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URL: https://dx.doi.org/10.17504/protocols.io.bnx5mfq6

Authors: Kami Ahmad, Steven Henikoff

Summary: This is a modification of the Benchtop CUT&Tag method (dx.doi.org/10.17504/protocols.io.bcuhiwt6) for epigenomic profiling of histone modifications and chromatin proteins in dissected Drosophila tissues. Intact unfixed tissues are permeabilized and incubated under conditions where a factor-specific antibody can infiltrate and bind a chromatin protein in situ. This antibody is then decorated with a secondary antibody, and this is used to tether a protein AG-Tn5 fusion protein loaded with sequencing adapters. Upon transposome activation by adding Mg2+ ions, DNA is tagmented around the chromatin binding site. Here, dissected imaginal discs are coated with magnetic beads for handling, and about 5 imaginal discs provides high-quality data for histone modifications or for chromatin factor for which high-quality antibodies are available. The protocol is scaled for 8 samples. Performing CUT&Tag takes 2 days from larvae to sequencing-ready libraries.

Affiliations: Fred Hutchinson Cancer Research Center, Fred Hutchinson Cancer Research Center

Version: 1

Publication Date: 2020

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