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We have developed Differential Specificity and Energy Landscape (DiSEL) analysis to comprehensively compare DNA–protein interactomes (DPIs) obtained by high-throughput experimental platforms and cutting edge computational methods. While high-affinity DNA binding sites are identified by most methods, DiSEL uncovered nuanced sequence preferences displayed by homologous transcription factors. Pairwise analysis of 726 DPIs uncovered homolog-specific differences at moderate- to low-affinity bindingdoi:10.1073/pnas.1811431115 fatcat:gubbnu5bybhpxdfcswcxerexz4