Journal: Genome Biology

Article Title: NicE-seq: high resolution open chromatin profiling

doi: 10.1186/s13059-017-1247-6

Figure Lengend Snippet: Nicking enzyme-mediated labeling of open chromatin. a Nicking of crosslinked chromatin using varying amounts of Nt.CviPII. A 1% agarose gel showing differential nicking of HCT116 genomic DNA based on amount of nicking enzyme (10 U, 5 U, 2.5 U, 1 U, 0.3 U, 0 U). M is a DNA molecular weight ladder. b Open chromatin labeling in fixed HeLa cells using dNTPs supplemented with TexasRed-dATP. Top panel : labeling reaction performed in the presence of Nt.CviPII and DNA polymerase I. Middle panel : labeling reaction performed in the presence of DNA polymerase I only. Bottom panel : labeling reaction performed in the absence of Nt.CviPII and DNA polymerase I. TexasRed-dATP was included in all reactions. DNA staining was performed using DAPI ( blue ) and TexasRed stain ( red ) represents labeled OCSs. Magenta stain ( Merge ) represents the colocalization. c Labeling efficiency of OCSs in all three assayed conditions. The y-axis represents the ratio of the intensity of the red pixels to the intensity of the blue pixels ( OCS labeling efficiency ). d Dot blot showing labeling of open chromatin by Nt.CviPII nicking enzyme in both native and formaldehyde-fixed HCT116 cells. The level of labeling was revealed using HRP-conjugated goat anti-biotin antibody

Article Snippet: Membrane was blocked by 5% non-fat milk and probed using an HRP-conjugated goat anti-biotin antibody (1:2000 dilution, CST, 7075).

Techniques: Labeling, Agarose Gel Electrophoresis, Molecular Weight, Staining, Dot Blot