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| Status |
Public on May 13, 2016 |
| Title |
Input_DNA_H3.3WT |
| Sample type |
SRA |
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| Source name |
C3H10T1/2 cells
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| Organism |
Mus musculus |
| Characteristics |
cell type: C3H embryo-derived mesenchymal progenitor cells
genotype: Stably expressing FLAG-HA-tagged wildtype H3.3
passages: Passage 10-15
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| Growth protocol |
C3H10T1/2 cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM, Invitrogen) with 10% fetal bovine serum (FBS, CellGro).
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
~2x10^7 cells were fixed with 1% formaldehyde for 10 min at room temperature and then quenched with 125 mM glycine for 5 min. Cells were then lysed in SDS lysis buffer (1% SDS, 10 mM EDTA, 50 mM Tris-HCl) and sonicated for 1 h with Bioruptor (Diagenode) to obtain mononucleosomes. After centrifugation, the lysates were diluted with dilution buffer (167 mM NaCl, 0.01% SDS, 1.1% TritonX-100, 1.2 mM EDTA, 16.7 mM Tris-HCl) and incubated with αCbx2 (Active Motif, 39663) antibody bound to 75 μl protein A Dynal magnetic beads (Invitrogen) and incubated overnight at 4°C, with 5% kept as input DNA. Magnetic beads were washed with low salt buffer (150 mM NaCl; 0.1% SDS; 1% Triton X-100; 1 mM EDTA; 50 mM Tris-HCl), high salt buffer (500 mM NaCl; 0.1% SDS; 1% Triton X-100; 1 mM EDTA; 50 mM Tris-HCl), LiCl buffer (150 mM LiCl; 0.5% Na deoxycholate; 0.1% SDS; 1% Nonidet P-40; 1 mM EDTA; 50 mM Tris-HCl) and TE buffer (1 mM EDTA; 10 mM Tris-HCl). ChIP DNA was eluted, treated with Proteinase K (Roche) and recovered using Qiagen PCR purification kit.
libraries were prepared according to the Illumina TruSeq protocol
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2500 |
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| Description |
5% input DNA
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| Data processing |
Reads that passed the quality score were aligned using bowtie 1.0.0 with default parameters
Peaks were called using mosaics (threshold = 20, FDR= 0.05, maxgap = 10000bp, minsize = 5000bp, binSize= 200bp, two component model, Input-Only analysis)
RPKM-normalized read intensities in the peaks were determined for the H3.3WT and H3.3K36M samples
Genome_build: mm9
Supplementary_files_format_and_content: tab-separated file containing genomic coordinates for Cbx2 peaks and rpkm-normalized read-counts for H3.3WT and H3.3K36M samples for each peak
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| Submission date |
Dec 09, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Peter W Lewis |
| E-mail(s) |
pwlewis2@wisc.edu
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| Phone |
608-316-4388
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| Organization name |
University of Wisconsin-Madison
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| Department |
Department of Biomolecular Chemistry
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| Lab |
Room 2174, Wisconsin Institute for Discovery
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| Street address |
330 N Orchard Street
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| City |
Madison |
| State/province |
Wisconsin |
| ZIP/Postal code |
53715 |
| Country |
USA |
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| Platform ID |
GPL17021 |
| Series (2) |
| GSE69291 |
H3K36 mutations promote sarcomagenesis through genome-wide remodeling of H3K36 and H3K27 methylation |
| GSE75856 |
H3K36 mutations promote sarcomagenesis through genome-wide remodeling of H3K36 and H3K27 methylation [Cbx2_ChIP] |
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| Relations |
| BioSample |
SAMN04328030 |
| SRA |
SRX1473024 |
| 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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