EXPERIMENT_TITLE: GSM3956923: HiC-K562-Cell-R1; Homo sapiens; Hi-C
EXPERIMENT_DESIGN_LIBRARY_DESCRIPTOR_LIBRARY_CONSTRUCTION_PROTOC: Hi-C was performed as described (Hi-C 2.0 Belaghzal et al., 2017) . For intact cells: 5 million K562 cells or nuclei were washed twice with 15 mL of HBSS and pelleted at 300 g for 10 min, then resuspended in 2.5 mL of HBSS. The sample was transferred to 20.625 mL crosslinking mix [625 μL of 37% formaldehyde + 20 mL of HBSS]. Samples were incubated at RT for 10 min on a rocking platform. Next, to stop cross-linking 1.25 mL of 2.5 M glycine was added to each sample and the mix was incubated at RT for 5 min on a rocking platform. To pellet the crosslinked cells the sample was centrifuged at 1,000 g for 10 min at 4°C. The supernatant was discarded and the pellet was washed twice with HBSS before going to the next step or storing samples at -80°C. Lysis For Hi-C with intact cells: the 5 million cross-linked cells were lysed by adding 1 mL cold lysis buffer [10 mM Tris-HCl (pH=8.0), 10 mM NaCl, 0.2% Igepal CA-630 (NP40)] and 10 µL of 100X Protease inhibitors. The sample was incubated on ice for 15 min to let the cells swell. The cells were lysed on ice using a dounce homogenizer with pestle A by moving the pest Hi-C 2.0: We prepare the library for sequencing as described in Hi-C 2.17
STUDY_DESCRIPTOR_STUDY_ABSTRACT: Chromosomes are folded so that active and inactive chromatin domains are spatially segregated to form a variety of sub-nuclear neighborhoods. Compartmentalization is thought to occur through polymer phase/microphase separation mediated by interactions between loci of similar type. The nature and dynamics of these interactions are not known. We developed liquid chromatin Hi-C to map the stability of associations between loci genome-wide. Before fixation and Hi-C, chromosomes are fragmented removing the strong polymeric constraint to enable detection of intrinsic locus-locus interaction stabilities. We find that chromosome compartmentalization is dependent on the length of chromatin fragments. Compartmentalization is stable when fragments are over 10-25 kb. Fragmenting chromatin into pieces smaller than 6 kb leads to gradual loss of spatial genome organization. In addition to fragmentation level dissolution kinetics of chromatin interactions vary for different chromatin domains. Lamin associated domains are most stable, and speckle-associated loci are more dynamic. The polycomb-enriched B1 sub-compartment also displays highly unstable interactions. Cohesin-mediated loops dissolve after fragmentation, possibly because cohesin rings slide off nearby DNA ends. Liquid chromatin Hi-C provides a genome-wide view of chromosome interaction dynamics, revealing a range of conformational stabilities at different sub-nuclear structures. Overall design: Two replicate timecourses of liquid chromatin Hi-C, Hi-C 2.0 and 5C in cells and mock nuclei, DpnII-seq timecourse
SAMPLE_TITLE: HiC-K562-Cell-R1
SAMPLE_SAMPLE_ATTRIBUTES_SAMPLE_ATTRIBUTE: [{"TAG": "source_name", "VALUE": "HiC-K562-Cell"}, {"TAG": "cell line", "VALUE": "K562"}, {"TAG": "cell type", "VALUE": "The continuous cell line K-562 was established by Lozzio and Lozzio from the pleural effusion of a 53-year-old female with chronic myelogenous leukemia in terminal blast crises."}, {"TAG": "protocol", "VALUE": "Hi-C 2.0"}, {"TAG": "enzyme", "VALUE": "HindIII"}]
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