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Status |
Public on Apr 29, 2021 |
Title |
WT_Input DNA |
Sample type |
SRA |
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|
Source name |
HAP1 Parental Cells
|
Organism |
Homo sapiens |
Characteristics |
cell type: HAP1 cells derived from male CML cell line KBM7 morphology: Fibroblast-like genotype: Wild-type chip antibody: None
|
Treatment protocol |
Cells were harvested by trypsinization followed by two washes with ice-cold PBS.
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Growth protocol |
Human HAP1 WT and DSETX cells were cultured in IMDM (ThermoFisher) supplemented with 10% FBS (Sigma-Aldrich) and 1% Penicillin/Streptomycin (Sigma-Aldrich). Cells were grown at 37°C and in a humidified atmosphere containing 5% CO2.
|
Extracted molecule |
genomic DNA |
Extraction protocol |
Genomic DNA was extracted and sonicated to ~150-300 bp using a Misonix Sonicator 300 equipped with a microtop. Illumina adaptors were added to the sonicated DNA and immunoprecipitated using a mouse monoclonal antibody against 5-methylcytosine (5meC; Clone 33D3, Active Motif) following the protocol described in the Active Motif MeDIP kit (Active Motif, Catalog No. 55009). Libraries were prepared according to Illumina's instructions. Deep sequencing libraries were generated from the DNA immunoprecipitates and input DNA by the standard consecutive enzymatic steps of end-polishing, dA-addition and adaptor ligation. After a final PCR amplification step using barcoded primers, the resulting DNA libraries were quantified and sequenced.
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|
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Library strategy |
MeDIP-Seq |
Library source |
genomic |
Library selection |
5-methylcytidine antibody |
Instrument model |
Illumina NextSeq 500 |
|
|
Description |
Input DNA sequencing
|
Data processing |
Sequenced reads were aligned to the human hg19 genome assembly using Burrows Wheeler Aligner (BWA, RRID:SCR_010910) with default settings. Only reads that aligned with no more than 2 mismatches and mapped uniquely to the genome were used for subsequent analyses. PCR duplicate reads were removed. The sequence reads represented the 5’-ends of the IP-fragments, hence the reads were extended in silico using proprietary software from Active Motif at their 3’-ends to a length of 150 bp to match the fragment length of the library. To identify the density of fragments, the genome was divided into 32-nt bins and the number of fragments in each bin was determined. The resulting metric (“fragment density”) was used for the peak metrics in further analyses. Genome_build: hg19 Supplementary_files_format_and_content: bigWig files representing normalised coverage, created using the standard Active Motif MeDIP-seq pipeline.
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|
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Submission date |
Jun 30, 2020 |
Last update date |
Apr 29, 2021 |
Contact name |
Steve West |
E-mail(s) |
stephen.west@crick.ac.uk
|
Organization name |
The Francis Crick Institute
|
Street address |
1 Midland Road
|
City |
London |
ZIP/Postal code |
NW1 1AT |
Country |
United Kingdom |
|
|
Platform ID |
GPL18573 |
Series (2) |
GSE143574 |
Integrated Genome and Transcriptome Analyses Reveal the Mechanism of Genome Instability in Ataxia with Oculomotor Apraxia 2 |
GSE153577 |
Integrated Genome and Transcriptome Analyses Reveal the Mechanism of Genome Instability in Ataxia with Oculomotor Apraxia 2 [MeDIP-Seq] |
|
Relations |
BioSample |
SAMN15408008 |
SRA |
SRX8642914 |