|
| Status |
Public on Dec 05, 2006 |
| Title |
Rainbow trout hepatocycte 10nM EE2 exposure Fish B microarray number FD009_018b |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
Fish B hepatocytes
|
| Organism |
Oncorhynchus mykiss |
| Characteristics |
0.1% DMSO Control exposed
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Qiagen RNeasy mini kit according to manufacturer's protocol
|
| Label |
Cy3
|
| Label protocol |
Labeled cDNA was generated using a SuperScript Indirect cDNA Labeling System kit (Invitrogen) according to the manufacturer's instructions. For each sample 5.0 ug of total RNA was used to generate modified cDNAs.
|
| |
|
| Channel 2 |
| Source name |
Fish B hepatocytes
|
| Organism |
Oncorhynchus mykiss |
| Characteristics |
10nM EE2 exposed
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Qiagen RNeasy mini kit according to manufacturers protocol
|
| Label |
Cy5
|
| Label protocol |
Labeled cDNA was generated using a SuperScript Indirect cDNA Labeling System kit (Invitrogen) according to the manufacturer's instructions. For each sample 5.0 ug of total RNA was used to generate modified cDNAs.
|
| |
|
| |
| Hybridization protocol |
Microarray was prepared for hybridization by washing 2 x 5 min with 0.2% SDS, 5 x 1 min with MilliQ water followed by drying using centrifugation (5 min, 514 x g in 50 mL tube). Array was pre-hybridized in 5X SSC, 0.1% SDS, 3% BSA for 1.5 h at 49C, briefly washed 3 X 20 sec in MilliQ water and dried by centrifugation. Labeled cDNAs were hybridized to pre-warmed microarray in a formamide based buffer (25% formamide, 4X SSC, 0.5% SDS, 2X Denhardt's solution, and 4 ul of Genisphere LNA dT blocker) for 16 h at 49C. Coverslips were removed in 2X SSC, 0.1% SDS at 49C. The arrays were washed 1 x 10 min at 49C (2X SSC, 0.1% SDS). All subsequent washes were at room temperature; 2 x 5 min in 2X SSC, 0.1% SDS, 2 x 5 min in 1X SSC and 4 x 5 min in 0.1X SSC and dried by centrifugation.
|
| Scan protocol |
Fluorescent array images were collected immediately for both Cy3 and Cy5 using ScanArray Express (PerkinElmer). Image intensity data were extracted using Imagene 5.5 software (Biodiscovery).
|
| Description |
Oncorhynchus mykiss 10nM EE2 exposed hepatocytes Replicate 2 dye flip
|
| Data processing |
Only elements deemed present by Imagene software and with control values greater then the average base/proportional value were used. Expression ratios were generated using background corrected lowess normalized data using GeneSpringGX 7 software (Agilent Technologies, Palo Alto, Ca, USA).
|
| |
|
| Submission date |
Nov 30, 2006 |
| Last update date |
Dec 05, 2006 |
| Contact name |
Ben F Koop |
| E-mail(s) |
bkoop@uvic.ca
|
| Phone |
(250) 472-4067
|
| Organization name |
The University of Victoria
|
| Department |
Biology
|
| Lab |
Centre for Biomedical Research
|
| Street address |
PO Box 3020 STN CSC
|
| City |
Victoria |
| State/province |
BC |
| ZIP/Postal code |
V8W 3N5 |
| Country |
Canada |
| |
|
| Platform ID |
GPL2716 |
| Series (1) |
| GSE6416 |
Toxicogenomic responses in rainbow trout hepatocytes exposed to model chemicals and a synthetic mixture |
|
| Data table header descriptions |
| ID_REF |
Unique identifier. |
| VALUE |
log2 of (CH2_SIGNAL_MEDIAN/CH1_SIGNAL_MEDIAN) |
| CH1_FLAG |
Flags as defined by Imagene 5.6 software (Biodiscovery). 0 = present, 1 = marginal (user defined), 2 = absent |
| CH1_SIGNAL_MEAN |
Pixel intensity averaged over the local signal region. |
| CH1_BKD_MEAN |
Pixel intensity averaged over the local background region. |
| CH1_SIGNAL_MEDIAN |
Median pixel intensity computed over the local signal region. |
| CH1_BKD_MEDIAN |
Median pixel intensity computed over the local background region. |
| CH1_SIGNAL_MODE |
Mode pixel intensity computed over the local signal region (mode corresponds to the pick location in intensity distribution). |
| CH1_BKD_MODE |
Mode pixel intensity computed over the local background region. |
| CH1_SIGNAL_AREA |
Number of pixels in the local signal region. |
| CH1_BKD_AREA |
Number of pixels in the local background region. |
| CH1_SIGNAL_TOTAL |
Total pixel intensity summed over the local signal region. |
| CH1_BKD_TOTAL |
Total pixel intensity summed over the local background region. |
| CH1_SIGNAL_STDEV |
Standard deviation of pixel intensities over the local signal region. |
| CH1_BKD_STDEV |
Standard deviation of pixel intensities over the local background region. |
| CH2_FLAG |
Flags as defined by Imagene 5.6 software (Biodiscovery). 0 = present, 1 = marginal (user defined), 2 = absent |
| CH2_SIGNAL_MEAN |
Pixel intensity averaged over the local signal region. |
| CH2_BKD_MEAN |
Pixel intensity averaged over the local background region. |
| CH2_SIGNAL_MEDIAN |
Median pixel intensity computed over the local signal region. |
| CH2_BKD_MEDIAN |
Median pixel intensity computed over the local background region. |
| CH2_SIGNAL_MODE |
Mode pixel intensity computed over the local signal region (mode corresponds to the pick location in intensity distribution). |
| CH2_BKD_MODE |
Mode pixel intensity computed over the local background region. |
| CH2_SIGNAL_AREA |
Number of pixels in the local signal region. |
| CH2_BKD_AREA |
Number of pixels in the local background region. |
| CH2_SIGNAL_TOTAL |
Total pixel intensity summed over the local signal region. |
| CH2_BKD_TOTAL |
Total pixel intensity summed over the local background region. |
| CH2_SIGNAL_STDEV |
Standard deviation of pixel intensities over the local signal region. |
| CH2_BKD_STDEV |
Standard deviation of pixel intensities over the local background region. |
