Figure 2 |.

Use of proximal and distal DHS read enrichment to determine optimal DNase I enzyme concentration. (a) Plot of the relative enrichment of normalized read density near transcription start sites (TSSs) that represent reads from proximal DHSs. 5,000 human CD8+ cells were collected by FACS sorting. DNase I was diluted as indicated to generate an enzyme concentration titration. Suboptimal concentrations (0.4 U/μl and above) have a broader distribution of reads near the TSS. The highest concentrations (0.5–1 U/μl) result in overdigestion of chromatin and show a depletion of reads proximal to the TSS. (b) Enrichment of reads near H3K27ac peaks, which serve as a marker of active distal enhancers, indicates that DNase I enzyme concentrations of 0.2–0.4 U/μl generate the highest enrichment at distal DHSs. On the basis of enrichment at proximal and distal DHSs, a DNase I enzyme concentration of 0.2–0.3 U/μl is optimal.