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| Status |
Public on Dec 31, 2020 |
| Title |
C4_RRBS |
| Sample type |
SRA |
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| Source name |
Control Biological Replicate 4
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| Organism |
Rattus norvegicus |
| Characteristics |
strain: Sprague Dawley
Sex: male
tissue: pre-frontal cortex
age: 3-5 wks (juvenile adolescent)
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| Treatment protocol |
prolonged unpredictable stress, two stressors per day including: restraint (4hr), forced swim (15C, 5min), elevated platform (30min), tilted cage with wet bedding on orbital shaking (45o angle, 100 cycle/min. 1hr)
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Genomic DNA was isolated using the ZR-Duet DNA/RNA MiniPrep kit (cat. D7001) from Zymo Research Corp. This kit is suitable for extraction of DNA and RNA molecules from a wide range of sources using trade-marked Clear-Spin column technology.
Libraries were prepared according to Diagenodes Premium RRBS Sequencing Kit (cat. C02030032). Briefly, genomic DNA was digested using MspI, restriction fragment ends were filled in to produce a 3`-"A" base for ligation of Illumina Sequencing Adapters, and size selection using AMPure XP beads. After size selection, samples were pooled and PCR-amplified for 25 cycles, and subsequently generated amplicons were treated with a Bisulfite Conversion Reagent. Following Bisulfite Conversion, further PCR amplification of 15 cycles was performed to enrich the library, and after amplification, AMPure XP Beads were used to remove remaining primer oligos and any library fragments digested as a result of Bisulfite Conversion.
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| Library strategy |
Bisulfite-Seq |
| Library source |
genomic |
| Library selection |
Reduced Representation |
| Instrument model |
Illumina HiSeq 2000 |
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| Description |
tissue punch
C7_CCAACA
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| Data processing |
Base-calling was performed by CASAVA. Resulting sequencing reads were demultiplexed using bcl2fastq_v1.8.4.
Sequenced reads were trimmed for adapter sequences, low-quality base-calls (< 20;phred 33), and too short reads (< 20 bases), and an artifact of RRBS library preparation (if captured during sequencing); then, the resulting trimmed reads were mapped to a bisulfite converted rn6 genome using Bismark_v0.19.0 with parameters: --directional -q --score-min L,0,-0.2 -p 2 --reorder --ignore-quals
Binary alignment mapping files were sorted using samtools and were processed using methylKit_v1.4.1 with the following parameters: mincov=10 minqual=20 read.context="CpG"
Proccessed files in methylKit were assessed for differential methylation using getMethDiff() with the following parameters: difference=15, qvalue=0.05, type="all"
Genome_build: rn6
Supplementary_files_format_and_content: For processed files, they are in .wig format and each sample has an individual .wig file. This format is a tab-delimited file with the following columns: chromosome, start position, end position, methylation percent. Methylation percent refers to number of methylated Cytosine calls / total number of cytosine calls.
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| Submission date |
Mar 31, 2020 |
| Last update date |
Jan 01, 2021 |
| Contact name |
Nicholas James Waddell |
| E-mail(s) |
nwaddell@fsu.edu
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| Organization name |
Florida State University
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| Department |
Biological Sciences
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| Lab |
Laboratory of Jian Feng
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| Street address |
319 Stadium Dr
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| City |
Tallahassee |
| State/province |
FL |
| ZIP/Postal code |
32306 |
| Country |
USA |
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| Platform ID |
GPL14844 |
| Series (2) |
| GSE147846 |
DNA Methyltransferase 3A Mediates the Sustained Effects of Stress on Synaptic Functions and Behaviors [RRBS] |
| GSE147847 |
DNA Methyltransferase 3A Mediates the Sustained Effects of Stress on Synaptic Functions and Behaviors |
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| Relations |
| BioSample |
SAMN14505973 |
| SRA |
SRX8037020 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM4447039_C4_CpG.wig.gz |
2.0 Mb |
(ftp)(http) |
WIG |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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