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| Status |
Public on Apr 30, 2020 |
| Title |
normal N7 WGBS |
| Sample type |
SRA |
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| Source name |
Adjacent normal tissue
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| Organism |
Homo sapiens |
| Characteristics |
tissue: adjacent normal esophageal tissue
tumor pathological stage: NA
tnm stage: NA
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Frozen tissues of surgically removed specimens were used for genomic DNA extraction. Genomic DNA was extracted with QIAmp DNA Mini kits (Qiagen) from fresh frozen tissue samples. 1μg gDNA was fragmented by sonication with a base pair peak of 300 bp for the resulting fragment, and adaptors were then ligated to both ends of the fragments. Bisulfite conversion was performed to whole genomes of ten pairs of ESCC and matched normal tissues, where converts cytosine residues of the dinucleotide CpG to uracil but leaves methylated cytosine unaffected. PCR amplification and purification were carried out. The uracil-binding pocket of KAPA HiFi DNA Polymerase has been inactivated, enabling amplification of uracil-containing DNA.
The high quality of the library was estimated by The Qubit® 3.0 Fluorometer. Bisulfite conversion success ratio is 99.18% and 99.49%, respectively in normal and ESCC samples
The WGBS library was sequenced on an Illumina HiSeq2500 sequencers and generated 400M of paired-end reads (2x125bp)
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| Library strategy |
Bisulfite-Seq |
| Library source |
genomic |
| Library selection |
RANDOM |
| Instrument model |
Illumina HiSeq 4000 |
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| Description |
normal
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| Data processing |
Illumina Casava1.7 software used for basecalling.
Firstly, we trimmed using Trim Galore! (v0.4.1) to remove Illumina adaptors with the options of ' --paired --length 50 --clip_1 6 --clip_2 6 '. After trimming, around 90 Gbp of data remained per sample. Then trimmed reads were aligned to the HG19 reference genome using BSMAP (v2.89) with the option of '-p 8 -R'. Then SAMtools (v. 1.3.1) was used to sort by genomic coordination and make a bam file index. Picard Tools (v.1.92) is used to remove PCR duplicates. After deduplication, ~70Gbp remained per sample. Lastly, we ran MOABS (v. 1.3.4) to compute the methylation ratio per CpG with the option of '--cytosineMinScore 20 --skipRandomChrom 1 -p 4 --keepTemp 0 --processPEOverlapSeq 1 --requiredFlag 2 --excludedFlag 256 --minFragSize 110 --reportCpX G --qualityScoreBase 0 --trimRRBSEndRepairSeq 0 --trimWGBSEndRepairPE1Seq 5 --trimWGBSEndRepairPE2Seq 5'. Around 95% of CpGs are covered by at least five reads.
Given the methylation ratio and coordination computed, we built a data matrix, whose columns are samples and rows are CpGs.
Genome_build: hg19
Supplementary_files_format_and_content: tab-delimited text files include methylation ration (methylated reads/total reads at a given CpG) for each Sample.
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| Submission date |
Apr 29, 2020 |
| Last update date |
Apr 30, 2020 |
| Contact name |
Wei Wu |
| E-mail(s) |
wei.wu@ucsf.edu
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| Phone |
4157669898
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| Organization name |
University of California, San Franciso
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| Department |
Department of Medicine
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| Lab |
Trever Bivona
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| Street address |
600 16th street
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| City |
San Francisco |
| State/province |
CA |
| ZIP/Postal code |
94143 |
| Country |
USA |
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| Platform ID |
GPL20301 |
| Series (2) |
| GSE149608 |
Multi-omics analysis reveals divergent epigenetic regulation of gene expression and drivers of esophageal squamous cell carcinoma (WGBS) |
| GSE149612 |
Multi-omics analysis reveals divergent epigenetic regulation of gene expression and drivers of esophageal squamous cell carcinoma |
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| Relations |
| BioSample |
SAMN14772592 |
| SRA |
SRX8208805 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM4505859_N7_CpG_methylation.bed.gz |
131.3 Mb |
(ftp)(http) |
BED |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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