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| Status |
Public on Apr 01, 2020 |
| Title |
TC-797_washout_replicate2 (HiC) |
| Sample type |
SRA |
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| Source name |
TC-797 cultured cells
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| Organism |
Homo sapiens |
| Characteristics |
cell type: NUT carcinoma
cell line: TC-797
spike-in: Drosophila melanogaster Kc167 cells
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| Treatment protocol |
Tetracycline hydrochloride (Sigma-Aldrich #T7660) was solubilized in water. 293TRex-Flag-BRD4-NUT-HA were induced to express BRD4-NUT with 1 µg/mL tetracycline for 8 hours. MZ1 (Cayman Chemical #21622) was solubilized in DMSO. TC-797 cells were treated with 100 nM MZ1 (0.01% final DMSO) for 4 hours. For washout experiments, TC-797 cells were treated with 100 nM MZ1 (0.01% final DMSO) for 4 hours, washed three times with PBS, and then returned to normal culture media (as described above) for 24 hours.
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| Growth protocol |
293TRex-Flag-BRD4-NUT-HA cells were a kind gift from Christopher A. French (Brigham and Women’s Hospital, Boston, MA). Cells were grown in DMEM with 4.5 g/L glucose and sodium pyruvate without L-glutamine (Corning #15013CV) + 10% Fetal Bovine Serum (Atlanta Biologicals #S10350) + 1% GlutaMAX (Gibco #35050061) + 1% Penicillin-Streptomycin (Gibco #15140122) + 15 µg/mL blasticidin (Gibco #A1113903) + 100 µg/mL hygromycin (Corning #30240CR) at 37° C with 5% CO2. TC-797 cells were a kind gift from Jeffrey A. Toretsky (Georgetown University, Washington, DC). Cells were grown in DMEM with 4.5 g/L glucose and sodium pyruvate without L-glutamine (Corning #15013CV) + 10% Fetal Bovine Serum (Atlanta Biologicals #S10350) + 1% GlutaMAX (Gibco #35050061) + 1% Penicillin-Streptomycin (Gibco #15140122) at 37° C with 5% CO2. Kc167 cells were obtained from the Drosophila Genomics Resource Center (#1; Bloomington, IN). Cells were grown in CCM3 Media (HyClone #SH30062.02) at 25° C.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Human cells were mixed with Drosophila cells at a 2:1 ratio and then cells were cross-linked with 1% EM-grade paraformaldehyde. Intact nuclei were crudely isolated using detergent lysis. DNA was digested with MboI (prior to digestion an aliquot was saved as an input control), DNA ends filled in with biotin-14-dATP, and free DNA ends ligated together in situ. DNA-protein cross-links were reversed and the DNA then purified.
DNA sheared to a size of approximately 500 bp. DNA was end-repaired and adaptor-ligated by following “NEBNext End Prep” and “Adaptor Ligation” in the NEBNext Ultra II DNA Library Prep Kit for Illumina (New England Biolabs). The biotinylated DNA was pulled down with streptavidin beads and PCR amplified by following "PCR Amplification" in the NEBNext Ultra II DNA Library Prep Kit for Illumina (New England Biolabs).
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| Library strategy |
Hi-C |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina NovaSeq 6000 |
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| Data processing |
Reads were aligned to concatenated hg38/Genome Reference Consortium Human Reference 38 Patch 15 (GRCh38.p15) and dm6/BDGP Release 6 + ISO1 MT assemblies of the human and Drosophila melanogaster genomes, respectively, using BWA-MEM v0.7.15-r1140.
Reads were then assigned to MboI restriction fragments, duplicates removed, reads with a MAPQ ≥ 30 were retained, and intra restriction fragment reads removed.
The genome was divided into equally spaced, non-overlapping bins and the number of reads counted in each pair of bins gave the number of contacts, generating a Hi-C contact map.
Hi-C contact maps were normalized by matrix balancing using the KR normalization algorithm (Durand et al., 2016; Knight and Ruiz, 2012; Rao et al., 2014) using Juicer Tools v1.8.9.
Biological replicates were combined after filtering and the combined contact map KR normalized. All subsequent analysis was performed on the combined, KR normalized contact map.
TADs were identified using the previously described Arrowhead algorithm (Durand et al., 2016; Rao et al., 2014) using Juicer Tools v1.8.9.
Loops were identified using the previously described HiCCUPs algorithm (Durand et al., 2016; Rao et al., 2014) using Juicer Tools v1.8.9.
All of the above steps were performed using the previously described Juicer pipeline (Durand et al., 2016; Rao et al., 2014) and Juicebox visualization interface (Durand et al., 2016).
Megadomain-megadomain interactions were identified using a model that accounted for enrichment relative to local background based on Poisson statistics.
Genome_build: GRCh38.p15 concatenated with dm6/BDGP Release 6 + ISO1 MT
Supplementary_files_format_and_content: *.hic file of Hi-C contacts in compressed format. Readable by open-source Juicebox software (http://aidenlab.org/juicebox/).
Supplementary_files_format_and_content: *_TADs.bedpe file of TAD coordinates in Juicebox format.
Supplementary_files_format_and_content: *_loops.bedpe file of loop coordinates in Juicebox format.
Supplementary_files_format_and_content: *_mega-mega.bedpe file of megadomain-megadomain interactions in Juicebox format.
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| Submission date |
Jun 21, 2019 |
| Last update date |
Apr 02, 2020 |
| Contact name |
Kyle Eagen |
| E-mail(s) |
kyle.eagen@bcm.edu
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| Organization name |
Baylor College of Medicine
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| Department |
Department of Molecular and Cellular Biology
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| Lab |
Eagen Lab
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| Street address |
1 Baylor Plaza
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| City |
Houston |
| State/province |
TX |
| ZIP/Postal code |
77030 |
| Country |
USA |
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| Platform ID |
GPL24676 |
| Series (2) |
| GSE133163 |
Hi-C to Identify Interactions between Distant Genomic Loci due to BRD4-NUT-mediated Megadomain Formation |
| GSE133165 |
Chromatin Hyperacetylation Impacts Chromosome Folding by Forming a Nuclear Subcompartment |
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| Relations |
| BioSample |
SAMN12108432 |
| SRA |
SRX6183141 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM3901278_tc-797_washout_replicate2.hic |
2.2 Gb |
(ftp)(http) |
HIC |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
| Processed data are available on Series record |
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