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Sample GSM4505860 Query DataSets for GSM4505860
Status Public on Apr 30, 2020
Title normal N8 WGBS
Sample type SRA
 
Source name Adjacent normal tissue
Organism Homo sapiens
Characteristics tissue: adjacent normal esophageal tissue
tumor pathological stage: NA
tnm stage: NA
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)
 
Library strategy Bisulfite-Seq
Library source genomic
Library selection RANDOM
Instrument model Illumina HiSeq 4000
 
Description normal
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.
 
Submission date Apr 29, 2020
Last update date Apr 30, 2020
Contact name Wei Wu
E-mail(s) wei.wu@ucsf.edu
Phone 4157669898
Organization name University of California, San Franciso
Department Department of Medicine
Lab Trever Bivona
Street address 600 16th street
City San Francisco
State/province CA
ZIP/Postal code 94143
Country USA
 
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
Relations
BioSample SAMN14772591
SRA SRX8208806

Supplementary file Size Download File type/resource
GSM4505860_N8_CpG_methylation.bed.gz 133.4 Mb (ftp)(http) BED
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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