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| Status |
Public on May 24, 2018 |
| Title |
5F UNGKO PD 1 |
| Sample type |
SRA |
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| Source name |
HEK293T
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| Organism |
Homo sapiens |
| Characteristics |
cell line: HEK293T
atcc number: CRL-3216
enrichment: dU enrichment
genotype: UNG-/-
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| Treatment protocol |
"5F" samples mean cells was treated with 5 μM 5-fluorodeoxyuridine for 48h.
None
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| Growth protocol |
cells was maintained in DMEM with 10% FBS and penicillin/streptomycin.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
DNA was extracted by Universal Genomic DNA Kit (CWbiotech, CW2298).
Genomic DNA was digested to 100-500 bp and purified with Agencourt AMPure XP beads (BECKMAN COULTER) following Spin-6 column (Bio-rad) purification. 2 μg DNA fragments and 15 pg spike_in sequences were used for end repair with NEBNext® End Repair Module (NEB, E6050) and 1 μl E.coli ligase was added to repair nicks in DNA. DNA was then purified with 1.8×AMPure XP beads. dA was added to the 3’ end of double-stranded DNA by NEBNext® dA-Tailing Module (NEB, E6053) and purified with 1.8×AMPure XP beads.
Damages that may interfere the following labeling step were repaired. DNA was purified and subjected to in vitro BER labeling. For samples named "biotin-dCTP", biotin-dCTP was utilized in the in vitro BER labeling;other samples utilized biotin-dUTP. After purification, fragments labeled with biotin were enriched by streptavidin C1 beads (Invitrogen) following the manufacturer’s guidelines.
Y adaptor was ligated to double-stranded DNA on streptavidin C1 beads so that free adaptor can be removed by beads washing. DNA was then eluted from beads by 95 °C 3 min using deionized water. Eluted DNA was subjected to PCR amplification and sequencing.
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| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
HiSeq X Ten |
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| Description |
5F_UNGKOPD.uniq.peaks.bed
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| Data processing |
150bp pairend Raw reads were firstly sent for adaptor and quality trimming using trim_galore, and reads shorter than 25 nt after trimming were excluded. For spike-in DNA sequences calculation, bowtie2 was used to reads mapping to corresponding sequences. For genome-wide dU sites identification, processed reads mapped to spike- in sequence were discarded, and then mapped to human reference genome (hg38) using bowtie2. By default, bowtie2 searches for multiple alignments bowtie2 search and only reports a random one in the best matches; for repeat sequences, bowtie reports the best matched locus or random one from the best-matched loci. After alignment, peaks were called using model-based analysis of ChIP-Seq (MACS2) using nonredundant reads. Peaks overlap with that in the control sample were removed. Genomic annotations were performed using Homer software. Gene annotations(RefSeq) were download from UCSC.
Genome_build: hg38
Supplementary_files_format_and_content: bed
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| Submission date |
Sep 01, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
zhike lu |
| E-mail(s) |
[email protected]
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| Organization name |
University of Chicago
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| Department |
department of chemistry
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| Street address |
5801 South Ellis Avenue
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| City |
Chicago |
| State/province |
IL |
| ZIP/Postal code |
60637 |
| Country |
USA |
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| Platform ID |
GPL20795 |
| Series (1) |
| GSE99011 |
Genome-wide mapping reveals that deoxyuridine is enriched in the human centromeric DNA |
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| Relations |
| BioSample |
SAMN07594243 |
| SRA |
SRX3153912 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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