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| Status |
Public on Jul 22, 2009 |
| Title |
M/E-cells expression constitutively active Mef2c |
| Sample type |
RNA |
| |
|
| Source name |
murine bone marrow cells, immortalized, 6h,24h,48h in expansion medium
|
| Organism |
Mus musculus |
| Characteristics |
cell type: bone marrow cells
genotype: M/E-Mef2c del/-
|
| Treatment protocol |
M/E-cells were harvested at three different time-points: 6h, 24h and 48h after seeding in fresh expansion-medium
|
| Growth protocol |
M/E-cells were grown in RPMI supplemented with 10% FCS, 1% Penicillin/Streptamycin, 1% Glutamine, 10 ng/ml recombinant murine IL6, GM-CSF, IL3 and 100 ng/ml murine recombinant SCF.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
extraction of total RNA was done at the facilities of Miltenyi Biotec, Bergisch-Gladbach, Germany. In Brief: Use of standard RNA extraction protocol (NucleoSpin® RNA II, Macherey-Nagel), the three samples of both M/E-celllines was pooled.
|
| Label |
Cy3
|
| Label protocol |
labeling of T7-based amplificated cRNA was done at the facilities of Miltenyi Biotec, Bergisch-Gladbach, Germany. In Brief: RNA samples were amplified and labeled using the Agilent Low RNA Input Linear Amp Kit (Agilent Technologies) following the manufacturer's protocol. Yield of cRNA and dye-incorporation was measured with ND-1000 Spectrophotometer (NanoDrop Technologies).
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| |
|
| Hybridization protocol |
hybridization was done at the facilities of Miltenyi Biotec, Bergisch-Gladbach, Germany. In Brief, the hybridization procedure was performed according to the Agilent 60-mer oligo microarray processing protocol using the Agilent Gene Expression Hybridization Kit (Agilent Technologies). Briefly, 1.65 μg Cy3-labeled fragmented cRNA in hybridization buffer was hybridized overnight (17 hours, 65 °C) to Agilent Whole Mouse Genome Oligo Microarrays 4x44K using Agilent’s recommended hybridization chamber and oven. Finally, the microarrays were washed once with 6x SSPE buffer containing 0.005% Nlauroylsarcosine for 1 min at room temperature followed by a second wash with preheated 0.06x SSPE buffer (37 °C) containing 0.005% N-lauroylsarcosine for 1 min. The last washing step was performed with acetonitrile for 30 sec.
|
| Scan protocol |
scanning was done at the facilities of Miltenyi Biotec, Bergisch-Gladbach, Germany. Fluorescence signals of the hybridized Agilent Microarrays were detected using Agilent’s Microarray Scanner System (Agilent Technologies).
|
| Description |
M/E-cells are rescued with an HDAC-binding mutant form of Mef2c (constitutively active).
murine bone marrow cells_immortalized by MLL/ENL-expression_6h,24h,48h pooled expansion medium
|
| Data processing |
data processing was done at the facilities of Miltenyi Biotec, Bergisch-Gladbach, Germany. The Agilent Feature Extraction Software (FES) was used to read out and process the microarray image files.
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| |
|
| Submission date |
Jul 21, 2009 |
| Last update date |
Jul 21, 2009 |
| Contact name |
Carol Stocking |
| E-mail(s) |
stocking@hpi.uni-hamburg.de
|
| Organization name |
Heinrich-Pette-Institut
|
| Street address |
Martinistrasse 52
|
| City |
Hamburg |
| ZIP/Postal code |
20251 |
| Country |
Germany |
| |
|
| Platform ID |
GPL4134 |
| Series (1) |
| GSE17231 |
Influence of MLL/ENL expression on murine bone marrow cells with a conditional Mef2c knockout or a Mef2c rescue |
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