Untargeted CUT&Tag and BG4 CUT&Tag are both enriched at G-quadruplexes and accessible chromatin.

G-quadruplex DNA structures (G4s) form within single-stranded DNA in nucleosome-free chromatin. As G4s modulate gene expression and genomic stability, genome-wide mapping of G4s has generated strong research interest. Recently, the Cleavage Under Targets and Tagmentation (CUT&Tag) method was performed with the G4-specific BG4 antibody to target Tn5 transposase to G4s. While this method generated a novel high-resolution map of G4s, we unexpectedly observed a strong correlation between the genome-wide signal distribution of BG4 CUT&Tag and accessible chromatin. To examine whether untargeted Tn5 cutting at accessible chromatin contributes to BG4 CUT&Tag signal, we examined the genome-wide distribution of signal from untargeted (i.e. negative control) CUT&Tag datasets. We observed that untargeted CUT&Tag signal distribution was highly similar to both that of accessible chromatin and of BG4 CUT&Tag. We also observed that BG4 CUT&Tag signal increased at mapped G4s, but this increase was accompanied by a concomitant increase in untargeted CUT&Tag at the same loci. Consequently, enrichment of BG4 CUT&Tag over untargeted CUT&Tag was not increased at mapped G4s. These results imply that either the vast majority of accessible chromatin regions contain mappable G4s or that the presence of G4s within accessible chromatin cannot reliably be determined using BG4 CUT&Tag alone.