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| Status |
Public on Nov 13, 2017 |
| Title |
RaeI shEBNA1 HiC |
| Sample type |
SRA |
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| Source name |
RaeI shEBNA1 HiC
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| Organisms |
Homo sapiens; human gammaherpesvirus 4 |
| Characteristics |
cell type: RaeI cells
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| Treatment protocol |
All cells except Akata-Zta were untreated. 5 million cells per experiment were fixed for in situ Hi-C as previously described (Rao et al., 2014). Akata-Zta cells were pretreated with 200 μM acyclovir for 1 hour before reactivation of the lytic cycle with 500 ng/mL doxycycline. After 1 day, cells were magnetically sorted using LNGFR Microbeads and LS columns (Miltenyi Biotech). 5 million cells each of LGNFR+ and LGNFR- were fixed for in situ Hi-C as previously described (Rao et al. 2014)
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| Growth protocol |
EBV-positive Daudi, KemIII, RaeI, and Raji cells were maintained under standard conditions (Phan et al., 2016). K562 (C. B. Lozzio and B. B. Lozzio, 1975), EBV- and KSHV-positive BC-1 (Cesarman et al., 1995) and EBV-positive Namalwa (Klein et al., 1972) cells were maintained in RPMI-1640 with 25 mM HEPES and 2 g/L NaHCO3 supplemented with 10% (v/v) fetal bovine serum (Invitrogen) in 5% CO2 at 37 °C. EBV-positive Akata-Zta cells (Ramasubramanyan et al., 2015) were maintained in RPMI-1640 with 25 mM HEPES and 2 g/L NaHCO3 supplemented with 10% (v/v) Tet System Approved fetal bovine serum (Clontech). The HPV16-positive 20863 (Jeon et al., 1995) and HPV31-positive 9E (De Geest et al., 1993) keratinocyte cell lines were grown in F-medium, 3:1 (v/v) F-12-DMEM, 5% fetal bovine serum, 400 ng/ml hydrocortisone, 5 µg/ml insulin, 8.4 ng/ml cholera toxin, 10 ng/ml epidermal growth factor, 24 µg/ml adenine, 100 U/ml penicillin, and 100 µg/ml streptomycin, in the presence of irradiated 3T3-J2 feeder cells, as described previously (Chapman et al., 2014). Irradiated feeder cells were removed by versene treatment before the cells were harvested.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
In situ Hi-C was performed with 5 million cells per experiment and restriction enzyme MboI as described (Rao et al., 2014) with slight modifications. After end-repair and washes, Dynabeads (Thermo Fisher Scientific) with bound DNA were resuspended in 10 mM Tris, 0.1 mM EDTA pH 8.0 and transferred to new tubes.
Sequencing libraries were created from bound DNA using the Ovation Ultralow Library System V2 kit (NuGEN) with one modification. After adapter ligation, because DNA is still attached to the beads, water instead of SPRI beads was added to the reaction. DNA bound to the beads was purified with a magnet, washed, and the beads were resuspended in 10 mM Tris, 0.1 mM EDTA pH 8.0. After library amplification, SPRI beads were added as directed to purify the amplified DNA.
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| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina HiSeq 4000 |
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| Data processing |
library strategy: HiC-seq
General HiC processing: Paired reads were aligned to combined human/viral reference genomes with HiCUP version 0.5.0 using default parameters (Wingett et al., 2015) to generate a set of interactions. We used the human hg19 sequence merged with the EBV NC_007605.1, KSHV NC_009333.1, HPV16 NC_001526.2, or HPV31 J04353.1 sequence. Only alignments with mapq scores equal to or greater than 30 were retained.
Chromosome-resolution heatmaps of interactions were determined from the HiCUP-filtered interchromosomal Hi-C interactions in sam format. Expected interactions were calculated using the following equation for each chromosome pair, where chrA represents the number of single end interchromosomal reads containing chrA, chrB represents the number of single end interchromosomal reads containing chrB, totalPairs represents the total number of interchromosomal paired end reads, and all represents the total number of interchromosomal single end reads, which is equal to 2 x totalPairs: (((chrA/all)*(chrB/(all-chrA)))+((chrB/all)*(chrA/(all-chrB)))) * total_pairs.
Lamin association analysis: The set of lamin interacting domains from Tig3 cells (Guelen et al., 2008) was downloaded from the UCSC genome browser (Kent et al., 2002). Hi-C interactions identified by HiCUP were processed into a format readable by the HiTC package (Servant, 2015) using 1 Mb bins for the human genome and 1 kb bins for the viral genome. We performed further analyses on these interaction matrices in R (R Core Team, 2016). First a full interaction matrix for all autosomes was constructed. Next, for each bin, the mean of interaction counts with LAD bins was calculated as was the mean of interaction counts with non-LAD bins. Correlations were then performed across all autosomal bins with the LAD mean vector and the non-LAD mean vector. A logistic regression was performed on the LAD and non-LAD correlation values estimate the probability with which each genome region would interact with lamin. Next, we calculated the probability that each viral bin interacted with the lamin by applying the logistic regression model of the autosomal LAD correlations to the viral interaction data.
Genome_build: human hg19 sequence merged with the EBV NC_007605.1, KSHV NC_009333.1, HPV16 NC_001526.2, or HPV31 J04353.1 sequence
Supplementary_files_format_and_content: Chromsome-resolution heatmap generation results in two plain text files *interchrom_raw_counts_label.txt and *obs_exp_log2_label.txt. The *interchrom_raw_counts_label.txt contains a labeled 26 x 26 matrix for the raw counts of interchromosomal interactions between whole chromosomes. The *obs_exp_log2_label.txt file contains the same labeled 26 x 26 matrix, with raw interchromosomal counts normalized as described above.
Supplementary_files_format_and_content: Lamin association analysis results in plain text files ending in *allCounts.txt, which contains three columns, the first two are individual genomic bins and the third column is the number of interactions between those two bins.
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| Submission date |
Apr 24, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
JJ Miranda |
| E-mail(s) |
jmiranda@barnard.edu
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| Organization name |
Barnard College, Columbia University
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| Street address |
3009 Broadway
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| City |
New York |
| ZIP/Postal code |
10027 |
| Country |
USA |
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| Platform ID |
GPL23362 |
| Series (2) |
| GSE98120 |
The Epstein-Barr virus episome maneuvers between nuclear chromatin compartments during reactivation [HiC-seq] |
| GSE98123 |
The Epstein-Barr virus episome maneuvers between nuclear chromatin compartments during reactivation |
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| Relations |
| BioSample |
SAMN06824396 |
| SRA |
SRX2755231 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM2587508_rae_shEBNA1_hg19chrebv_h1000000_v1000_allCounts.txt.gz |
17.4 Mb |
(ftp)(http) |
TXT |
| GSM2587508_rae_shEBNA1_interchrom_raw_counts_label.txt.gz |
1.7 Kb |
(ftp)(http) |
TXT |
| GSM2587508_rae_shEBNA1_obs_exp_log2_label.txt.gz |
1.8 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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