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| Status |
Public on Jul 18, 2016 |
| Title |
NHEK_bless_seq_rep2 |
| Sample type |
SRA |
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| Source name |
NHEK_bless_seq
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| Organism |
Homo sapiens |
| Characteristics |
cell line: NHEK
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| Growth protocol |
HeLa cells were cultured in DMEM (Sigma, D6429) supplemented with 10% FBS (Gibco, U120D). U2OS AID-DIvA cells were cultured in DMEM supplemented with 10% FBS and 800μg/mL G418 (Gibco, 10131). To induce nuclear localisation of AsiSI, AID-DIvA cells were treated with 300 nM 4OHT (Sigma, H7904) for 4 hours. Primary keratinocytes (Gibco, A13401) were cultured in EpiLife Medium (Gibco, M-EPI-500-CA) supplemented with human keratinocyte growth supplement (Gibco, S-001-5) and were detached using accutase (Sigma, A6964).
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
The BLESS extraction protocol was implemented as published in Crosetto et al., 2013 (http://breakome.utmb.edu).
The TruSeq Nano DNA Library Prep Kit (Illumina, FC-121-4001) was used to generate libraries for sequencing (size selection for 550bps). The BREAk-seq protocol was implemented as described in the Methods sectin of the associated paper (Lensing et al., 2016).
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| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina NextSeq 500 |
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| Description |
BLESS extracted DNA
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| Data processing |
Library strategy: Break-Seq
fastq files containing sequencing reads were pre-processed to remove the Illumina adapters and to trim for low quality tails by using trim_galore (http://www.bioinformatics.babraham.ac.uk/projects/trim_galore/)
reads were aligned to the human reference genome (hg19) using the bwa mem aligner (http://sourceforge.net/projects/bio-bwa/files/) and cleaned for low quality alignments (mapQ < 10) using samtools (http://samtools.sourceforge.net)
Duplicates were identified using picard tools (https://github.com/broadinstitute/picard/) and removed. Only reads from read 1, which are proximal to the break site, are then retained for the downstream analysis and shown in the bedGraph coverage plots.
Specific for the BLESS libraries: during adapter trimming, the BLESS linker sequences are trimmed (TCGAGGTAGTA and TCGAGACGACG for proximal and distal linkers, respectively), and only read pairs showing both linkers are retained. Trimmed fastq files then undergo the same processing pipeline as for BREAk-seq libraries. At the end, only sequencing reads that originally contained the proximal linker (they could be from either read 1 or read 2) are retained for subsequent analysis and shown in the coverage plots
Genome_build: hg19
Supplementary_files_format_and_content: bedGraph, TDF
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| Submission date |
Feb 22, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Giovanni Marsico |
| E-mail(s) |
persego@gmail.com
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| Organization name |
CRUK Cambridge Institute
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| Street address |
Robinson Way
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| City |
Cambridge |
| ZIP/Postal code |
CB2 0RE |
| Country |
United Kingdom |
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| Platform ID |
GPL18573 |
| Series (1) |
| GSE78172 |
DSBCapture: in situ capture and direct sequencing of dsDNA breaks |
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| Relations |
| BioSample |
SAMN04506973 |
| SRA |
SRX1596478 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM2068758_BLESS_n2.bedGraph.gz |
1.2 Gb |
(ftp)(http) |
BEDGRAPH |
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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