|
| Status |
Public on Jan 27, 2015 |
| Title |
d10PREd10CRE |
| Sample type |
genomic |
| |
|
| Channel 1 |
| Source name |
K27 ChIP from iPS [ChIPped DNA]
|
| Organism |
Mus musculus |
| Characteristics |
tissue: Embryonic brain
age: E13.5
antibody: H3K27me3 (17-622, Millipore)
|
| Growth protocol |
Embryonic brain dissection / standard ES cell growth conditions
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Forebrains were dissected from E13.5 mouse embryos, fixed in 1% formaldehyde for 15 minutes at room temperature, washed three times with cold phosphate buffer solution (PBS) and stored at -80°C. One forebrain was used for each experiment. ChIP was performed according to (Lee et al., 2006) using 2 μg of anti-H3K4me3 (ab8580, Abcam), H3K27me3 (17-622, Millipore), 4μg Suz12 (ab12073, abcam) and 4μg Ring1B (39663, Active Motif) and EZview Red protein G/A Affinity Gel (Sigma). Immunoprecipitated and whole cell extract DNA (input) were treated with RNAseA, proteinase K and purified by two rounds of extraction with phenol/chloroform/isoamyl alcohol. ChIP and input DNA were amplified using ligation-mediated PCR (Lee et al. 2006)
|
| Label |
Biotin
|
| Label protocol |
1 ug of ChIP and input DNA was fragmented and labeled using GeneChip WT Double-Stranded DNA Terminal Labeling Kit (Affymetrix)
|
| |
|
| Channel 2 |
| Source name |
Input DNA
|
| Organism |
Mus musculus |
| Characteristics |
age: e13.5
tissue: Embryonic brain
|
| Growth protocol |
Embryonic brain dissection / standard ES cell growth conditions
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Forebrains were dissected from E13.5 mouse embryos, fixed in 1% formaldehyde for 15 minutes at room temperature, washed three times with cold phosphate buffer solution (PBS) and stored at -80°C. One forebrain was used for each experiment. ChIP was performed according to (Lee et al., 2006) using 2 μg of anti-H3K4me3 (ab8580, Abcam), H3K27me3 (17-622, Millipore), 4μg Suz12 (ab12073, abcam) and 4μg Ring1B (39663, Active Motif) and EZview Red protein G/A Affinity Gel (Sigma). Immunoprecipitated and whole cell extract DNA (input) were treated with RNAseA, proteinase K and purified by two rounds of extraction with phenol/chloroform/isoamyl alcohol. ChIP and input DNA were amplified using ligation-mediated PCR (Lee et al. 2006)
|
| Label |
Biotin
|
| Label protocol |
1 ug of ChIP and input DNA was fragmented and labeled using GeneChip WT Double-Stranded DNA Terminal Labeling Kit (Affymetrix)
|
| |
|
| |
| Hybridization protocol |
Affymetrix hybridization protocol
|
| Scan protocol |
Arrays were scanned on an Affymetrix Scanner 3000 7G
|
| Description |
Sonicated genomic DNA
Sample name: d10PREd10CRE
|
| Data processing |
Array data were quantile normalized with input replicate groups and scaled to medial feature intensity of 500 using TAS software (Affymetrix, www.affymetrix.com). For each genomic position, a data set was generated consisting of all (PM-MM) pairs mapping within a sliding window of 250 bp. Exported text file was submitted to an in house algorithm including setting probes mapping within the deletions to 0 before applying a running mean approach with a higher pnderation for significant pvalues.
|
| |
|
| Submission date |
Jan 30, 2013 |
| Last update date |
Jan 27, 2015 |
| Contact name |
Patrick Schorderet |
| E-mail(s) |
patrick.schorderet@gmail.com
|
| Organization name |
EPFL
|
| Department |
ISREC
|
| Lab |
UPDUB
|
| Street address |
SV 2842 (Bâtiment SV) Station 19
|
| City |
Lausanne |
| State/province |
Vaud |
| ZIP/Postal code |
1015 |
| Country |
Switzerland |
| |
|
| Platform ID |
GPL14183 |
| Series (1) |
| GSE43915 |
In embryo analysis of H3K27 trimethylation suggests a two-step process involving PRC2 interacting sequences and high GC content |
|
