|
| Status |
Public on Jun 24, 2013 |
| Title |
mt DP with Dex+ culture, biological rep1 |
| Sample type |
RNA |
| |
|
| Source name |
Flk1+/PDGFRa+ mesoderm cells derived from Dnmt3a-/-Dnmt3b-/- ES cells, with further 4-days culture in the presence of dexamethasone to activate Gata4GR transgene
|
| Organism |
Mus musculus |
| Characteristics |
background strain: 129sv/Jae ES cell line (J1)
genotype: Dnmt3a-/-Dnmt3b-/-, stably expressing Gata4-glucocorticoid receptor ligand-binding domain fusion gene
clone name: 16G4.3
cell type: Flk1+/PDGFRa+ mesoderm cells
gata4gr induction: yes
|
| Treatment protocol |
The cells were collected using 0.25% trypsin-EDTA, and single cell suspensions were stained using anti-Flk1 and anti-PDGFRa antibodies. Flk1/PDGFRa double-positive cells were sorted by FACS-Aria (BD Biosciences). The cells were further cultured on type IV collagen-coated dishes in the differentiation medium (alpha-MEM supplemented with 10% FCS and 50 uM 2-mercaptoethanol) for 4 days in the presence of dexamethasone.
|
| Growth protocol |
ES cells were plated onto a type IV collagen-coated 10-cm dish in the differentiation medium (alpha-MEM supplemented with 10% FCS and 50 uM 2-mercaptoethanol) for 4 days.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated by TRIZOL Reagent (Invitrogen) and cleaned up by the RNeasy Mini Kit (Qiagen) according to the manufacturers’ protocols.
|
| Label |
biotin
|
| Label protocol |
Biotinylated cRNA were prepared using a two-cycle target-labeling assay according to the Affymetrix protocol from 50 ng total RNA (Small Sample Labeling Protocol Version II).
|
| |
|
| Hybridization protocol |
The 10 micro-g cRNA was hybridized at 45 C for 16 hours on the GeneChip Mouse Genome 430 2.0 array (Affymetrix) according to the Affymetrix Standard Protocol. GeneChips were washed and stained using the EukGE-Ws2v5_450 protocol in the Affymetrix Fluidics Station 450.
|
| Scan protocol |
GeneChips were scanned using the GeneChip Scanner 3000 7G (Affymetrix).
|
| Description |
Dex+_mutant_1
Flk1+/PDGFRa+ mesoderm cells derived from Dnmt3a-/-Dnmt3b-/- ES cells, with further 4-days culture in the presence of dexamethasone to activate Gata4GR transgene.
|
| Data processing |
Data values were calculated using the Bioconductor implementation (affy module) of the RMA algorithm in conjunction with quantiles normalisation of perfect match values (mismatches ignored). The specific function call used was: expresso(data, bgcorrect.method=rma, normalize.method=quantiles, pmcorrect.method=pmonly, summary.method=median.polish).
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| |
|
| Submission date |
Mar 26, 2012 |
| Last update date |
Jul 29, 2013 |
| Contact name |
Masaki Okano |
| E-mail(s) |
okano@cdb.riken.jp
|
| Phone |
81-78-306-3164
|
| Fax |
81-78-306-3167
|
| Organization name |
RIKEN
|
| Department |
Center for Developmental Biology
|
| Lab |
Laboratory for Mammalian Epigenetic Studies
|
| Street address |
2-2-3 Minatojima-Minamimachi, Chuo-ku
|
| City |
Kobe |
| State/province |
Hyogo |
| ZIP/Postal code |
650-0047 |
| Country |
Japan |
| |
|
| Platform ID |
GPL1261 |
| Series (1) |
| GSE36814 |
Role for DNA methylation in response to Gata4 activation in embryonic stem cell-derived mesoderm |
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