Figures
X-ChIP, native ChIP, and CUT&RUN.
(A) In X-ChIP, cells are first crosslinked (red crosses) with formaldehyde to freeze the interactions between the DNA (black line) and a chromatin-binding protein of interest (CP; blue). Sonication fragments the chromatin and makes it soluble. Antibodies are used to recognize the protein–DNA fragments, which are then ‘pulled’ out of the solution using antibody-binding beads, in a process called immunoprecipitation. The histones are shown in yellow. (B) In native ChIP, chromatin is fragmented and solubilized by treating cells with an enzyme called micrococcal nuclease (MNase; small brown shapes). The natural affinity of the protein for its DNA target keep them together during the immunoprecipitation process. (C) In CUT&RUN, antibodies direct the activity of the MNase enzyme to ensure that chromatin cleavage happens close to the protein of interest. A protein called protein A (brown ovals) binds the MNase enzyme to the antibody. The resulting small DNA fragments can be isolated as they diffuse out of the nuclei.
