Mouse methylome studies SRP082360 Track Settings
 
Epigenetic memory of obesity in mouse colonic epithelium [Colon]

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Study title: Epigenetic memory of obesity in mouse colonic epithelium
SRA: SRP082360
GEO: GSE85731
Pubmed: 29346762

Experiment Label Methylation Coverage HMRs HMR size AMRs AMR size PMDs PMD size Conversion Details
SRX2033416 Colon 0.696 5.1 47243 1475.1 17 1226.8 1075 18334.5 0.996 title: GSM2283298 LF1, WGBS, Mus musculus, Bisulfite-Seq; {"source_name": "colonic epithelium", "sex": "male", "age": "six-week old", "strain": "C57BL/6", "tissue": "colon", "diet": "10% fat diet (LF)"}
SRX2033417 Colon 0.690 5.4 48550 1466.8 13 1103.3 892 20236.9 0.996 title: GSM2283299 LF2, WGBS, Mus musculus, Bisulfite-Seq; {"source_name": "colonic epithelium", "sex": "male", "age": "six-week old", "strain": "C57BL/6", "tissue": "colon", "diet": "10% fat diet (LF)"}
SRX2033418 Colon 0.694 5.3 48306 1449.0 15 1120.1 937 19691.8 0.996 title: GSM2283300 LF3, WGBS, Mus musculus, Bisulfite-Seq; {"source_name": "colonic epithelium", "sex": "male", "age": "six-week old", "strain": "C57BL/6", "tissue": "colon", "diet": "10% fat diet (LF)"}
SRX2033419 Colon 0.688 5.1 45794 1515.9 16 1033.6 817 20603.6 0.996 title: GSM2283301 LF4, WGBS, Mus musculus, Bisulfite-Seq; {"source_name": "colonic epithelium", "sex": "male", "age": "six-week old", "strain": "C57BL/6", "tissue": "colon", "diet": "10% fat diet (LF)"}
SRX2033420 Colon 0.695 5.5 49553 1419.1 18 921.5 1012 19094.6 0.996 title: GSM2283302 LF5, WGBS, Mus musculus, Bisulfite-Seq; {"source_name": "colonic epithelium", "sex": "male", "age": "six-week old", "strain": "C57BL/6", "tissue": "colon", "diet": "10% fat diet (LF)"}
SRX2033421 Colon 0.691 6.1 51077 1376.0 22 1222.0 801 20851.1 0.996 title: GSM2283303 HF1, WGBS, Mus musculus, Bisulfite-Seq; {"source_name": "colonic epithelium", "sex": "male", "age": "six-week old", "strain": "C57BL/6", "tissue": "colon", "diet": "60% fat diet (HF)"}
SRX2033422 Colon 0.692 5.5 49932 1416.8 14 1168.6 832 20746.2 0.996 title: GSM2283304 HF2, WGBS, Mus musculus, Bisulfite-Seq; {"source_name": "colonic epithelium", "sex": "male", "age": "six-week old", "strain": "C57BL/6", "tissue": "colon", "diet": "60% fat diet (HF)"}
SRX2033423 Colon 0.686 5.8 49230 1415.3 20 1157.8 824 20233.3 0.996 title: GSM2283305 HF3, WGBS, Mus musculus, Bisulfite-Seq; {"source_name": "colonic epithelium", "sex": "male", "age": "six-week old", "strain": "C57BL/6", "tissue": "colon", "diet": "60% fat diet (HF)"}
SRX2033424 Colon 0.694 5.3 48528 1447.4 14 1095.6 938 19604.9 0.996 title: GSM2283306 HF4, WGBS, Mus musculus, Bisulfite-Seq; {"source_name": "colonic epithelium", "sex": "male", "age": "six-week old", "strain": "C57BL/6", "tissue": "colon", "diet": "60% fat diet (HF)"}
SRX2033425 Colon 0.694 5.2 47256 1483.1 14 1194.4 1021 19068.3 0.996 title: GSM2283307 HF5, WGBS, Mus musculus, Bisulfite-Seq; {"source_name": "colonic epithelium", "sex": "male", "age": "six-week old", "strain": "C57BL/6", "tissue": "colon", "diet": "60% fat diet (HF)"}
SRX2033426 Colon 0.696 6.0 49383 1412.1 24 1178.0 805 20123.0 0.996 title: GSM2283308 HFLF1, WGBS, Mus musculus, Bisulfite-Seq; {"source_name": "colonic epithelium", "sex": "male", "age": "six-week old", "strain": "C57BL/6", "tissue": "colon", "diet": "HF diet then switched to LF diet"}
SRX2033427 Colon 0.699 4.8 47445 1501.1 13 1375.5 733 21560.0 0.996 title: GSM2283309 HFLF2, WGBS, Mus musculus, Bisulfite-Seq; {"source_name": "colonic epithelium", "sex": "male", "age": "six-week old", "strain": "C57BL/6", "tissue": "colon", "diet": "HF diet then switched to LF diet"}
SRX2033428 Colon 0.694 5.1 46214 1507.4 17 1290.3 689 21524.8 0.996 title: GSM2283310 HFLF3, WGBS, Mus musculus, Bisulfite-Seq; {"source_name": "colonic epithelium", "sex": "male", "age": "six-week old", "strain": "C57BL/6", "tissue": "colon", "diet": "HF diet then switched to LF diet"}
SRX2033429 Colon 0.696 5.3 47023 1482.1 15 1395.1 1048 18655.6 0.996 title: GSM2283311 HFLF4, WGBS, Mus musculus, Bisulfite-Seq; {"source_name": "colonic epithelium", "sex": "male", "age": "six-week old", "strain": "C57BL/6", "tissue": "colon", "diet": "HF diet then switched to LF diet"}
SRX2033430 Colon 0.692 5.7 48933 1439.3 16 1328.9 924 19611.3 0.996 title: GSM2283312 HFLF5, WGBS, Mus musculus, Bisulfite-Seq; {"source_name": "colonic epithelium", "sex": "male", "age": "six-week old", "strain": "C57BL/6", "tissue": "colon", "diet": "HF diet then switched to LF diet"}

