Introduction of Some New Technologies (2)

November 16, 2018

ATAC-seq, or Assay for Transposase-Accessible Chromatin coupled with NGS, is a technique to locate accessible chromatin regions. The method is a fast and sensitive alternative to DNase-seq (DNase I hypersensitive sites sequencing) or MNase-seq (micrococcal nuclease sensitive sites sequencing).By using this service, we can detect genome-wide profiles of open and accessible regions of chromatin that are indicative of active regulatory regions. The eukaryotic genome is highly packaged to fit into the very limited nuclear space. As a result, access to genomic information is tightly regulated based on cellular state. What regions of the genome are accessible reveals a great deal about the state of the cell.

The key part of the ATAC-seq procedure is the action of the transposase Tn5 on the genomic DNA of the sample. Transposases are enzymes catalyzing the movement of transposons to other parts of the genome. While naturally occurring transposases have a low level of activity, ATAC-seq employs a mutated hyperactive transposase. The high activity allows for highly efficient cutting of exposed DNA and simultaneous ligation of specific sequences, called adapters. Adapter-ligated DNA fragments are then isolated, amplified by PCR and used for next generation sequencing. The experimental pipeline of ATAC-seq is demonstrated in Figure 1.

Figure 1. Schematic workflow of ATAC-seq process.

Sequencing Strategy and Recommended Depth:

• Illumina HiSeq SE50 or SE75. For some applications such as nucleosome mapping, paired end sequencing is preferred.
• ≥50 M clean reads

Sample Requirements:

• Sample type:
  Live cells, not genomic DNA
  Human and animal tissues
  Primary cells (including T and B cells)
  FACS sorted cells
  Most rare cell populations
• cell number : ≥ 50,000 cells
• Healthy cells in a homogeneous single-cell suspension work the best.

1. Gain mechanistic insight into gene regulation, cellular response to treatment or disease
2. Identify which transcription factors are driving cell fate, disease, or response
3. Primary tissues or cells such as pancreatic Beta cells
4. Limited patient samples
5. Stratify patients or sample groups based on open chromatin signatures
6. Low requirements on the amount of the biological sample, and the whole protocol requires 3 hours in total.