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| Status |
Public on Aug 25, 2015 |
| Title |
H3K27ac_SF268_rep2 |
| Sample type |
SRA |
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| Source name |
SF268 glioblastoma cells
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| Organism |
Homo sapiens |
| Characteristics |
cell line: SF268
cell type: Glioblastoma cells
chip antibody: Histone H3K27ac antibody
chip antibody vendor: Active Motif
chip antibody cat. #: 39133
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| Growth protocol |
SF268 cells were maintained in RPMI 1640 Medium, GlutaMAXT Supplement, 25mM HEPES (72400, Life Technologies) supplemented with 10% (v/v) fetal calf serum and 1 mM sodium pyruvate in a humidified incubator 37°C, 5% CO2 incubator.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Cells were cross-linked in the presence of 1% formaldehyde for 10 min at at room temperature. Crosslinking was stopped by addition of 125mM Glycine for 5 min. Cells were rinsed with 1x PBS and scraped off. Pellets were resuspended in buffer 1 (10 mM Tris (pH 8.0), 10 mM EDTA, 0.5 mM EGTA, 0.25% Triton X-100) and washed once buffer 2 (10 mM Tris (pH 8.0), 1 mM EDTA, 0.5 mM EGTA, 200 mM NaCl). Then cells were lysed in lysis buffer (50 mM HEPES/KOH (pH 7.5), 500 mM NaCl, 1 mM EDTA, 1% Triton X-100, 0.1% DOC, 0.1% SDS, protease inhibitors) and sonicated for 15min using a Covaris E210 (TR24-M (500035) and TR12x12 Tubes (520080), Settings: 5% duty cycle, intensity 4). 70ug of chromatin were incubated over night at 4°C with 7ul YAP antibody (Abcam, [EP1674Y] (ab52771) or 20ul TEAD1 antibody (BD Transduction LaboratoriesT (610922)), and for 2h with 25ul preblocked (tRNA, BSA) Dynabeads protein G. Beads were washed twice with lysis buffer, once with DOC buffer (10 mM Tris (pH 8.0), 0.25 M LiCl, 0.5% NP-40, 0.5% deoxycholate, 1 mM EDTA) and once with TE pH8.0, and bound chromatin was eluted in 1% SDS/0.1 M NaHCO3. After RNase A treatment, cross-linking was reversed by overnight incubation at 65 °C followed by proteinase K digestion. DNA was purified using the Minielute PCR purification kit (Qiagen). A sample of the input chromatin was treated in the same way to generate total input DNA. Input and ChIP-samples in duplicate were sequenced.
Libraries were prepared using the NEBNext ChIP-seq library reagent set for Illumina (New England BioLabs Inc.) omitting the size selection step before PCR enrichment.
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2500 |
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| Data processing |
Basecalling was performed with the Illumina RTA 1.17.21.3 software with default parameters
Fastq files were generated with the Illumina bcl2fastq-1.8.4 software with options --use-bases-mask Y50n,I6n --fastq-cluster-count 100000000 --no-eamss
Read mapping was performed using Bowtie version 1.0.0 with parameters -v 3 -m 1 --best --strata
Bigwig files were generated by extending the reads to 150bp and calculating for each genomic position the read density normalized to one million reads in the library
Peak calling was performed using peakzilla with default parameters
Genome_build: hg19
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| Submission date |
Sep 29, 2014 |
| Last update date |
May 15, 2019 |
| Contact name |
Anais Flore Bardet |
| Organization name |
IGBMC
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| Street address |
1 rue Laurent Fries
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| City |
Illkirch |
| ZIP/Postal code |
67404 |
| Country |
France |
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| Platform ID |
GPL16791 |
| Series (1) |
| GSE61852 |
YAP1 exerts its transcriptional control via TEAD-mediated activation of enhancers |
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| Relations |
| BioSample |
SAMN03083669 |
| SRA |
SRX716562 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM1515745_H3K27ac_SF268_rep2.bw |
525.2 Mb |
(ftp)(http) |
BW |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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