|
| Status |
Public on Mar 28, 2014 |
| Title |
G1_2 |
| Sample type |
RNA |
| |
|
| Source name |
FL-HSPCs, transduced with LeGO-iG-Gata1/shG1, grown 4 days in myeloid differentiation medium, duplicate 2
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: wild-type CD34+-hematopoietic stem and progenitor cells
genotype/variation (vector): LeGO-iG-Gata1/shG1
days: 4
|
| Treatment protocol |
FL-HSPCs were transduced with the indicated virus.
|
| Growth protocol |
FL-HSPCs cells were grown in RPMI, 20% FCS, 1% P/S, SCF (5ng/ml), GM-CSF (10ng/ml), G-SCF (10ng/ml), IL-3 (5ng/ml)
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNeasy Mini Kit (Qiagen)
|
| Label |
Cy3
|
| Label protocol |
Cyanine-3 (Cy3) labeled cRNA was prepared from 0.5 ug RNA using the One-Color Low RNA Input Linear Amplification PLUS kit (Agilent) according to the manufacturer's instructions, followed by RNAeasy column purification (QIAGEN, Valencia, CA). Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.
|
| |
|
| Hybridization protocol |
1.5 ug of Cy3-labelled cRNA (specific activity >10.0 pmol Cy3/ug cRNA) was fragmented at 60°C for 30 minutes in a reaction volume of 250 ml containing 1x Agilent fragmentation buffer and 2x Agilent blocking agent following the manufacturers instructions. On completion of the fragmentation reaction, 250 ml of 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to Agilent Whole Human Genome Oligo Microarrays (G4112A) for 17 hours at 65°C in a rotating Agilent hybridization oven. After hybridization, microarrays were washed 1 minute at room temperature with GE Wash Buffer 1 (Agilent) and 1 minute with 37°C GE Wash buffer 2 (Agilent), then dried immediately by brief centrifugation.
|
| Scan protocol |
Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner (G2505B) using one color scan setting for 1x44k array slides (Scan Area 61x21.6 mm, Scan resolution 10um, Dye channel is set to Green and Green PMT is set to 100%).
|
| Data processing |
The scanned images were analyzed with Feature Extraction Software 10.5 (Agilent) using default parameters to obtain background subtracted and spatially detrended Processed Signal intensities. Features flagged in Feature Extraction as Feature Non-uniform outliers were excluded.
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| |
|
| Submission date |
Mar 28, 2014 |
| Last update date |
Mar 28, 2014 |
| Contact name |
Jan-Henning Klusmann |
| E-mail(s) |
[email protected]
|
| Organization name |
Goehte University Frankfurt
|
| Department |
Pediatric Hematology and Oncology
|
| Street address |
Theodor-Stern-Kai 7
|
| City |
Frankfurt |
| ZIP/Postal code |
60590 |
| Country |
Germany |
| |
|
| Platform ID |
GPL6480 |
| Series (1) |
| GSE56332 |
GATA1s induces hyperproliferation of eosinophil precursors in Down syndrome transient leukemia |
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