|
| Status |
Public on Dec 01, 2005 |
| Title |
Pilot_cold-shock_experiment. |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
dp males, untreated
|
| Organism |
Drosophila melanogaster |
| Characteristics |
Strain: dp cn bw cl
Age: 6-7 days post-eclosion
Sex: Male
Tissue: Whole organism
|
| Biomaterial provider |
Wensheng Qin and Scott J. Neal
|
| Treatment protocol |
Control flies were maintained at 25ºC in the treatment vessel for the duration of the experimental procedure. The flies were collected, snap frozen in liquid nitrogen and stored at –70ºC.
|
| Growth protocol |
Flies were cultured in a standard yeast sucrose medium, at 25ºC in a 12 h light/ 12 h darkness (L: D, 8 am to 8 pm) photoperiod. Newly emerged adults were removed from bottles after eclosion and transferred to fresh medium on which they were allowed to feed and oviposit for 1-2 days. Adult males (< 2 days post eclosion) were selected and transferred to fresh food bottles for 5 more days before being treated.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was isolated with TRIzol as described in Riedl et al., 2005.
|
| Label |
Cy5
|
| Label protocol |
The direct labeling protocol was performed as described by Neal et al., 2003.
|
| |
|
| Channel 2 |
| Source name |
dp males, 2 h cold shock at 0ºC, 30 min recovery
|
| Organism |
Drosophila melanogaster |
| Characteristics |
Strain: dp cn bw cl
Age: 6-7 days post-eclosion
Sex: Male
Tissue: Whole organism
|
| Biomaterial provider |
Wensheng Qin and Scott J. Neal
|
| Treatment protocol |
Flies were treated for 2 h at 0 +/- 0.5ºC, followed by a 30 min recovery at 25ºC (during the light portion of their photoperiod). The flies were collected, snap frozen in liquid nitrogen and stored at –70ºC.
|
| Growth protocol |
Flies were cultured in a standard yeast sucrose medium, at 25ºC in a 12 h light/ 12 h darkness (L: D, 8 am to 8 pm) photoperiod. Newly emerged adults were removed from bottles after eclosion and transferred to fresh medium on which they were allowed to feed and oviposit for 1-2 days. Adult males (< 2 days post eclosion) were selected and transferred to fresh food bottles for 5 more days before being treated.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was isolated with TRIzol as described in Riedl et al., 2005.
|
| Label |
Cy3
|
| Label protocol |
The direct labeling protocol was performed as described by Neal et al., 2003.
|
| |
|
| |
| Hybridization protocol |
Hybridizations were performed as in Neal et al., 2003. The respective Cy3 and Cy5 targets were suspended in 75 uL DIG EasyHyb solution (Roche, Mississauga, ON) and were competitively hybridized to the array for 16 hours in a CMT-GAPS hybridization chamber (Corning, Acton, MA).
|
| Scan protocol |
Scanning was performed with a ScanArray 4000 system (Perkin Elmer, Boston, MA) using the following settings: 10 um scan: Cy3(543 nm) 100-65, Cy5(633 nm) 100-58 (laser power-PMT Gain). Quantification was performed using QuantArray v3 software (Perkin Elmer) using the adaptive algorithm without background subtraction.
|
| Description |
Array Number: 12291544
The data in the file is not normalized. The ratio in the Value column was derived from normalized data (not shown) as discussed in the manuscript.
Keywords = cold hardening
|
| Data processing |
Data were normalized using the Lowess algorithm implemented in GeneTraffic DUO while ignoring the flagged values. Refer to the manuscript for further discussion of data handling.
|
| |
|
| Submission date |
Jul 24, 2005 |
| Last update date |
Jul 26, 2005 |
| Contact name |
Zak Razak |
| E-mail(s) |
z.razak@utoronto.ca
|
| Phone |
905-569-4664
|
| URL |
http://www.flyarrays.com
|
| Organization name |
University of Toronto
|
| Department |
Zoology Department
|
| Lab |
Canadian Drosophila Microarray Centre
|
| Street address |
3359 Mississauga Road
|
| City |
Mississauga |
| State/province |
ON |
| ZIP/Postal code |
L5L 1C6 |
| Country |
Canada |
| |
|
| Platform ID |
GPL311 |
| Series (1) |
| GSE2989 |
Cold hardening gene expression |
|
| Data table header descriptions |
| ID_REF |
|
| CH1_RAW |
Raw fluorescence intensity - Cy5 |
| CH1_BKD_RAW |
Raw background fluorescence intensity - Cy5 |
| CH1_AREA |
Spot area - Cy5 |
| Ch1_Signal_Noise_Ratio |
Signal to noise ratio - Cy5 |
| CH2_RAW |
Raw fluorescence intensity - Cy3 |
| CH2_BKD_RAW |
Raw background fluorescence intensity - Cy3 |
| CH2_AREA |
Spot area - Cy3 |
| Ch2_Signal_Noise_Ratio |
Signal to noise ratio - Cy3 |
| Ignore |
Quality measure exported from GeneTraffic; flag=1, good=0 |
| Ratio |
The ratio of CH2_Raw to CH1_Raw |
| VALUE |
The log2-transformed ratio of the Lowess-normalized fluorescence values (Ch2/Ch1) exported from GeneTraffic |
