Center for Precision Medicine
Advancing research, education, and clinical practice in precision medicine

Chromatin Profiling Analysis @ CRS

Workflow

3 Figures: #1 Primary anitbody binds specifically to the target chromatin-binding protein, #2 a scondary antibody is added to enhance binding of the Tn% transposase, which is fused to Protein A to bind the antibodies at the target site. #3 The Protein A-Tn5 transposase is pre-loaded iwth sequencing adapters that will be integrated at the ends of the cut DNA fragments. These fragments can tehn be amplified by PCR to prepare DNA libraries.
Image from http://www.abclonal.com/

Cleavage Under Targets & Tagmentation (CUT&Tag)

A targeted chromatin profiling technique that maps protein-DNA interactions, including histone modifications and transcription factor binding, at high resolution. An antibody binds a chromatin-associated protein of interest, and a tethered transposase inserts sequencing adapters at nearby DNA sites, allowing precise mapping of the protein’s genomic locations. CUT&Tag is highly sensitive and works well with low-input or rare samples, providing a more efficient alternative to traditional ChIP-Seq.

This workflow is from experiences in the Jessica Lang Lab at CPM, based on a protocol modified from the whole-cell CUT&Tag protocol described by Stephen Henikoff and colleagues (https://dx.doi.org/10.17504/protocols.io.x54v9mkmzg3e/v5).

Potential Applications

  • Profiling histone post-translational modifications
  • Mapping transcription factor binding sites
  • Studying chromatin accessibility at specific regulatory regions
  • Epigenetic landscape characterization in low-input or single-cell samples
  • Investigating dynamic chromatin changes during development or disease

 

Workflow

Assay of Transposase Accessible Chromatin (ATAC-Seq)

Measures genome-wide chromatin accessibility, providing insights into regulatory regions such as promoters and enhancers. A hyperactive transposase inserts sequencing adapters into open chromatin regions, allowing direct sequencing of accessible DNA. This technique requires minimal input material and enables rapid profiling of chromatin states, making it a powerful tool for studying transcriptional regulation and epigenetic dynamics.

This workflow is from experiences in the Jessica Lang Lab at CPM, based on a modified protocol from Corces et al. Nature Methods 2017 (https://www.nature.com/articles/nmeth.4396).

 

Potential ApplicationsA chart in 3 parts from Nature Protocols., 2022 Part 1 shows Nucleosome ATAC-seq fragments with isolate nuclei, exposed to Tn5 transposase, isloated transposed fragments and amplify and sequence and identify accessible regions with ATAC seq peak. Chart b shows transposition into native chromatin, post transposition DNA fragment, Tn5-induced nick, initial extension at 72 deg C amplification and addition of barcodes and adapter components, purification of ATAC-seq library sites of chromatin accessibility defined by Tn5 insertion, wiht each part showing different colored bars of the proportion in each image. Part C shows ATAC-seq signal by cell type X vs Y with X having a peak at both enhance and promoter and a larger Gene A expression than type Y with GATAA of the DNA sequence modified for TF b

  • Mapping open chromatin regions genome-wide
  • Identifying active promoters, enhancers, and regulatory elements
  • Studying transcriptional regulation in different cell types or conditions
  • Profiling chromatin accessibility in rare or low-input samples
  • Integrating with other epigenomic assays to link accessibility with histone modifications or transcription factor binding

 

 

 

Schematic of the ATAC-seq transposition reaction and library preparation.
Corces, et al. Nature Protocols 2022
https://www.nature.com/articles/s41596-022-00692-9

Epigenomics Contacts

Jessica Lang, Ph.D

Credentials: Department of Pathology and Laboratory Medicine

Position title: Assistant Professor

Email: jessica.lang@wisc.edu

Address:
CRS Faculty Affiliate

Kristin Adams

Credentials: Lang Lab

Position title: Researcher I and Laboratory Manager

Email: researchservice@precisionmedicine.wisc.edu

Address:
CRS Scientific Lead

Price/Sample

CUT&Tag Member Non-member
Library prep, QC, P1 100c flow cell $774 $1,509
ATAC-seq: basic chromatin analysis Member Non-member
Library prep, QC, P2 100c flow cell  $1,185 $1,920
ATAC-seq: transcription factor foot printing Member Non-member
Library prep, QC, P4 100c flow cell $2,762 $3,497
Member  Non-member
DNA Isolation $16 $16