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SRX2548113: GSM2481496: CBP57; Homo sapiens; OTHER
1 ILLUMINA (Illumina HiSeq 2500) run: 11.8M spots, 495.3M bases, 273.3Mb downloads

Submitted by: NCBI (GEO)
Study: Sexually dimorphic methylation of CD3+ T-lymphocyte DNA in offspring of overweight and obese mothers in a high risk, minority population in the Bronx
show Abstracthide Abstract
Maternal obesity impacts the health of offspring, increasing the risk of developing obesity and/or other metabolic dysregulation in childhood or later in life. Using a genome-wide methylation assay, we identified sex-dependent dysregulation of the methylome of CD3+ T-lymphocytes, a cell type that plays an important role in obesity and inflammatory diseases, in newborn offspring of overweight and obese mothers. Furthermore, the differentially methylated loci were targeted to regulatory regions of the genome, in imprinted genes and genes identified from GWAS as being related to type 2 diabetes and obesity. Overall design: DNA methylation was assessed globally at specific CpG sites in CD3+ T cell from 76 consenting women who delivered healthy, non-anomalous singleton AGA term neonates following an uncomplicated intrapartum course at the Weiler Division of Montefiore Medical Center.
Sample: CBP57
SAMN06318006 • SRS1966763 • All experiments • All runs
Organism: Homo sapiens
Library:
Instrument: Illumina HiSeq 2500
Strategy: OTHER
Source: GENOMIC
Selection: other
Layout: SINGLE
Construction protocol: High molecular weight genomic DNA was extracted from CD3+ T-cell from human cord blood. Custom adapters were created for HELP-tagging, referred to as AE and AS Index. As well as an Illumina adapter sequence, adapter AE contains an EcoP15I recognition site and a T7 promoter sequence, and adapter AS contains index sequences for multiplexing . Adapter AS contains an Illumina sequencing primer sequence. Five hundred nanograms of genomic DNA were digested with HpaII in separate 50 μl reactions and purified by phenol/chloroform extraction followed by ethanol precipitation. The digested genomic DNA was ligated to adapter AE using an NEB Quick Ligation Kit (25 μl of 2x Quick ligase buffer, HpaII-digested DNA, 0.5 μl of Adapter AE (0.1 μM), 1 μl of Quick Ligase in a final volume of 10 μl). After AE ligation, the products were purified using AMpure XP (Agencort), then digested with EcoP15I (NEB). The restriction fragments were end-repaired to inhibit to dimerization of adapters, and tailed with a single dA, at the 3’ end. After the dA tailing reaction, adapter AS was ligated to the dA-tailed fragments using an NEB Quick Ligation Kit (12.5 μl of 2x Quick ligase buffer, 1.25 μl of Adapter AS (1 μM), 1.25 μl of Quick Ligase in a final volume 25 μl). After ligation, products were purified using the MinElute PCR purification kit (Qiagen) and in vitro-transcribed using MEGAshortscript (Ambion). Following in vitro transcription, products were purified with an RNeasy clean-up kit (Qiagen) before reverse transcription was performed using the SuperScript III kit (Invitrogen). The first strand cDNA produced was used as a template for PCR using the following conditions: 96°C for 2 minutes, then 18 cycles of 96°C for 15 seconds and 72°C for 15 seconds followed by 5 minutes at 72°C for the final extension. After PCR, the library was purified using a QIAQuick PCR clean-up kit (Qiagen).
Experiment attributes:
GEO Accession: GSM2481496
Links:
Runs: 1 run, 11.8M spots, 495.3M bases, 273.3Mb
Run# of Spots# of BasesSizePublished
SRR524125411,793,492495.3M273.3Mb2020-02-20

ID:
3688931

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