title: GSM7903381: Bone marrow-derived macrophage-BMDMCTR_Rep1; Mus musculus; Hi-C
library_name: GSM7903381
library_construction_protocol: Hi-C was performed by following the reported HiChIP protocol and omitting the protein immunoprecipitation step42. Briefly, 106 bone marrow-derived macrophages were crosslinked with 1% formaldehyde for 10-minutes. Formaldehyde was quenched by the addition of glycine. Nuclei were isolated in lysis buffer (1% Triton x-100, 0.1% SDS, 150 mM NaCl, 1mM EDTA, and 20 mM Tris, pH 8.0) by rotating the cells for 1-hour at 4C. Nuclear pellet was resuspended in 0.5% SDS solution and incubated at 62C for 10 minutes. SDS was quenched by the addition of 10% Triton-X. Chromatin was digested with 8U of MboI enzyme overnight at 37C. Biotin fill-in was performed for 1-hour at 37C, followed by proximity ligation for 6-hours at room temperature. After proximity ligation, nuclei were subjected to Proteinase K digestion and decrosslinking overnight at 68C. DNA was column purified (Minelute, Qiagen), and sonicated to a size distribution of 200-500bp. DNA was subjected to streptavidin bead binding to pull-down biotinylated DNA that represents ligation junctions. Tn5 was used to create libraries by on bead tagmentation with sequencing adapters, followed by PCR. Libraries were quantified by Qubit and size distribution was assessed with Bioanalyzer.