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| Status |
Public on Jul 03, 2013 |
| Title |
CMS-IP_mm_fc_10wk_IP |
| Sample type |
SRA |
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| Source name |
Frontal cortex from 10 week old male mouse brain
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| Organism |
Mus musculus |
| Characteristics |
strain: C57Bl/6
genotype: wild type
tissue: brain (frontal cortex)
cell type(s): all
age: 10 weeks (P70)
gender: male
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| Growth protocol |
The mouse lines C57Bl/6 and S100b-eGFP (B6;D2-Tg(S100B-EGFP)1Wjt/J), both from Jackson Laboratories, ME were bred and maintained in our animal facility in 12 h light/dark cycles with food ad libitum. To produce the S100b-eGFP animals used for FACS, animals were crossed to C57BL/6 to produce heterozygotes for eGFP expression. Tet2-/- mice were produced by targeted disruption of the Tet2 gene. Animals were weaned in groups of 3-4 per cage, and used at the postnatal time-points indicated. Three animals obtained from separate litters were processed together for DNA and RNA isolations.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Genomic DNA was purified from cells using the DNeasy Blood and Tissue Kit (Qiagen, Valencia, CA, USA) according to the manufacturer’s instructions. Cytosine 5-methylenesulphonate immunoprecipitation (CMS-IP) sequencing libraries were made as follows. Genomic DNA was fragmented to 100-200 bp using a Covaris S2 (Covaris, Woburn, MA, USA), end repaired, 3’ adenylated, ligated to methylated Illumina adaptor oligonucleotides, and bisulfite converted as per the MethylC-Seq protocol, descibed previously [Lister R. et al., Hotspots of aberrant epigenomic reprogramming in human induced pluripotent stem cells. Nature 471, 68-73 (2011)]. Bisulfite conversion of hmC to cytosine 5-methylenesulphonate (CMS) was followed by immunoprecipitation of genomic DNA fragments containing CMS with a specific antiserum to CMS. Bisulfite converted genomic DNA was first denatured for 10 minutes at 95°C (0.4 M NaOH, 10 mM EDTA), neutralized by addition of cold 2 M Ammonium Acetate pH 7.0, incubated with anti-CMS antiserum in 1x IP buffer (10 mM sodium phosphate pH 7.0, 140 mM NaCl, 0.05% Triton X-100) for 2 h to overnight at 4°C, and then precipitated with Protein G beads. Precipitated DNA was washed 3 times with 1x IP buffer and eluted with Proteinase K, then purified with Phenol Chloroform. Following immunoprecipitation, sequencing libraries were amplified by 8 cycles of PCR.
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| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina HiSeq 2000 |
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| Description |
Other: CMS-IP
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| Data processing |
Data were mapped to the Homo sapiens reference genome (hg18) or the Mus musculus reference genome (mm9).
MethylC-Seq, TAB-Seq, CMS-IP and hmC-gluc-IP-Seq analyses were performed as described in Lister, Mukamel et al. Global epigenomic reconfiguration during mammalian brain development. Science (2013).
RNA-Seq RPKM data were generated using Bowtie2 and Tophat2. Gene expression values were calculated using Cufflinks2.
genome build: mm9
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| Submission date |
Jun 25, 2013 |
| Last update date |
May 15, 2019 |
| Contact name |
Joseph R Ecker |
| E-mail(s) |
ecker@salk.edu
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| Phone |
8584534100
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| Organization name |
HHMI-Salk-Institute
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| Department |
Genomic Analysis Laboratory
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| Lab |
Ecker lab
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| Street address |
10010 North Torrey Pines Road
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| City |
La Jolla |
| State/province |
CA |
| ZIP/Postal code |
92037 |
| Country |
USA |
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| Platform ID |
GPL13112 |
| Series (1) |
| GSE47966 |
Global epigenomic reconfiguration during mammalian brain development |
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| Relations |
| SRA |
SRX314963 |
| BioSample |
SAMN02213462 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM1173798_CMS-IP_mm_fc_10wk_ip.mapped.txt.gz |
11.4 Gb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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