ATACSeq

ATAC-seq, short for Assay for Transposase-Accessible Chromatin using sequencing, is a genome-wide method for mapping open chromatin regions. It identifies regulatory elements such as promoters and enhancers by exploiting the activity of a hyperactive Tn5 transposase that simultaneously cuts accessible DNA and inserts sequencing adapters, a process known as tagmentation.

In bulk ATAC-seq, input material can be from thousands to millions of cells, while single-cell ATAC-seq (scATAC-seq) inventories chromatin accessibility at the level of individual cells, enabling the study of heterogeneous tissues and rare cell types. The protocol generally yields fragment libraries that reflect chromatin structure: short fragments (roughly under 100 base pairs) indicate nucleosome-free regions, whereas fragments around 200 base pairs or longer correspond to mono- and di-nucleosome protection patterns. Sequencing these libraries allows the identification of accessible regions and regulatory elements across the genome.

Data analysis involves aligning reads to a reference genome, calling accessible regions or peaks, and assessing quality metrics such as the fraction of reads in peaks (FRiP) and transcription start site (TSS) enrichment. Additional analyses include nucleosome occupancy patterns and transcription factor footprinting using tools designed for ATAC-seq data (and with methods to correct for Tn5 bias). In scATAC-seq, computational approaches cluster cells by chromatin accessibility profiles.

Applications span developmental biology, cancer and immune research, and epigenomic mapping of regulatory landscapes. Limitations include sequence bias from Tn5, GC-content effects, and the indirect nature of accessibility measurements, which do not directly measure DNA methylation.