Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Genome-wide DNA methylation analyses by the HELP assay

ABSTRACT: The HELP tagging assay was performed on purified genomic DNAs. The protocol was modified for NEBNext Multiplex Oligos for Illumina (NEB) from the original protocol4 (http://wasp.einstein.yu.edu/index.php/Protocol:HELP_tagging). Briefly, genomic DNA was digested by HpaII or MspI, the former only cutting at CCGG sequences where the central CG dinucleotide is unmethylated. AS and AE adapters were prepared by annealing two oligo DNAs separately. The first Illumina AE adapter was ligated to the compatible cohesive end created, juxtaposing an EcoP15I site beside the HpaII/MspI digestion site and allowing EcoP15I to digest within the flanking genomic DNA sequence. An A-overhang was created, allowing the ligation of the second Illumina AS adapter. This created both AE-insert-AS products and AS-insert-AS molecules. By performing a T7 polymerase-mediated in vitro transcription from a promoter sequence located on the AE adapter, we selectively enriched for the AE-insert-AS product. PCR amplification by NEBNext Multiplex Oligos was performed to generate a single-sized amplicons for Illumina sequencing. DNA methylation in Zscan4 positive and negative cells

ORGANISM(S): Mus musculus

SUBMITTER: Minoru Ko

PROVIDER: E-GEOD-51680 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Transient bursts of Zscan4 expression are accompanied by the rapid derepression of heterochromatin in mouse embryonic stem cells.

Akiyama Tomohiko T Xin Li L Oda Mayumi M Sharov Alexei A AA Amano Misa M Piao Yulan Y Cadet J Scotty JS Dudekula Dawood B DB Qian Yong Y Wang Weidong W Ko Shigeru B H SB Ko Minoru S H MS

DNA research : an international journal for rapid publication of reports on genes and genomes 20150831 5

Mouse embryonic stem cells (mESCs) have a remarkable capacity to maintain normal genome stability and karyotype in culture. We previously showed that infrequent bursts of Zscan4 expression (Z4 events) are important for the maintenance of telomere length and genome stability in mESCs. However, the molecular details of Z4 events remain unclear. Here we show that Z4 events involve unexpected transcriptional derepression in heterochromatin regions that usually remain silent. During a Z4 event, we se ...[more]

PMID: 26324425

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Dataset Information

Genome-wide DNA methylation analyses by the HELP assay

Publications

Transient bursts of Zscan4 expression are accompanied by the rapid derepression of heterochromatin in mouse embryonic stem cells.

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