milk.


A Dot Blot analysis was performed to test the cross-reactivity of the Bioss antibody against H3K79me1 (Cat. No. bs-53126R) with peptides containing other histone modifications and the unmodified H3K79. One hundred to 0.2 pmol of the respective peptides were spotted on a membrane. The antibody was used at a dilution of 1:20,000.


To determine the titer of the antibody, an ELISA was performed using a serial dilution of the Bioss antibody directed against H3K79me1 (Cat. No. bs-53126R). The antigen used was a peptide containing the histone modification of interest. By plotting the absorbance against the antibody dilution, the titer of the antibody was estimated to be 1:11,500.


ChIP was performed with 1 μg of the Bioss antibody against H3K79me1 (Cat. No. bs-53126R) and the IP’d DNA was subsequently analyzed on an Illumina Genome Analyzer. Library preparation, cluster generation, and sequencing were performed according to the manufacturer’s instructions. The 36 bp tags were aligned to the human genome using the ELAND algorithm. The figure shows the peak distribution along the complete sequence and a 1 Mb region of the X-chromosome (figure A and B), in 100 kb regions surrounding the GAPDH positive control and EIF4A2 genes (figure C and D).