Commentary on 'Untargeted CUT&Tag reads are enriched at accessible chromatin and restrict identification of potential G4-forming sequences in G4-targeted CUT&Tag experiments'.

In a recent study (Nucleic Acids Research, Volume 53, Issue 14, 12 August 2025, gkaf678), Thompson and Byrd examine potential issues in the mapping of DNA G-quadruplexes (G4s) by CUT&Tag and related methods [1]. The authors conclude that detection of G4s in chromatin is significantly confounded by artefacts arising from background Tn5 enzymatic activity independent of the primary G4-targeting probe. We highlight several concerns regarding the authors’ meta-analysis of published data, including the problematic use of computational tools developed initially for other experimental modalities, such as ATAC-seq and ChIP-seq, for G4-CUT&Tag data without explicit disclosure of the analytical methods and code. Given that CUT&Tag is widely used and the technology is rapidly evolving, it is timely to promote best practices in the profiling of G4s and other features.