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Sample GSM11111 Query DataSets for GSM11111
Status Public on Dec 02, 2003
Title Apterous[56f] vs Apterous[56f]-49
Sample type RNA
 
Channel 1
Source name Apterous[56f] adult flies
Organism Drosophila melanogaster
Extracted molecule total RNA
 
Channel 2
Source name Apterous[56f] adult flies
Organism Drosophila melanogaster
Extracted molecule total RNA
 
 
Description Adult Drosophila melanogaster apterous[56f] strain flies were obtained from Carolina Biological Supply (Burlington, NC). Flies were quick frozen on dry ice and total RNA was extracted (Andrews et al, Genome Research 10:2030-2043) using Trizol reagent (GibcoBRL, Gaithersburg, MD). Probes were labeled with Cy3 (Probe 1) or Cy5 (Probe 2). To synthesize probes, mRNA was isolated by a single round of poly(A) selection using Oligotex resin (Qiagen, Valencia, CA). The purified mRNA was quantified using RiboGreen dye (Molecular Probes) in a fluorescent assay. RiboGreen dye was diluted 1:200 (v/v final) and mixed with known RNA concentrations (determined by absorbance at 260 nm) ranging from 1 to 5000 ng/ml. A Millennium RNA size ladder (Ambion, Austin, TX) was used to generate standard curves and unknown samples were diluted as necessary. Fluorescence was measured in 96-well plates with a FLUOstar fluorometer (BMG Lab Technologies, Germany) fitted with 485 nm (excitation) and 520 nm (emission) filters. Between 25 and 100 ng mRNA were separated on an Agilent 2100 Bioanalyzer (Agilent Technologies, Palo Alto, CA), to examine the mRNA size distribution. 200 ng of purified mRNA were converted to either a Cy3- or Cy5-labeled cDNA probe using a custom labeling kit (Incyte Genomics). Each reaction contained 50 mM Tris–HCl pH 8.3, 75 mM KCl, 15 mM MgCl2, 4 mM DTT, 2 mM dNTPs (0.5 mM each), 2 µg Cy3 or Cy5 random 9mer (Trilink, San Diego, CA), 20 U RNase inhibitor (Ambion), 200 U MMLV RNase H-free reverse transcriptase (Promega, Madison, WI) and mRNA. Correspondingly labeled Cy3 and Cy5 cDNA products were combined and purified on a size exclusion column, concentrated by ethanol precipitation and resuspended in hybridization buffer. Probes were added between two subarray slides and developed as described (Yue et al.2001, NAR. 29:e41). Microarrays were scanned on a GenePix 4000A scanner (Axon Instruments, Foster City, CA) at 535 nm. Signal intensities were initially stored and analyzed using GEMTools software (Incyte Genomics, Palo Alto CA). Background corrections were done by calculating the average Cy3/Cy5 signal to produce a balanced coefficient (Yue et al., 2001).
 
Submission date Oct 10, 2003
Last update date Apr 25, 2012
Contact name Brian Oliver
E-mail(s) briano@nih.gov
Phone 301-204-9463
Organization name NIDDK, NIH
Department LBG
Lab Developmental Genomics
Street address 50 South Drive
City Bethesda
State/province MD
ZIP/Postal code 20892
Country USA
 
Platform ID GPL20
Series (1)
GSE757 Drosophila melanogaster strain comparison

Data table header descriptions
ID_REF
DiffExpr Ratio of uncorrected Probe 1/Probe 2 signal intensity.
VALUE Ratio of uncorrected Probe 1/balanced Probe 2 signal intensity
Probe 1Signal Raw signal intensity data from Cy3 (Probe 1) channel.
Probe 1S/B Probe 1 signal minus background/background.
Probe 1Area% Percentage of area read from element spot.
Probe 2 BalancedSignal Raw signal intensity data from Cy5 channel balanced against Probe 1 signal. Intensity is adjusted by a balancing coefficient that takes the average Cy3 signal on an array/the average Cy5 signal on the array.
Probe 2Signal Raw signal intensity data from Cy5 (Probe 2) channel.
Probe 2S/B Probe 2 signal-background/background.
Probe 2Area% Percentage of area read from element spot.

Data table
ID_REF DiffExpr VALUE Probe 1Signal Probe 1S/B Probe 1Area% Probe 2 BalancedSignal Probe 2Signal Probe 2S/B Probe 2Area%
1 1.6 1.4 16192 159.7 62 11779 9816 161.9 62
2 2.9 2.4 7676 72.1 67 3186 2655 40 67
3 2.2 1.8 4756 43.1 73 2594 2162 31 73
4 2.8 2.3 17455 158.3 71 7534 6278 92 71
5 1.2 -1 13017 122.7 62 13486 11238 173.9 62
6 1.1 -1.1 11500 111.6 59 12528 10440 164.1 59
7 1.6 1.3 13581 135.5 64 10436 8697 143.6 64
8 1.2 1 5039 53.5 63 5008 4173 71.7 63
9 1.5 1.3 1996 22.5 82 1554 1295 23.3 82
10 1.4 1.2 469 5.8 94 404 337 6.7 94
11 1 -1.2 379 6.1 91 445 371 8.9 91
12 1.4 1.2 607 9.2 83 527 439 10.8 83
13 1.5 1.2 434 6.6 95 355 296 7.3 95
14 1.4 1.1 618 8.9 91 545 454 10.1 91
15 1.4 1.2 534 8.2 85 461 384 8.8 85
16 1.5 1.3 1464 20.3 73 1165 971 20 73
17 1.2 -1 133 2.6 100 134 112 3.2 100
18 1.2 1 144 2.8 100 139 116 3.2 100
19 -1 -1.2 1008 13.3 83 1250 1042 19.6 83
20 1.2 -1 205 3.7 100 212 177 4.3 100

Total number of rows: 31464

Table truncated, full table size 1231 Kbytes.




Supplementary data files not provided

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