|
| Status |
Public on Apr 30, 2015 |
| Title |
CG13624_KO_male_4d_rep1 |
| Sample type |
RNA |
| |
|
| Source name |
REPTOR_KO adults, 4d old, replicate 1
|
| Organism |
Drosophila melanogaster |
| Characteristics |
Sex: male
developmental stage: 4d old adults
genotype: w[1118];;CG13624-/-
|
| Treatment protocol |
All flies were grown on standard laboratory food at 25 degrees in a humidified room
|
| Growth protocol |
4d old male adults, growth controlled, synchronized Drosophila melanogaster flies
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Trizol extraction of total RNA was performed according to the manufacturer's instructions.
|
| Label |
Cy3
|
| Label protocol |
Cyanine-3 (Cy3) labeled cRNA was prepared from 0.5 ug RNA using the One-Color Microarray-Based Gene Expression Analysis Low Input Quick Amp Labeling kit (Agilent, version 6.6) according to the manufacturer's instructions, followed by RNAeasy column purification (QIAGEN, Valencia, CA, p/n 74104). Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 UV-VIS Spectrophotometer.
|
| |
|
| Hybridization protocol |
1.65 ug of Cy3-labelled cRNA (specific activity >6.0 pmol Cy3/ug cRNA) was fragmented at 60°C for 30 minutes in a reaction volume of 55 µl containing 1x Agilent fragmentation buffer and 2x Agilent blocking agent following the manufacturers instructions. On completion of the fragmentation reaction, 55 µl of 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to Agilent Drosophila (V2) Gene Expression Microarrays for 17 hours at 65ºC 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 (G2505C) using one color scan setting for 4x44k array slides (Scan Area 61x21.6 mm, Scan resolution 5µm, Dye channel is set to Green and Green PMT is set to 100%).
|
| Description |
REPTOR = CG13624
|
| Data processing |
The scanned images were analyzed with Feature Extraction Software 10.7 (Agilent) using default parameters (protocol GE1-v1_91 and Grid: 012391_D_20060331) to obtain background subtracted and spatially detrended Processed Signal intensities. Features flagged in Feature Extraction as Feature Non-uniform outliers were excluded.
|
| |
|
| Submission date |
Jan 09, 2015 |
| Last update date |
May 01, 2015 |
| Contact name |
Aurelio A Teleman |
| Organization name |
German Cancer Research Center (DKFZ)
|
| Street address |
Im Neuenheimer Feld 580
|
| City |
Heidelberg |
| ZIP/Postal code |
69120 |
| Country |
Germany |
| |
|
| Platform ID |
GPL16820 |
| Series (2) |
| GSE55221 |
REPTOR and REPTOR-BP regulate organismal metabolism and transcription downstream of mTORC1 |
| GSE64846 |
Gene expression data from control and REPTOR (=CG13624) KO male adults |
|
