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| Status |
Public on Oct 11, 2012 |
| Title |
tgf beta H3K9ac |
| Sample type |
SRA |
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| Source name |
lung adenocarcinoma cells:A549, the histone was h3k9ac,treated by tgf-beta
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| Organism |
Homo sapiens |
| Characteristics |
cell line: A549
treatment: TGFbeta
chip antibody: H3K9ac
chip antibody manufacturer: Millipore
chip antibody catalog #: 17-658
chip antibody lot #: NG1839561
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| Treatment protocol |
A549 cells were treated with TGF-β1 (5 ng/mL) and/or sorafenib (5 μM) for 48 h. Cells treated with vehicle only (DMSO) served as controls.
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| Growth protocol |
Human alveolar epithelial A549 cells were obtained from ATCC (Manassas, VA, USA) and cultured in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% fetal bovine serum (FBS) (Biochrom, Berlin, Germany) at 37°C with 5% CO2.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
ChIP-seq libraries were prepared using the following the protocol.The cells were cross-linked by a final concentration of 1% formaldehyde following incubation for 10 min at 37°C. Next, the cells were collected by centrifugation, washed twice with ice cold PBS, and dissolved in 500 μl SDS lysis buffer (1% SDS; 10 mM EDTA; protease inhibitors; 50 mM Tris-HCl, pH 8.0) followed by incubation for 30 min on ice. The lysates were sonicated at high power with 8x30 sec pulses in a BioruptorTM200 to generate chromatin fragments between 100 and 500 bp. Cellular debris was removed by centrifugation. Aliquots of 100 μl of the lysate were diluted 1:10 in ChIP dilution buffer (0.01% SDS; 1.1% Triton X-100; 1.2 mM EDTA; 167 mM NaCl; protease inhibitors; 16.7 mM Tris-HCl, pH 8.0). Then, 4ug of each antibody against H3K9ac, H3K4me3, H3K27me3 and H3K9me3 was added to 40ul of protein A/G Dynabeads (equal volume) and incubated for 3 h at 4°C with rotation. The beads were washed with cold PBS to remove free antibodies and added to diluted lysate followed by an overnight incubation at 4°C on a rotating platform. Sequentially, the immunocomplex plus beads were washed with 1 ml of the following buffers: low salt wash buffer (0.1% SDS; 1% Triton X-100; 2 mM EDTA; 150 mM NaCl; 20 mM Tris-HCl, pH 8.0), high salt wash buffer (0.1% SDS; 1% Triton X-100; 2 mM EDTA; 500 mM NaCl; 20 mM Tris-HCl, pH 8.0) and LiCl wash buffer (0.25 M LiCl; 1% Nonidet P-40; 1% sodium deoxycholate; 1 mM EDTA; 10 mM Tris-HCl, pH8.0). Finally, the beads were washed twice with 1 ml TE buffer (1 mM EDTA; 10 mM Tris-HCl, pH 8.0), and the immune complexes were reverse cross-linked directly by adding 200 μl of elution buffer (1% SDS, 100 mM NaHCO3) in the presence of proteinase K to a final concentration of 0.1 mg/ml and incubated for 3 h at 65°C. DNA in the supernatant was extracted with equal volumes of phenol/chloroform, precipitated with ethanol in the presence of glycogen and eluted in 30ul of nuclease-free H2O. The purified DNA was used for cluster generation and standard single-end sequencing with 50 bp reads (E50) using Illumina Hiseq 2000.
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2000 |
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| Description |
A549,lung adenocarcinoma cells treated by tgf-beta,H3K9ac
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| Data processing |
Illumina BclConverter-1.9.0 software was used for basecalling.
Adaptor sequences were removed, and low-quality sequence reads were trimmed. The clean ChIP-seq reads were mapped to hg19 reference with Soap aligner 2.21, parameters used the defaults.
further analisys used unique mapped reads after duplication, differential histone modification regions(DHMR) were found by t-test(p<0.01) and twice difference of signal in enriched region with poission test P<0.001
Genome_build: hg19
Supplementary_files_format_and_content: bed files contained chromsome,reads strart site,reads end sites;wig files include each samples ChIP signal on enriched sites.
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| Submission date |
Oct 10, 2012 |
| Last update date |
May 15, 2019 |
| Contact name |
Desheng Gong |
| E-mail(s) |
gds19870718@163.com
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| Organization name |
Agricultural Genomes Institute at Shenzhen
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| Street address |
No.7 PengFei road
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| City |
Shenzhen |
| ZIP/Postal code |
518120 |
| Country |
China |
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| Platform ID |
GPL11154 |
| Series (1) |
| GSE41479 |
Sorafenib inhibits epithelial-mesenchymal transition through an epigenetic-based mechanism in human lung epithelial cells |
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| Relations |
| SRA |
SRX193428 |
| BioSample |
SAMN01760812 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM1018050_sora_H3K9me3.bed.gz |
114.7 Mb |
(ftp)(http) |
BED |
| GSM1018050_tgf-beta_H3K9ac.wig.gz |
31.9 Mb |
(ftp)(http) |
WIG |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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