ChIP-exo is a specialized version of ChIP used to very specifically map protein of interest (POI) binding sites in the genome. ChIP-exo adds an additional DNA digestion step to ChIP-seq. When the antibody is bound to the POI-chromatin complex, a 3’ exonuclease is used to digest the DNA protruding from the POI binding site. This reduces the resolution from hundreds of nucleotides to as little as one (Rhee and Pugh, 2012). The 5’ strands remain, and are used to detect the region by microarray, PCR, or most commonly sequencing.
The major advantage of this technology is clearly its increase in resolution, but it also reduces noise by digesting contaminating DNA, therefore less sequencing depth is needed (Rhee and Pugh, 2012). Due to this sensitivity, ChIP-exo has been used to map novel transcriptional initiation sites and other discovery-based studies (Venters and Pugh, 2013). The only real disadvantage is that any complex three dimensional interactions of the protein are lost. For example, if a transcription factor simultaneously binds two genomic regions, this will be read as two distinct binding events, not one.
Recently, ChIP-exo has been adapted to use the common Illumina sequencing platform. This adaptation acutally outperformed ChIP-seq on Illumina in all metrics, and is specially designed for efficient human studies (Serandour et al., 2013).
ChIP-exo Additional Reading
This paper is from the group that developed ChIP-exo and describes the workflow of the process in detail. The authors also apply the technology to characterize previously unknown transcription factor binding sties.
- Rhee, H.S., and Pugh, B.F. (2012). ChIP-exo method for identifying genomic location of DNA-binding proteins with near-single-nucleotide accuracy. Curr. Protoc. Mol. Biol. Chapter 21, Unit 21.24.
- Serandour, A.A., Brown, G.D., Cohen, J.D., and Carroll, J.S. (2013). Development of an Illumina-based ChIP-exonuclease method provides insight into FoxA1-DNA binding properties. Genome Biol. 14, R147.
- Venters, B.J., and Pugh, B.F. (2013). Genomic organization of human transcription initiation complexes. Nature 502, 53-58.