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Status |
Public on Nov 10, 2017 |
Title |
MBDseq DMSO rep 4 |
Sample type |
SRA |
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Source name |
Gonadal adipose tissue
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Organism |
Mus musculus |
Characteristics |
strain: C57BL/6J age: 33 weeks generation: F4
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Treatment protocol |
Female C57BL/6J mice (25 females per treatment group) were randomly assigned to the different treatment groups and exposed via drinking water to 50 nM TBT or 0.1% DMSO vehicle (both diluted in 0.5% carboxymethyl cellulose in water to maximize solubility) for 7 days prior to mating. Chemicals were administered to the dams throughout pregnancy and lactation.We chose only 1 male and 2 females per litter for breeding. To randomize the breeding process as much as possible, we did not breed siblings and did not breed females from the same litter with the same male. Unexposed animals were bred to each other and TBT-exposed bred to each other. A similar approach was followed in generations F2-F4. Animals from F0-F4 were maintained on a standard diet (SD - Rodent Diet 20 5053*; PicoLab) throughout the experiment. F4 males and females (avoiding siblings within the same gender) were maintained on a higher fat diet (HFD - Mouse Diet 20 5058*; PicoLab) between weeks 19-25. Mice were subsequently returned to the SD for 8 more weeks prior to euthanasia at 33 weeks of age
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Growth protocol |
Male (n=25) and female (n=50) C57BL/6J mice (7 weeks of age) were purchased from the Jackson Laboratory (Sacramento, CA). Mice were housed in micro-isolator cages in a temperature-controlled room (21–22°C) with a 12 hr light/dark cycle. Water and food was provided ad libitum unless otherwise indicated. Animals were treated humanely and with regard for alleviation of suffering. All procedures conducted in this study were approved by the Institutional Animal Care and Use Committee of the University of California, Irvine. All tissue harvesting was performed with the dissector blinded to which groups the animals belonged to.
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Extracted molecule |
genomic DNA |
Extraction protocol |
Genomic DNA (gDNA) and RNA were isolated from male white gonadal adipose tissue using ZR-Duet™ DNA/RNA MiniPrep kit (Zymo Research) of animals sacrificed at 33 weeks of age (after the diet challenge). Nucleic acid samples from 4 randomly selected mice from control or TBT groups were submitted for DNA methylome and transcriptome analyses Sonicated gDNA (100-200 bp, 500 ng) was subjected to methylated DNA enrichment using the MethylMiner kit (Life Technologies), followed by deep sequencing library construction using the NEBNext Ultra kit (New England Biolabs). Sonicated gDNA without methylation enrichment was used as input control. Following the ENCODE recommendation for ChIP-seq, we aimed for 20 million mapped reads (75 nt single) for each gDNA sample
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Library strategy |
MBD-Seq |
Library source |
genomic |
Library selection |
MBD2 protein methyl-CpG binding domain |
Instrument model |
Illumina NextSeq 500 |
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Description |
MBD-seq_processed data.txt
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Data processing |
Statistical evaluation of TBT-dependent methylome and transcriptome variation in F4 male gonadal white adipose tissue was performed using R (version 3.3), and Bioconductor (version 3.3) packages MEDIPS (version 1.22), Rsubread (version 1.22), and edgeR (version 3.14). MEDIPS function MEDIPS.meth was used to estimate the statistical significance of MBD-seq reads differential coverage for GRCm38/mm10 mouse reference genomic 100-bp consecutive, non-overlapping windows with the following parameters: diff.method = edgeR, minRowSum = 10, and diffnorm = quantile. Rsubread function featureCounts was used to assigned uniquely mapped RNA-seq reads to GRCm38/mm10 mouse reference genome gene models and count reads with the following parameters: GTF.featureType = exon, GTF.attrType = gene_id, allowMultiOverlap = TRUE, nthreads = 24, and strandSpecific = 0 . edgeR functions cpm and glmQLFTest were used to estimate the number of counts per million per gene model and the statistical significance of RNA-seq reads differential coverage. Genome_build: GRCm38/mm10 Supplementary_files_format_and_content: RNA-seq_processed data.txt includes RNA-seq normalized counts per gene model for each sample, and results of the statistical analyses performed as described above. Supplementary_files_format_and_content: MBD-seq_processed data.txt includes MBD-seq read counts, and rpkm for each sample, and results of the statistical analyses performed as described above for GRCm38/mm10 mouse reference genomic 100-bp consecutive, non-overlapping windows for which P value was lower than 0.001. Supplementary_files_format_and_content: ATAC-seq_isochores_f3_processed data.txt and ATAC-seq_isochores_f4_processed data.txt include ATAC-seq read counts, and rpkm for each sample, and results of the statistical analyses performed as described above for GRCm38/mm10 mouse isochores obtained from isoFinder for sperm samples from F3 and F4 generations, respectively. ATAC-seq_SICER_f3_processed data.txt and ATAC-seq_SICER_f4_processed data.txt include ATAC-seq read counts, and normalized counts for each group of samples, and results of the statistical analyses performed as described above for GRCm38/mm10 mouse SICER islands for sperm samples from F3 and F4 generations, respectively.
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Submission date |
Oct 16, 2017 |
Last update date |
May 15, 2019 |
Contact name |
Raquel Chamorro-Garcia |
E-mail(s) |
rchamorr@ucsc.edu
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Organization name |
University of California Irvine
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Street address |
4351 Natural Sciences 2
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City |
Irvine |
ZIP/Postal code |
92697 |
Country |
USA |
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Platform ID |
GPL19057 |
Series (1) |
GSE105051 |
Ancestral perinatal obesogen exposure results in a transgenerational thrifty phenotype in mice |
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Relations |
BioSample |
SAMN07791970 |
SRA |
SRX3289550 |
Supplementary data files not provided |
SRA Run Selector |
Raw data are available in SRA |
Processed data are available on Series record |
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