|
| Status |
Public on Mar 02, 2012 |
| Title |
G9a_minus_minus_L2K_PolII_total |
| 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: G9a -/-, ERT2-Cre
passages: Passaged 12-15 times
chip antibody: Pol II
vendor: Abcam (ab5408)
lot: 722997
treatment: treated with transfection reagent only
|
| Treatment protocol |
Primary MEFs were immortalized at passage 2. At passage 5, MEFs were treated with either 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 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 Genome Analyzer II |
| |
|
| Description |
chromatin IP against Pol II in G9a -/- MEFs treated with transfection reagent only
|
| Data processing |
Raw data processed using onboard SCS/RTA 2.6, alignments performed using Bowtie keeping only unique reads with 2 or fewer mismatches in 36bp. Alignments summed in 100bp windows for wig files and integrated profiles.
|
| |
|
| Submission date |
Oct 19, 2010 |
| Last update date |
May 15, 2019 |
| Contact name |
Terry Fang |
| E-mail(s) |
[email protected]
|
| 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 |
GPL9250 |
| 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 |
SRX029285 |
| BioSample |
SAMN00116693 |
