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| Status |
Public on May 16, 2018 |
| Title |
3aKO H3K27me3_SpikeIn_ChIPSeq |
| Sample type |
SRA |
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| Source name |
embryonic stem cells
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| Organism |
Mus musculus |
| Characteristics |
strain background: 129S4/SvJae
genotype/variation: Dnmt3a-/-
cell type: embryonic stem cells
chip antibody: anti-H3K27me3 (Millipore, 07-449)
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| Growth protocol |
ES cells were cultured on 0.1% gelatin-coated dishes in DMEM (Gibco, 11965-092) supplemented with 15% Fetal Bovine Serum (Foundation, 900-108), 1×Non-Essential Amino Acids (Gibco, 11140-050), 1×GlutaMAX (Gibco, 35050-061), 1 μM 2-Mercaptoethanol (Sigma, M7522), 1×Sodium pyruvate (Sigma, S8636), 1×Penicillin/Streptomycin (Gibco, 15140-122) and 1000 U/ml LIF (Millipore, ESG1106).
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
2×10^6 cells were fixed for 10 minutes with 1% formaldehyde, followed by chromatin extraction and sonication to generate 200-500bp fragments. The sheared chromatin was incubated overnight at 4°C with H3K4me3 or H3K27me3 antibody. After incubation, 10 µl of Dynabeads Protein A magnetic beads previously washed in RIPA buffer was added, and samples were incubated for an additional 2 hours at 4°C. Bead-protein complexes were washed three times with RIPA buffer and twice with TE buffer. Genomic DNA was eluted for 2 hours at 68°C in 100 µl of Complete Elution Buffer (20 mM Tris, pH7.5, 5 mM EDTA, 50 mM NaCl, 1% SDS, 50 µg/ml proteinase K) and purified with MinElute PCR purification kit (Qiagen, 28004).
For ChIP-Seq, sequencing libraries were prepared using ThruPLEX DNA-seq 48D kit (Rubicon Genomics, R400406) following standard protocols.
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina NextSeq 500 |
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| Description |
3aKO H3K27me3
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| Data processing |
Raw reads treated by bisulfite were quality trimmed (Trimgalore), and uniquely mapped to the mm9 reference genome using BWA .
Chromatin marks were compared with Danpos2. Briefly, the experiments were input background-subtracted and normalized to the same read depth.
Genome_build: mm9
Supplementary_files_format_and_content: The bigWig files were reads density, created from Danpos2 wiggle files. More information may be found in the Danpos2 documentation.
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| Submission date |
Mar 25, 2018 |
| Last update date |
May 16, 2018 |
| Contact name |
xueqiu lin |
| E-mail(s) |
[email protected]
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| Organization name |
Stanford University
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| Street address |
450 Serra Mall
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| City |
Stanford |
| State/province |
CA |
| ZIP/Postal code |
94305 |
| Country |
USA |
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| Platform ID |
GPL19057 |
| Series (2) |
| GSE100957 |
The role of DNMT3A and TET1 in regulating promoter epigenetic landscapes |
| GSE112311 |
Genome-wide maps of histone modifications (SpikeIn) and Suz12 in WT, Dnmt3a-/-, Tet1-/- J1 and DKO ES cells. |
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| Relations |
| BioSample |
SAMN08793994 |
| SRA |
SRX3844086 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM3067533_3aKO_SpikeIn_K27me3_dup2_broad_treat_pileup.Fnor.smooth.bw |
127.6 Mb |
(ftp)(http) |
BW |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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