Targeted deaminase-accessible chromatin sequencing (TDAC-seq) measures chromatin accessibility across long chromatin fibers at targeted loci using double-stranded DNA cytidine deaminases. When combined with pooled CRISPR mutational screening, TDAC-seq enables the high-throughput detection of changes in chromatin accessibility following CRISPR perturbations, allowing fine mapping of sequence–function relationships within endogenous cis-regulatory elements.