|
| Status |
Public on Oct 22, 2010 |
| Title |
Cut wing discs 24-72h (Biological Replicate1, Dye-swap A) |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
cut wing discs 24 hours after implantation
|
| Organism |
Drosophila melanogaster |
| Characteristics |
strain: w118iso; 2iso; 3iso isogenic line from the DrosDel collection (Ryder et al. 2007)
tissue: wing imaginal discs and a 90º sector from the posterior (P) compartment, leaving a 3/4 anterior (3/4 A) fragment (Bosch et al. 2005)
developmental stage: third instar larvae implanted into recently eclosed Canton S females and kept at 25ºC for 24h (Bosch et al. 2005)
|
| Growth protocol |
Flies were kept on standard media at 25ºC.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA extraction was performed with RNeasy Protect Mini Kit (QIAGEN Inc.). Quality was assessed using the Eukaryote Total RNA Nano Assay on a 2100 Bioanalyzer (Agilent Technologies).
|
| Label |
Cy3
|
| Label protocol |
1.5ug total RNA was amplified and indirectly labeled with Amino-Allyl Messageamp II aRNA Amplification Kit (Ambion, Inc). Fragmentation was done with RNA Fragmentation Reagents #8740(Ambion, Inc).
|
| |
|
| Channel 2 |
| Source name |
cut wing discs 72 hours after implantation
|
| Organism |
Drosophila melanogaster |
| Characteristics |
strain: w118iso; 2iso; 3iso isogenic line from the DrosDel collection (Ryder et al. 2007)
tissue: wing imaginal discs and a 90º sector from the posterior (P) compartment, leaving a 3/4 anterior (3/4 A) fragment (Bosch et al. 2005)
developmental stage: third instar larvae implanted into recently eclosed Canton S females and kept at 25ºC for 72h (Bosch et al. 2005)
|
| Growth protocol |
Flies were kept on standard media at 25ºC.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA extraction was performed with RNeasy Protect Mini Kit (QIAGEN Inc.). Quality was assessed using the Eukaryote Total RNA Nano Assay on a 2100 Bioanalyzer (Agilent Technologies).
|
| Label |
Cy5
|
| Label protocol |
1.5ug total RNA was amplified and indirectly labeled with Amino-Allyl Messageamp II aRNA Amplification Kit (Ambion, Inc). Fragmentation was done with RNA Fragmentation Reagents #8740(Ambion, Inc).
|
| |
|
| |
| Hybridization protocol |
Samples were diluted in 2X Hybridization Buffer #5185-5973 (Agilent Technologies) and hybridization was carried out at 60ºC for 18 hours with a G2534A SureHyb Chamber in a G2545A Hybridization Oven (Agilent Technologies).
|
| Scan protocol |
GenePix Results (GPR) data files were obtained for each microarray with an Axon 4000B scanner and GenePix Pro 6 (Axon Instruments, Inc).
|
| Description |
The four microarrays hybridized for each pair of conditions were done in biological replicate pairs. Both arrays from each pair were hybridized with the same amplified RNA from sample and common reference (obtained using the Amino-Allyl Messageamp II aRNA Amplification Kit from Ambion, Inc) but with dyes (Cy3 and Cy5 from Amersham, Inc) swapped to take dye-bias into account.
|
| Data processing |
GPR files were analyzed with Limma package from BioConductor (Gentleman et al., 2004; Genome Biol 5, R80; Smyth, 2004; Statistical Applications in Genetics and Molecular Biology 3, Article 3) using the same criteria. Data was background corrected with the "normexp" method and normalized with OLIN (Futschik and Crompton, 2005; Bioinformatics 21, 1724-1726). Quality of spots was assesed by spot size, foreground versus background signals, saturation and coincidence between differently calculated ratio measures and R2 of regression ratio.
|
| |
|
| Submission date |
Jul 30, 2009 |
| Last update date |
Oct 22, 2010 |
| Contact name |
Sergi Beltran |
| Organization name |
Universitat de Barcelona
|
| Department |
Serveis Cientificotècnics
|
| Lab |
Unitat de Bioinformàtica
|
| Street address |
Baldiri Reixac 10
|
| City |
Barcelona |
| ZIP/Postal code |
08028 |
| Country |
Spain |
| |
|
| Platform ID |
GPL3797 |
| Series (1) |
| GSE17408 |
Transcriptional signature of Drosophila wing disc regeneration |
|
| Data table header descriptions |
| ID_REF |
|
| VALUE |
Log2 OLIN (bioconductor) normalized ratio signal intensity (Cy5/Cy3); represents 72h/24h |
| F635_MEDIAN |
Median feature pixel intensity at wavelength 635 nm (Cy5). |
| B635_MEDIAN |
Median feature background intensity at wavelength 635 nm (Cy5). |
| F532_MEDIAN |
Median feature pixel intensity at wavelength 532 nm (Cy3). |
| B532_MEDIAN |
Median feature background intensity at wavelength 532 nm (Cy3). |
| RATIO_OF_MEDIANS_(635/532) |
The ratio of the median intensities of each feature for each wavelength, with the median background subtracted.Not normalized. |
| MEDIAN_OF_RATIOS_(635/532) |
The median of pixel-by-pixel ratios of pixel intensities, with the median background subtracted.Not normalized. |
| RGN_RATIO_(635/532) |
The regression ratio of every pixel in a 2-feature-diameter circle around the center of the feature.Not normalized. |
| RGN_R2_(635/532) |
The coefficient of determination for the current regression value. |
| WEIGHT |
0.01 indicates a negative control or bad quality spot. 0.02 indicates a good quality spike-in control spot. |
