|
| Status |
Public on Apr 29, 2011 |
| Title |
Bmi1 embryonic stem cells sample_2 (Bmi1_ESC_2) |
| Sample type |
RNA |
| |
|
| Source name |
Bmi1 transduced embryonic stem cells (Bmi1 ES cells).
|
| Organism |
Mus musculus |
| Characteristics |
strain: 129/Sv
|
| Treatment protocol |
Bmi1 transduced embryonic stem cells were cultured on gelatin coated dish in DMEM medium with 15% FSC and leukemia inhibitory factor (LIF).
|
| Growth protocol |
Bmi1 transduced embryonic stem cells were cultured on gelatin coated dish in DMEM medium with 15% FSC and leukemia inhibitory factor (LIF). Lentivirus containing mouse BMI1 cDNA was used to infect CCE ES cells in the presence of 8 microgram polybrene. Empty vector served as control. After two passages eGFP positive CCE ES cells were sorted by flow cytometry and cells were further grown in culture medium as above.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using RNeasy Mini kit (Qiagen, Hilden, Germany) with DNAse digestion using the maufacturer's buffers and protocols. RNA quality was assessed by Agilent 2100 Bioanalyzer System.
|
| Label |
biotin
|
| Label protocol |
Probe preparation was with 1 microgram RNA using the Affymetrix One-Cycle Target Labeling Kit according to the manufacturer's instructions. Labeled RNA was fragmented by heating to 94°C for 35 minutes in Fragmentatin buffer of the compnay.
|
| |
|
| Hybridization protocol |
Affymetrix arrays were prehybridized with 200 microliter MES hybridization buffer for 10 minutes at 45˚C with rotation (60 U/min) in an Affymetrix Hybridization Oven 640. After removing the prehybridization solution, 10 microgram of labeled cRNA in 200 microliter MES hybridization buffer were applied to the array leaving a small air bubble. Arrays were incubated overnight at 45˚C with rotation (60U/min) in the hybridization chamber.
|
| Scan protocol |
Arrays were scanned using Affymetrix GeneChip Scanner 3000.
|
| Data processing |
CEL files were processed using the BioConductor software package (www.bioconductor.org). Normalization was done employing RMA (Irizarry et al., Biostatistics 4, 249-264, 2003).
|
| |
|
| Submission date |
Apr 12, 2011 |
| Last update date |
Apr 30, 2011 |
| Contact name |
Martin Zenke |
| E-mail(s) |
Martin.Zenke@rwth-aachen.de
|
| Phone |
+49-241-80 80760
|
| Organization name |
Institute for Biomedical Engineering
|
| Department |
Cell Biology
|
| Street address |
Universitatsklinikum Aachen, RWTH
|
| City |
Aachen |
| State/province |
NRW |
| ZIP/Postal code |
52074 |
| Country |
Germany |
| |
|
| Platform ID |
GPL1261 |
| Series (1) |
| GSE20958 |
Polycomb Group Protein Bmi1 Promotes Hematopoietic Cell Development from ES Cells |
|
