|
| Status |
Public on Sep 26, 2016 |
| Title |
REH-fusionKD-2 |
| Sample type |
RNA |
| |
|
| Source name |
Cell line
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: REH
shRNA target: ETV6/RUNX1 fusion
|
| Treatment protocol |
Cells were seeded at 1*10^6 cells/ml and treated with either shRNA against the ETV6/RUNX1 fusion or non-targeting shRNA.
|
| Growth protocol |
Cells were grown in RPMI supplemented with 10% fetal bovine serum, 1% glutamin and 1% penicilin/streptomycin
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was harvested with the miRNeasy mini kit (Qiagen) with Dnase treatment on-column.
|
| Label |
Cy3
|
| Label protocol |
Cyanine-3 (Cy3) labeled cRNA was prepared from 100 ng RNA using the One-Color Low Input Quick Amp Labling kit (Agilent) according to the manufacturer's instructions, followed by miRNeasy column purification (Qiagen). Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.
|
| |
|
| Hybridization protocol |
600 ng of Cy3-labelled cRNA (specific activity >6 pmol Cy3/ug cRNA) was fragmented at 60°C for 30 minutes in a reaction volume of 25µl containing 1x Agilent fragmentation buffer and 2x Agilent blocking agent following the manufacturers instructions. On completion of the fragmentation reaction, 25 µl of 2x GEx Hybridization buffer HI-RPM was added to the fragmentation mixture and hybridized to the array for 17 hours at 65°C in a rotating 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).
|
| Scan protocol |
Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner with SureScan High-Resolution Technology using one color scan setting for 8x60k array slides (Scan Area 61x21.6 mm, Scan resolution 3µm, Dye channel is set to Green).
|
| Data processing |
The scanned images were analyzed with Feature Extraction Software (Agilent) using default parameters (protocol GE1-105_Dec08 and Grid: 041648_D_F_20120615) to obtain background subtracted and spatially detrended Processed Signal intensities.
|
| |
|
| Submission date |
Apr 04, 2016 |
| Last update date |
Sep 26, 2016 |
| Contact name |
Farzaneh Ghazavi |
| E-mail(s) |
farzaneh.ghazavi@ugent.be
|
| Organization name |
Gent university hospital
|
| Department |
Center of medical genetics
|
| Street address |
De Pintelaan 185
|
| City |
Ghent |
| ZIP/Postal code |
9000 |
| Country |
Belgium |
| |
|
| Platform ID |
GPL19197 |
| Series (2) |
| GSE79868 |
Identification of ETV6/RUNX1 regulated lncRNAs |
| GSE79873 |
Unique long non-coding RNA expression signature in ETV6/RUNX1-driven B-cell precursor acute lymphoblastic leukemia |
|
