Homo sapiens
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA386890 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR5560400.fastq.gz | Fastqsanger.gz | |||
| SRR5560400_1.fastq.gz | Fastqsanger.gz | |||
| SRR5560400_2.fastq.gz | Fastqsanger.gz | |||
| SRR5560410.fastq.gz | Fastqsanger.gz | |||
| SRR5560410_1.fastq.gz | Fastqsanger.gz |
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Project description:Cytosine methylation of DNA CpG dinucleotides in gene promoters is an epigenetic modification that regulates gene transcription. While many methods exist to interrogate methylation states, no current methods offer large-scale, targeted, single CpG resolution. We report an approach combining bisulfite treatment followed by RainDance microdroplet PCR with next-generation sequencing to assay the methylation state of 50 genes in the regions 1 kb upstream and downstream of their transcription start sites. Wildtype and hypermethylated Jurkat DNA (New Englad Biolabs) was treated with bisulfite to convert all unmethylated cytosines to uracil. Following bisulfite treatment, targeted amplification was carried out using a custom primer library and microdroplet PCR. PCR product was sheared to 200 bp and ligated to sequencing adapters following standard protocols. Sequencing was conducted with single-end 100 bp reads on an Illumina GAIIx for wild type Jurkat DNA or Jurkat CpG DNA with a single sample per lane.
2011-06-25 | E-GEOD-30179 | biostudies-arrayexpress
Project description:Cytosine methylation of DNA CpG dinucleotides in gene promoters is an epigenetic modification that regulates gene transcription. While many methods exist to interrogate methylation states, no current methods offer large-scale, targeted, single CpG resolution. We report an approach combining bisulfite treatment followed by RainDance microdroplet PCR with next-generation sequencing to assay the methylation state of 50 genes in the regions 1 kb upstream and downstream of their transcription start sites.
2011-06-25 | GSE30179 | GEO
Project description:We report a method for specific capture of an arbitrary subset of genomic targets for single molecule bisulfite sequencing, and for digital quantitation of DNA methylation at a single nucleotide resolution. We used targeted bisulfite sequencing to characterize the changes of DNA methylation during the de-differentiation of human fibroblasts into hybrid stem cells, and into induced pluripotent stem cells. We compared the methylation level of approximately 66,000 CpG sites within 2020 CpG islands on chromosome 12, chromosome 20, and 34 selected regions. A total of 288 differentially methylated regions were identified between fibroblasts and pluripotent cells. Methylation cluster analysis revealed distinct methylation patterns between fibroblasts and pluripotent cells. Furthermore iPS cells are globally more methylated than human embryonic stem cells, which could be due to the reprogramming process. This targeted bisulfite sequencing method is particularly useful for efficient and large-scale analysis of DNA methylation in organisms with large genomes. Experiment Overall Design: Comparison of DNA methylation on 2020 CpG islands and 34 other selected regions among eleven human ES, iPS and fibroblast lines.
2009-08-28 | E-GEOD-15007 | biostudies-arrayexpress
Project description:DNA methylation stabilizes developmentally programmed gene expression states. Aberrant methylation is associated with disease progression and is a common feature of cancer genomes. Presently, few methods enable quantitative, large-scale, single-base resolution mapping of DNA methylation states in desired regions of a complex mammalian genome. Here, we present an approach that combines array-based hybrid selection and massively parallel bisulfite sequencing to profile DNA methylation in genomic regions spanning hundreds of thousands of bases. This single molecule strategy enables methylation variable positions to be quantitatively examined with high sampling precision. Using bisulfite capture, we assessed methylation patterns across 324 randomly selected CpG islands (CGI) representing more than 25,000 CpG sites. A single lane of Illumina sequencing permitted methylation states to be definitively called for >90% of target sties. The accuracy of the hybrid-selection approach was verified using conventional bisulfite capillary sequencing of cloned PCR products amplified from a subset of the selected regions. This confirmed that even partially methylated states could be successfully called. A comparison of human primary and cancer cells revealed multiple differentially methylated regions. More than 25% of islands showed complex methylation patterns either with partial methylation states defining the entire CGI or with contrasting methylation states appearing in specific regional blocks within the island. We observed that transitions in methylation state often correlate with genomic landmarks, including transcriptional start sites and intron-exon junctions. Methylation, along with specific histone marks, was enriched in exonic regions, suggesting that chromatin states can foreshadow the content of mature mRNAs. Keywords: DNA methylation profiling by massively parallel sequencing Keywords: Epigenetics Targeted examination of DNA methylation in two human cell types by combining array capture and bisulfite sequencing. In addition, this study examined two histone marks in the breast tumor cell line MDA-MB-231.
2009-07-08 | E-GEOD-17001 | biostudies-arrayexpress
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2015-07-28 | E-GEOD-70317 | biostudies-arrayexpress
Project description:We report a method for specific capture of an arbitrary subset of genomic targets for single molecule bisulfite sequencing, and for digital quantitation of DNA methylation at a single nucleotide resolution. We used targeted bisulfite sequencing to characterize the changes of DNA methylation during the de-differentiation of human fibroblasts into hybrid stem cells, and into induced pluripotent stem cells. We compared the methylation level of approximately 66,000 CpG sites within 2020 CpG islands on chromosome 12, chromosome 20, and 34 selected regions. A total of 288 differentially methylated regions were identified between fibroblasts and pluripotent cells. Methylation cluster analysis revealed distinct methylation patterns between fibroblasts and pluripotent cells. Furthermore iPS cells are globally more methylated than human embryonic stem cells, which could be due to the reprogramming process. This targeted bisulfite sequencing method is particularly useful for efficient and large-scale analysis of DNA methylation in organisms with large genomes.
2009-04-27 | GSE15007 | GEO
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