|
| Status |
Public on Mar 07, 2012 |
| Title |
MEFs_H3K9me2_DMSO |
| Sample type |
SRA |
| |
|
| Source name |
MEF
|
| Organism |
Mus musculus |
| Characteristics |
strain: C57BL/6
cell type: Fibroblasts from day 13.5 embryos immortalized with SV40-LTA
genotype: Wild-type
passages: Passaged 12-15 times
chip antibody: H3K9me2
vendor: Abcam (ab1220)
lot: GR38973
|
| Treatment protocol |
Primary MEFs were immortalized at passage 2. At passage 5, MEFs were treated witheither EtOH as a vehicle control or Tamoxifen to induce deletion of G9a. MEFs were either left untreated, treated with Lipofectamine 200, the tranfection reagent or tranfected with PolyIC.
|
| Growth protocol |
MEFs were grown in 10 cm plates with DMEM medium containing 15% FBS, Pen/Strep, Glutamine, NEAA and 2-mercaptoethanol. After immortalization with SV40-LTA, MEFs were grown in 15 cm plates with the same medium except FBS was 10%. DC were isolated from WT C57Bl/6, G9af/f, G9a;Vav-cre or IFNaR1 -/- mice that subcutaneously injected with B16-Flt3L cells. 2 weeks later splenic DCs were enriched for the CD11c+ population using CD11c MACS beads. ChIP was carried out directly on ex vivo isolated cells.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Lysates were clarified from sonicated nuclei and histone-DNA complexes were isolated with antibody. Libraries were prepared according to Illumina's instructions accompanying the DNA Sample Kit (Part# 0801-0303). Briefly, DNA was end-repaired using a combination of T4 DNA polymerase, E. coli DNA Pol I large fragment (Klenow polymerase) and T4 polynucleotide kinase. The blunt, phosphorylated ends were treated with Klenow fragment (3' to 5' exo minus) and dATP to yield a protruding 3- 'A' base for ligation of Illumina's adapters which have a single 'T' base overhang at the 3’ end. After adapter ligation DNA was PCR amplified with Illumina primers for 15 cycles and library fragments of ~250 bp (insert plus adaptor and PCR primer sequences) were band isolated from an agarose gel. The purified DNA was captured on an Illumina flow cell for cluster generation. Libraries were sequenced on the Genome Analyzer following the manufacturer's protocols.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
chromatin IP against H3K9me2 in WT MEFs treated with DMSO
|
| Data processing |
Raw data processed using onboard SCS/RTA, alignments performed using Bowtie keeping only unique reads with 2 or fewer mismatches in 36bp. Alignments summed in 100bp windows for wig files
|
| |
|
| Submission date |
Mar 06, 2012 |
| Last update date |
May 15, 2019 |
| Contact name |
Terry Fang |
| E-mail(s) |
tfang@rockefeller.edu
|
| Phone |
212-327-8265
|
| Fax |
212-327-8258
|
| Organization name |
Rockefeller University
|
| Street address |
1230 York Ave
|
| City |
New York |
| State/province |
NY |
| ZIP/Postal code |
10065 |
| Country |
USA |
| |
|
| Platform ID |
GPL13112 |
| Series (2) |
| GSE22102 |
Histone H3 lysine 9 di-methylation as an epigenetic signature of the interferon response (sequencing) |
| GSE24826 |
Histone H3 lysine 9 di-methylation as an epigenetic signature of the interferon response |
|
| Relations |
| SRA |
SRX128094 |
| BioSample |
SAMN00809340 |
