In this episode of the Epigenetics Podcast, we caught up with Steven Henikoff from the Fred Hutchinson Cancer Research Center in Seattle to talk about his work on Chromatin Profiling: From ChIP to CUT&RUN, CUT&Tag and CUTAC.
In the last few years Steven Henikoff has been developing methods which profile the chromatin landscape by using enzyme tethering. The quest first started with ChEC-Seq, which improved on Uli Laemmli's method of Chromatin endogenous cleavage (ChEC) but used sequencing as a read-out rather than southern blotting. Next, Cleavage Under Targets & Release Using Nuclease (CUT&RUN) was developed by making a fusion protein of Protein A and micrococcal nuclease (MNase), making it possible to achieve antibody-targeted cleavage of chromatin fragments. And finally, Cleavage Under Targets & Tagmenation (CUT&Tag) was developed by using Transposase Tn5 instead of MNase, which adds sequencing adapters and fragments chromatin at the same time, streamlining the protocol even further.
In this episode we discuss how working on centromeres set the stage for Steven Henikoff’s subsequent work, how he developed CUT&RUN and CUT&Tag, what the advantages and disadvantages of those methods are and how he developed all those experiments at home in his garage.
 
References

Takehito Furuyama, Steven Henikoff (2009) Centromeric nucleosomes induce positive DNA supercoils (Cell) DOI: 10.1016/j.cell.2009.04.049


Kristina Krassovsky, Jorja G. Henikoff, Steven Henikoff (2012) Tripartite organization of centromeric chromatin in budding yeast (Proceedings of the National Academy of Sciences of the United States of America) DOI: 10.1073/pnas.1118898109



Jorja G. Henikoff, Jitendra Thakur, … Steven Henikoff (2015) A unique chromatin complex occupies young α-satellite arrays of human centromeres (Science Advances) DOI: 10.1126/sciadv.1400234



Epigenomics Methods: ChEC-Seq, CUT&RUN, AutoCUT&RUN, Improved CUT&RUN, CUT&Tag, Automated CUT&Tag, CUTAC, scCUT&Tag.
 


Related Episodes

The Role of Non-Histone Proteins in Chromosome Structure and Function During Mitosis (Bill Earnshaw)


Hi-C and Three-Dimensional Genome Sequencing (Erez Lieberman Aiden)


In Vivo Nucleosome Structure and Dynamics (Srinivas Ramachandran)
 


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