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| Status |
Public on Apr 25, 2019 |
| Title |
Fetal hypoxia RRBS 3 |
| Sample type |
SRA |
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| Source name |
Heart
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| Organism |
Rattus norvegicus |
| Characteristics |
strain: Sprague Dawley
tissue: heart
age: E21
treatment: hypoxia
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
For genomic DNA (gDNA) extraction, heart tissues were minced and digested with 20 μL proteinase K (20 mg/mL) in digestion buffer (10mM Tris-Hcl, pH 8.0 containing 0.5 M EDTA, 1% SDS, 10 m NaCl, and 5mM CaCl2) for 3 h at 55°C. Next the tissue lysate was twice phenol/chloroform/isoamyl: (25/24/1) alcohol extracted and the aqueous layer was treated with RNAse AþT1 for 1 h at 37°C. The lysate was again phenol/chloroform/isoamyl alcohol extracted and the gDNA was precipitated with equal volume of isopropanol in presence of 0.3 M sodium acetate at -20°C overnight. The gDNA was finally centrifuged, washed with 70% ethanol, air dried and reconstituted in 10 mM Tris-HCl, pH 8.0. And then, gDNA was denatured with 2 N NaOH at 42°C for 15 min, treated with sodium bisulfite at 55°C for 16 h, and purified by EZ DNA Methylation-Gold KitTM.
RRBS libraries were constructed by standard protocols of the NuGEN Ovation® Ultralow Methyl-Seq Library Systems.
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| Library strategy |
Bisulfite-Seq |
| Library source |
genomic |
| Library selection |
RANDOM |
| Instrument model |
NextSeq 550 |
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| Data processing |
Illumina Casava1.8 software used for basecalling.
Adaptor sequences were removed from the reads using trim_galore v0.3.7 with option ‘--rrbs’ disabled due to special design of NuGEN Ovation RRBS system. After adaptor trimming, NuGEN’s diversity trimming was performed to remove diversity adaptors. The trimmed reads were aligned to rat genome NCBI Rnor6.0 by using Bismark v0.16.3 with all default parameter settings. The methylation call files including the location of each CpG sites and the methylation percentage (beta value) were generated by the bismark_methylation_extractor
Differential methylation sites (DMC) analysis was performed by methylKit. CpGs with coverage ≥ 10 were used for DMC analysis. DMCs were identified when the difference of methylation percentages between control and hypoxia were greater than 10% and q-value < 0.05.
Differential methylation region (DMR) analysis was performed by DMAP and methylKit. We first used DMAP to generate CpG region profiles. CpG regions with coverage ≥ 20 were used for DMR analysis. methylKit was used to perform DMR analysis. Same criteria of DMCs selection was applied for DMRs identification.
Genome_build: Rnor6.0
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| Submission date |
Apr 16, 2019 |
| Last update date |
Apr 26, 2019 |
| Contact name |
Charles Wang |
| E-mail(s) |
[email protected]
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| Organization name |
Loma Linda University
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| Street address |
11021 Campus St
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| City |
Loma Linda |
| State/province |
CA |
| ZIP/Postal code |
92350 |
| Country |
USA |
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| Platform ID |
GPL25029 |
| Series (2) |
| GSE129874 |
Prenatal hypoxia exposure induced methylomic and transcriptomic alterations in rat hearts [BiSulfite-seq] |
| GSE129876 |
Prenatal hypoxia exposure induced methylomic and transcriptomic alterations in rat hearts |
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| Relations |
| BioSample |
SAMN11433282 |
| SRA |
SRX5693651 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM3723821_R1_H5_3.bismark.cov.gz |
18.7 Mb |
(ftp)(http) |
COV |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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