EXPERIMENT_TITLE: GSM4306485: Cremer-2019-HIC005; Homo sapiens; Hi-C
EXPERIMENT_DESIGN_LIBRARY_DESCRIPTOR_LIBRARY_CONSTRUCTION_PROTOC: Cells were crosslinked and then lysed with nuclei permeabilized but still intact. DNA was then restricted and the overhangs filled in incorporating a biotinylated base. Free ends were then ligated together in situ. Crosslinks were reversed, the DNA was sheared to 300-500bp and then biotinylated ligation junctions were recovered with streptavidin beads. standard Illumina library construction protocol, Briefly, DNA was end-repaired using a combination of T4 DNA polymerase, E. coli DNA Pol I large fragment (Klenow polymerase) and T4 polynucleotide kinase. The blunt, phosphorylated ends were treated with Klenow fragment (32 to 52 exo minus) and dATP to yield a protruding 3- 'A' base for ligation of Illumina's adapters which have a single 'T' base overhang at the 3' end. After adapter ligation DNA was PCR amplified with Illumina primers for 8-12 cycles and library fragments of 400-600 bp (insert plus adaptor and PCR primer sequences) were purified using SPRI beads. The purified DNA was captured on an Illumina flow cell for cluster generation. Libraries were sequenced on the NextSeq 500 following the manufacturer's protocols
STUDY_DESCRIPTOR_STUDY_ABSTRACT: Nuclear functions are essentially linked to nuclear compartmentalization. This study demonstrates that cohesin rings as anchors of chromatin loops are dispensable to rebuild a functional nuclear compartmentalization in cohesin depleted cells after passing through mitosis and formation of one daughter cell with a multilobulated nucleus (MLN). Super-resolved microscopy reveals co-aligned active and inactive nuclear compartments (ANC/INC) in these postmitotic nuclei, likely corresponding to A and B compartments, detected with Hi-C. MLN carry chromosome territories, built from chromatin domain clusters, pervaded by a system of interchromatin channels harboring splicing speckles. Channels are lined by transcriptionally competent chromatin, whereas repressed chromatin with higher compaction locates in the interior of chromatin clusters. MLN pass through S-phase with typical early and mid-to-late replication patterns. Sites of DNA synthesis become physically larger, consistent with a model where cohesin dependent loop extrusion tends to compact intervals of replicating chromatin, whereas their genomic boundaries are associated with compartmentalization. Overall design: Microscopy, Hi-C, and Repli-Seq in HCT-116 cmvOsTir1 RAD21-mAC cells
SAMPLE_TITLE: Cremer-2019-HIC005
SAMPLE_SAMPLE_ATTRIBUTES_SAMPLE_ATTRIBUTE: [{"TAG": "source_name", "VALUE": "HCT-116 cmvOsTIR1 RAD21-mAC"}, {"TAG": "cell line background", "VALUE": "HCT-116"}, {"TAG": "genotype/variation", "VALUE": "cmvOsTIR1, RAD21-mAC"}, {"TAG": "treatment", "VALUE": "500uM IAA, 6hrs"}]
Pool_Member_@attributes_sample_title: Cremer-2019-HIC005
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