Framework for experimental design and analysis for low input/single cell chromatin profiling: CUT&Tag

CUT&Tag recovers up to half of ENCODE ChIP-seq peaks

Want to conduct genome wide profiling of histone marks and transcription binding, but don't have hundreds of thousands of cells needed for ChIP-seq? CUT&Tag may offer a solution, but because it is a relatively new method (since 2019), there lacked a framework for experimental design and analysis. This is where I step in: https://www.biorxiv.org/content/10.1101/2022.03.30.486382v1

Article TitleCUT&Tag recovers up to half of ENCODE ChIP-seq peaks

DOI
Published
03/31/2022
License
Non CC - Author held copyright

Abstract

Techniques for genome-wide epigenetic profiling have been undergoing rapid development toward recovery of high quality data from bulk and single cell samples. DNA-protein interactions have traditionally been profiled via chromatin immunoprecipitation followed by next generation sequencing (ChIP-seq), which has become the gold standard for studying histone modifications or transcription factor binding. Cleavage Under Targets & Tagmentation (CUT&Tag) is a promising new technique, which enables profiling of such interactions _in situ_at high sensitivity and is adaptable to single cell applications. However thorough evaluation and benchmarking against established ChIP-seq datasets are still lacking. Here we comprehensively benchmarked CUT&Tag for H3K27ac and H3K27me3 against published ChIP-seq profiles from ENCODE in K562 cells. Across a total of 30 new and 6 published CUT&Tag datasets we found that no experiment recovers more than 50% of known ENCODE peaks, regardless of the histone mark. We tested peak callers MACS2 and SEACR, identifying optimal peak calling parameters. Balancing both precision and recall of known ENCODE peaks, SEACR without retention of duplicates showed the best performance. We found that reducing PCR cycles during library preparation lowered duplication rates at the expense of ENCODE peak recovery. Despite the moderate ENCODE peak recovery, peaks identified by CUT&Tag represent the strongest ENCODE peaks and show the same functional and biological enrichments as ChIP-seq peaks identified by ENCODE. Our workflow systematically evaluates the merits of methodological adjustments and will facilitate future efforts to apply CUT&Tag in human tissues and single cells.


Login or Signup to leave a comment
Di Hu4 months ago

any questions feel free to reach out to me!

Find your community. Ask questions. Science is better when we troubleshoot together.
Find your community. Ask questions. Science is better when we troubleshoot together.

Have a question?

Contact support@scifind.net or check out our support page.