|
| Status |
Public on Feb 01, 2013 |
| Title |
0.12mg/L TNT, replicate 6 |
| Sample type |
RNA |
| |
|
| Source name |
Whole daphnia (20 daphnia/replicate)
|
| Organism |
Daphnia magna |
| Characteristics |
agent: 0.12mg/L TNT
|
| Treatment protocol |
U.S. EPA. 1996. OPPTS 850.1300 Daphnid Chronic Toxicity Test
|
| Growth protocol |
U.S. EPA. 1996. OPPTS 850.1300 Daphnid Chronic Toxicity Test
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Qiagen Rneasy
|
| Label |
Cy3
|
| Label protocol |
Cyanine-3 (Cy3) labeled cRNA was prepared from 0.4 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 |
600 ng of Cy3-labelled cRNA (specific activity >9.0 pmol Cy3/ug cRNA) was fragmented at 60°C for 30 minutes in a reaction volume of 25 µl containing 1x Agilent fragmentation buffer and 2x Agilent blocking agent following the manufacturers instructions. On completion of the fragmentation reaction, 25 µl of 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to the custom-designed Agilent test array 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), 1 minute with 37°C GE Wash buffer 2 (Agilent), 30 seconds with Acetonitrile, and dried with Agilent Stabilization and Drying Solution.
|
| Scan protocol |
Slides were scanned immediately after washing on a GenePix 4200AL scanner at PMT gain level 420 using default setting (Scan resolution 5um, One channel-Green, Laser power 100%).
|
| Data processing |
The scanned images were analyzed with GenePix Pro 6.1 (Molecular Devices, Sunnyvale, CA) using default parameters and Agilient-provided GEML grid file 023710_D_F_20090417.xml to obtain signal intensities. GeneSpring GX (Agilent Technologies) was used to normalize data by quantile normalizaton and baseline transformation to the median of all samples. Statistical analysis was also performed in GeneSpring utilizing data from all microarray probes and consisted of one-way ANOVA to test for significant differential expression of microarray targets.
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| |
|
| Submission date |
Jan 31, 2013 |
| Last update date |
Feb 01, 2013 |
| Contact name |
Tanwir Habib |
| E-mail(s) |
thabib@sidra.org
|
| Organization name |
Sidra Medicine
|
| Street address |
Al Luqta Street
|
| City |
Doha |
| ZIP/Postal code |
26999 |
| Country |
Qatar |
| |
|
| Platform ID |
GPL16579 |
| Series (1) |
| GSE43960 |
THE GOOD, THE BAD, AND THE TOXIC: DECIPHERING HORMESIS IN DAPHNIA MAGNA EXPOSED TO AN ENERGETIC COMPOUND |
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