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GEO help: Mouse over screen elements for information. |
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| Status |
Public on Oct 31, 2016 |
| Title |
genomic DNA from CD3+ blood cell donor T-cell 10 |
| Sample type |
SRA |
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| Source name |
Peripheral blood
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| Organism |
Homo sapiens |
| Characteristics |
disease state: normal CD3+ cells
immunophenotype: NA
subtype: NA
cell type: normal
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
DNA was extracted using the AllPrep DNA/RNA Mini Kit (QIAGEN)
Whole-genome bisulfite sequencing libraries were prepared using the Illumina TruSeq DNA Library preparation kit starting with 1 µg of DNA. End-repair, A-tailing and ligation of adaptors was performed according to the Illumina TruSeq protocol. Subsequent bisulfite conversion of the sequencing libraries was performed using reagents from the EpiTect Bisulfite kit (Qiagen). Bisulfite converted adaptor-ligated fragments were amplified by PCR using Pfu TurboCx Hot-start DNA polymerase (Stratagen).
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| Library strategy |
Bisulfite-Seq |
| Library source |
genomic |
| Library selection |
RANDOM |
| Instrument model |
Illumina Genome Analyzer IIx |
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| Data processing |
Bisulfite converted reads were aligned to reference hg19 as in Johnson MD, Mueller M, Game L, Aitman TJ: Single Nucleotide Analysis of Cytosine Methylation by Whole-Genome Shotgun Bisulfite Sequencing. In Current Protocols in Molecular Biology. John Wiley & Sons, Inc.; 2001
BisSNP was used for DNA methylation calling (Liu Y, Siegmund KD, Laird PW, Berman BP: Bis-SNP: Combined DNA methylation and SNP calling for Bisulfite-seq data. Genome Biology 2012, 13:R61)
The R-package bsseq (Hansen KD, Langmead B, Irizarry RA: BSmooth: from whole genome bisulfite sequencing reads to differentially methylated regions.) was used to smoothen methylation calls from WGBS data, segment data and call differentially methylated regions between sample pairs. To call a segment as a differentially methylated region (DMR) ? 3 consecutive CpG sites and an average difference > 0.3 in the DNA methylation levels between an ALL sample and the control were required. DMRs with an average mapability score (wgEncodeCrgMapabilityAlign100mer, download from UCSC table browser) of < 0.5 across segments and DMRs where the average coverage in any of the compared pair of samples was less than 3 or larger than 50 (except in pooled samples), were removed to ensure that potential mapping errors did not inflate DMR detection.
RNA-seq reads were mapped to the reference genome (human_g1k_v37) using Tophat2 (version 2.0.4) with default settings.
The number of fragments per transcript was calculated from Tophat bam files using the function featureCounts in Rsubread R-package and fragments per transcript per kilobase per million (FPKM) was computed by the function rpkm in the package edgeR. Transcript fragment counts were normalized to log2 counts using the voom function in the limma R-package
Genome_build: hg19
Supplementary_files_format_and_content: cpg site methylation level bedgraph: 4th column is methylation level (0-100) over a CpG dinucleotide
Supplementary_files_format_and_content: cpg site read coverage bedgraph: 4th column is read coverage over a CpG di nucleotide
Supplementary_files_format_and_content: expression bed: name column is ensemble gene id, score column is expression in fpkm
Supplementary_files_format_and_content: normalized expression bed: name column is ensemble gene id, score column is voom normalized expression.
Supplementary_files_format_and_content: [Table_S2_dmrs.txt] list of differentially methylated regions (tab delimited text). Columns are: comparison: samples compared, chr: chromosome, start: start of region, end: end of region, avgdiff(smoothed_meth): average difference in smoothed methylation in region, nCpGsites: number of CpG sites considered in region, rawdiff: average difference in non-smoothed methylation in region.
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| Submission date |
Dec 22, 2015 |
| Last update date |
Oct 31, 2016 |
| Contact name |
Jessica Nordlund |
| Organization name |
Uppsala University
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| Department |
Department of Medical Sciences
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| Lab |
Molecular Precision Medicine
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| Street address |
Box 1432, BMC
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| City |
Uppsala |
| ZIP/Postal code |
SE-75144 |
| Country |
Sweden |
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| Platform ID |
GPL10999 |
| Series (1) |
| GSE76270 |
DNA methylome analysis of pediatric acute lymphoblastic leukemia cells reveals stochastic de novo DNA methylation in CpG islands. |
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| Relations |
| BioSample |
SAMN04362298 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM1978800_T-cell_10_cpg_site_coverage.bedgraph.gz |
163.1 Mb |
(ftp)(http) |
BEDGRAPH |
| GSM1978800_T-cell_10_cpg_site_methylation_level.bedgraph.gz |
173.5 Mb |
(ftp)(http) |
BEDGRAPH |
| Raw data not provided for this record |
| Processed data provided as supplementary file |
| Processed data are available on Series record |
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