The main controversy over DNase-seq is the ability for DNase I to introduce cleavage bias [31, 79, 81, 82, 84], thus affecting its use as a reliable TF footprint detection assay. Two recent publications clearly demonstrate that cleavage signatures traditionally attributed to protein protection of underlying nucleotides, are detected even in the absence of TF binding as a result of DNase I inherent sequence preferences that span over more than two orders of magnitude [84, 85]. This observation is strongly supported by frequent lack of correspondence between TF binding events detected with ChIP-seq versus DGF [85]. Also, TFs with transient DNA binding times in living cells leave minimal to no detectable footprints at their sites of recognition, making the quality of footprinting highly factor-dependent [84, 85]. Collectively, these findings challenge previous DGF research on TF footprinting and its applicability as a reliable recognition assay of complex factor-chromatin interactions in a dynamic timescale.
