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| Status |
Public on Jan 31, 2017 |
| Title |
HTGTS_KO-DMSO rep3 |
| Sample type |
SRA |
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| Source name |
CSR activated B cells
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| Organism |
Mus musculus |
| Characteristics |
strain: 129
tissue: spleen
age: 8-12 weeks
genotype: c-myc 25xI-SceI AID -/-
cell type: splenic B cells activated with anti-CD40 plus IL-4 for 4 days
treatment: DMSO
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| Treatment protocol |
1 micromolar DMSO
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| Growth protocol |
Splenic naïve B cells were purified via an anti-CD43 negative selection approach. The purified naïve B cells were cultured in RPMI 1640 medium supplimented with 10% FBS and activated with anti-CD40 plus IL-4 for 4 days to generate CSR-activated B cells, and simultaneously treated with control DMSO. Cells were transduced with pMX-I-SceI retrovirus to induce double strand breaks 24 h post activation.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Cells were lysed in lysis buffer (NaCl 200 mM, Tris.HCl 100 mM, EDTA 5mM, SDS 0.2%) containing 10 μg/ml Proteinase K at 56°C overnight. Genomic DNA was precipitated with isopropanol at room temperature, washed in ethanol 70% and resuspended in TE buffer (10mM Tris-HCl plus 1mM EDTA)
See Supplemental Experimental procedures from Chiarle et al. Cell 2011, paragraph Generation of HTGTS libraries by adapter-PCR.
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| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina MiSeq |
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| Data processing |
For HTGTS data, we applied prinseq 0.2 to remove sequences with exact PCR duplicates, mean quality score < 20 and length < 50. Next, effect reads for each sample were recognized by designed barcode, primer and bait portion. Lastly,the barcode, primer and bait portion of the remained sequences were masked for alignment analysis to determine translocation junctions as previously described(Chiarle et al.,Cell 2011). Briefly, we aligned sequences to the mouse reference genome (GRCm37/mm9) or human genome (GRCh38/hg38) using BLAT, and then filtered artificial junctions by removing PCR repeats (reads with same junction position in alignment to the reference genome and a start position in the read less than 3 bp apart), invalid alignments (including alignment scores < 30, reads with multiple alignments having a score difference < 4 and alignments having 10-nucleotide gaps) and ligation artifacts (for example, random HaeIII restriction sites ligated to bait breaksite). Translocation junction position was determined based on the genomic position of the 5’ end of the aligned read.
Genome_build: mm9
Supplementary_files_format_and_content: For HTGTS data, BED files contain all obtained translocation junctions information including chromosome,corrdinate,read name,align score and strand.
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| Submission date |
Feb 10, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Roberto Chiarle |
| E-mail(s) |
Roberto.Chiarle@childrens.harvard.edu
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| Organization name |
Children’s Hospital Boston and Harvard Medical School
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| Department |
Department of Pathology
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| Lab |
Roberto Chiarle
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| Street address |
300 Longwood Avenue
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| City |
Boston |
| State/province |
Massachusetts |
| ZIP/Postal code |
02115 |
| Country |
USA |
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| Platform ID |
GPL16417 |
| Series (1) |
| GSE77788 |
Phosphatidylinositol 3-Kinase (PI3K) delta blockade increases genomic instability in B cells |
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| Relations |
| BioSample |
SAMN04485258 |
| SRA |
SRX1570118 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM2059308_HTGTS_KO-DMSO_3.bed.gz |
247.0 Kb |
(ftp)(http) |
BED |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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