Methods

All analysis was done using a bisulfite sequnecing data analysis pipeline DNMTools developed in the Smith lab at USC.

Mapping reads from bisulfite sequencing: Bisulfite treated reads are mapped to the genomes with the abismal program. Input reads are filtered by their quality, and adapter sequences in the 3' end of reads are trimmed. This is done with cutadapt. Uniquely mapped reads with mismatches/indels below given threshold are retained. For pair-end reads, if the two mates overlap, the overlapping part of the mate with lower quality is discarded. After mapping, we use the format command in dnmtools to merge mates for paired-end reads. We use the dnmtools uniq command to randomly select one from multiple reads mapped exactly to the same location. Without random oligos as UMIs, this is our best indication of PCR duplicates.

Estimating methylation levels: After reads are mapped and filtered, the dnmtools counts command is used to obtain read coverage and estimate methylation levels at individual cytosine sites. We count the number of methylated reads (those containing a C) and the number of unmethylated reads (those containing a T) at each nucleotide in a mapped read that corresponds to a cytosine in the reference genome. The methylation level of that cytosine is estimated as the ratio of methylated to total reads covering that cytosine. For cytosines in the symmetric CpG sequence context, reads from the both strands are collapsed to give a single estimate. Very rarely do the levels differ between strands (typically only if there has been a substitution, as in a somatic mutation), and this approach gives a better estimate.

Bisulfite conversion rate: The bisulfite conversion rate for an experiment is estimated with the dnmtools bsrate command, which computes the fraction of successfully converted nucleotides in reads (those read out as Ts) among all nucleotides in the reads mapped that map over cytosines in the reference genome. This is done either using a spike-in (e.g., lambda), the mitochondrial DNA, or the nuclear genome. In the latter case, only non-CpG sites are used. While this latter approach can be impacted by non-CpG cytosine methylation, in practice it never amounts to much.

Identifying hypomethylated regions (HMRs): In most mammalian cells, the majority of the genome has high methylation, and regions of low methylation are typically the interesting features. (This seems to be true for essentially all healthy differentiated cell types, but not cells of very early embryogenesis, various germ cells and precursors, and placental lineage cells.) These are valleys of low methylation are called hypomethylated regions (HMR) for historical reasons. To identify the HMRs, we use the dnmtools hmr command, which uses a statistical model that accounts for both the methylation level fluctations and the varying amounts of data available at each CpG site.

Partially methylated domains: Partially methylated domains are large genomic regions showing partial methylation observed in immortalized cell lines and cancerous cells. The pmd program is used to identify PMDs.

Allele-specific methylation: Allele-Specific methylated regions refers to regions where the parental allele is differentially methylated compared to the maternal allele. The program allelic is used to compute allele-specific methylation score can be computed for each CpG site by testing the linkage between methylation status of adjacent reads, and the program amrfinder is used to identify regions with allele-specific methylation.

For more detailed description of the methods of each step, please refer to the DNMTools documentation.