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Sample GSM3585211 Query DataSets for GSM3585211
Status Public on Jan 31, 2020
Title SEQ811
Sample type SRA
 
Source name Plasma
Organism Homo sapiens
Characteristics tissue: Plasma
disease diagnosis: Type 2 Diabetes
population: Chinese
sample: F25
batch: 1
t2dm_complications: 3
t2dm_nephropathy: 0
age: 62
binary for gender: 2
athero: 1
t2dm_comp: Yes
age_bin: 1
multiple_single: Multiple
Extracted molecule genomic DNA
Extraction protocol CfDNA samples from T2D patients were prepared from peripheral blood with EDTA anticoagulation. Briefly, 4 mL of peripheral blood was collected from each subject, and plasma was separated within 4 h by centrifuging twice, one at 1 350× g for 12 min, and another at 13 500× g for 5 min. The processed plasma samples were stored at –80 °C immediately. CfDNA was extracted using the Circulating Nucleic Acid Kit (Qiagen, Germany) according to the manufacturer’s instructions. On average, 1-2 ng of cfDNA was obtained from each cfDNA sample. The quality of cfDNA was checked using the standard molecular biology approach.
Each cfDNA sample was first repaired and ligated with adaptors. Next, the T4 bacteriophage enzyme β-glucosyltransferase (β-GT) was used to transfer an engineered glucose moiety containing an azide-group to 5hmC in duplex DNA. A biotin tag was then installed onto the azide group using Huisgen cycloaddition (click) chemistry, followed by capturing of 5hmC-containing DNA fragments using avidin beads. The 5hmC-Seal library for each cfDNA sample was then constructed through PCR amplification, followed by paired-end sequencing (PE38) at Shanghai Epican Genetech, Co. Ltd. (Shanghai, China), using the Illumina NextSeq500 platform according to Illumina’s protocol. We randomized cfDNA samples when constructing the 5hmC-Seal libraries and sequencing.
 
Library strategy OTHER
Library source genomic
Library selection other
Instrument model Illumina NextSeq 500
 
Description F25
cfDNA
processed data file: 5hmC_Count_Matrix.csv
Data processing Library strategy: 5hmC-Seal
Raw sequencing reads were trimmed for adaptor sequences using Trimmomatic.
Low quality bases were also trimmed to a minimum length of 30 bp, followed by alignment to the current human genome reference (hg19) using Bowtie2 with the end-to-end alignment mode. Read pairs were concordantly aligned with fragment length ≤ 500 bp and with average ≤ 1 ambiguous base and up to four mismatched bases per 100 bp length.
Alignments with Mapping Quality Score ≥ 10 were counted for gene bodies, according to the gene start and gene end annotations by the GENCODE Project (release 19), using featureCounts without strand information.
Peaks were called using PeaksFind version 2.2 with the following setting: ChIP threshold (0.2), Enrichment Fold (2.5), Rescue Fold (3).
The 5hmC-Seal libraries were sequenced to produce a median number of ~25 million reads in cfDNA samples, and a median number of ~13.5 million unique reads (~55% of total reads) were mapped to the ~22,000 gene body regions.
Genome_build: hg19 (GRCh37)
Supplementary_files_format_and_content: 5hmC_Count_Matrix.csv: Comma-separated text file
 
Submission date Jan 30, 2019
Last update date Jan 31, 2020
Contact name Wei Zhang
E-mail(s) wei.zhang1@northwestern.edu
Organization name Northwestern University
Department Preventive Medicine
Street address 680 N Lake Shore Drive, Suite 1400
City Chicago
State/province IL
ZIP/Postal code 60611
Country USA
 
Platform ID GPL18573
Series (1)
GSE125929 5-Hydroxymethylcytosines in Circulating Cell-free DNA Reveal Vascular Complications of Type 2 Diabetes
Relations
BioSample SAMN10849433
SRA SRX5313348

Supplementary data files not provided
SRA Run SelectorHelp
Raw data are available in SRA
Processed data are available on Series record

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