Dataset Information

Defining Regulatory Elements in the Human Genome Using Nucleosome Occupancy and Methylome Sequencing (NOMe-Seq).

ABSTRACT: NOMe-seq (nucleosome occupancy and methylome sequencing) identifies nucleosome-depleted regions that correspond to promoters, enhancers, and insulators. The NOMe-seq method is based on the treatment of chromatin with the M.CviPI methyltransferase, which methylates GpC dinucleotides that are not protected by nucleosomes or other proteins that are tightly bound to the chromatin (GpCm does not occur in the human genome and therefore there is no endogenous background of GpCm). Following bisulfite treatment of the M.CviPI-methylated chromatin (which converts unmethylated Cs to Ts and thus allows the distinction of GpC from GpCm) and subsequent genomic sequencing, nucleosome-depleted regions can be ascertained on a genome-wide scale. The bisulfite treatment also allows the distinction of CpG from CmpG (most endogenous methylation occurs at CpG dinucleotides) and thus the endogenous methylation status of the genome can also be obtained in the same sequencing reaction. Importantly, open chromatin is expected to have high levels of GpCm but low levels of CmpG; thus, each of the two separate methylation analyses serve as independent (but opposite) measures which provide matching chromatin designations for each regulatory element.NOMe-seq has advantages over ChIP-seq for identification of regulatory elements because it is not reliant upon knowing the exact modifications on the surrounding nucleosomes. Also, NOMe-seq has advantages over DHS (DNase hypersensitive site)-seq, FAIRE (Formaldehyde-Assisted Isolation of Regulatory Elements)-seq, and ATAC (Assay for Transposase-Accessible Chromatin)-seq because it also gives positioning information for several nucleosomes on either side of each open regulatory element. Here, we provide a detailed protocol for NOMe-seq that begins with the isolation of chromatin, followed by methylation of GpCs with M.CviPI and treatment with bisulfite, and ending with the creation of next generation sequencing libraries. We also include sequencing QC analysis metrics and bioinformatics steps that can be used to identify nucleosome-depleted regions throughout the genome.

SUBMITTER: Rhie SK

PROVIDER: S-EPMC6019634 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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Publications

Defining Regulatory Elements in the Human Genome Using Nucleosome Occupancy and Methylome Sequencing (NOMe-Seq).

Rhie Suhn Kyong SK Schreiner Shannon S Farnham Peggy J PJ

Methods in molecular biology (Clifton, N.J.) 20180101

NOMe-seq (nucleosome occupancy and methylome sequencing) identifies nucleosome-depleted regions that correspond to promoters, enhancers, and insulators. The NOMe-seq method is based on the treatment of chromatin with the M.CviPI methyltransferase, which methylates GpC dinucleotides that are not protected by nucleosomes or other proteins that are tightly bound to the chromatin (GpC<sup>m</sup> does not occur in the human genome and therefore there is no endogenous background of GpC<sup>m</sup>). ...[more]

PMID: 29605855

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Dataset Information

Defining Regulatory Elements in the Human Genome Using Nucleosome Occupancy and Methylome Sequencing (NOMe-Seq).

Publications

Defining Regulatory Elements in the Human Genome Using Nucleosome Occupancy and Methylome Sequencing (NOMe-Seq).

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