Mapping 3D genome organization at nucleosome-scale with Micro-C and Region Capture Micro-C (RCMC).

Micro-C and Region Capture Micro-C (RCMC) are developments of chromosome conformation capture (3C) genomics methods used to identify regions of the genome in close 3D proximity at nucleosome resolution. Micro-C uses in situ MNase digestion of crosslinked chromatin, biotin labeling, proximity ligation and biotin pulldown to recover ligated nucleosomal DNA fragments, with RCMC including a subsequent capture step to enrich for specific regions of the genome. This presents an advantage over previous approaches, such as Hi-C, which rely on restriction enzymes: by digesting the genome without requiring sequence motifs and performing size selection for ligated mononucleosomes, Micro-C can achieve nucleosome-scale resolution of 3D genome organization. The addition of tiled capture of a region allows for deeper sequencing of a location of interest or for experiments with many conditions to be performed while avoiding costly sequencing of the entire genome. Micro-C enables the identification of all known 3D genome features, including interactions between enhancers and promoters that have been missed by previous methods. RCMC can achieve even greater depth and the identification of even more finely resolved interactions. The Micro-C protocol can be completed in 4 d, with an extra 2 d to complete capture, and can be performed by a competent wet laboratory scientist with experience in molecular biology.