|
| Status |
Public on Dec 01, 2005 |
| Title |
hsf4_heat_shock_hyb_1 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
hsf4 larvae, untreated
|
| Organism |
Drosophila melanogaster |
| Characteristics |
Strain: cn bw hsf4
Stage: Late third instar
Stage Determination: blue-gut method
Tissue: Whole organism
|
| Biomaterial provider |
Scott Neal
|
| Treatment protocol |
Approximately 30 larvae were collected in 2 mL screwcap plastic vials and were returned to the 25ºC incubator for 90 min. Larvae were snap-frozen in liquid nitrogen immediately following the treatment.
|
| Growth protocol |
D. melanogaster strains were grown on standard yeast-agar medium supplemented with 0.05% (w/v) bromophenol blue at 25ºC with a 12 h/12 h light/dark cycle. Adult flies were allowed to lay eggs for 3 days on fresh media and wandering third instar larvae were collected upon their emergence from the food during the illuminated period. Staging was verified by the blue-gut method (Maroni and Stamey, 1983).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was isolated with TRIzol as described in Riedl at al., 2005.
|
| Label |
Cy3
|
| Label protocol |
The direct labeling protocol was performed as described by Neal et al., 2005.
|
| |
|
| Channel 2 |
| Source name |
hsf4 larvae, heat shock
|
| Organism |
Drosophila melanogaster |
| Characteristics |
Strain: cn bw hsf4
Stage: Late third instar
Stage Determination: blue-gut method
Tissue: Whole organism
|
| Biomaterial provider |
Scott Neal
|
| Treatment protocol |
Approximately 30 larvae were collected in 2 mL screwcap plastic vials and were heat shocked in an air incubator at 36ºC for 60 min and were returned to the 25ºC incubator for 30 min. Larvae were snap-frozen in liquid nitrogen immediately following the treatment.
|
| Growth protocol |
D. melanogaster strains were grown on standard yeast-agar medium supplemented with 0.05% (w/v) bromophenol blue at 25ºC with a 12 h/12 h light/dark cycle. Adult flies were allowed to lay eggs for 3 days on fresh media and wandering third instar larvae were collected upon their emergence from the food during the illuminated period. Staging was verified by the blue-gut method (Maroni and Stamey, 1983).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was isolated with TRIzol as described in Riedl at al., 2005.
|
| Label |
Cy5
|
| Label protocol |
The direct labeling protocol was performed as described by Neal et al., 2005.
|
| |
|
| |
| 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) 100-74, Cy5 (633 nm) 90-67 (laser power-PMT Gain). Quantification was performed using QuantArray v3 software (Perkin Elmer) using the adaptive algorithm without background subtraction.
|
| Description |
Array Number: 12281106
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 = neuromuscular junction
Keywords = heat stress
Keywords = thermotolerance
|
| Data processing |
Data were normalized using the Lowess algorith implemented in GeneTraffic DUO while ignoring the flagged values. Refer to the manuscript for further discussion of data handling.
|
| |
|
| Submission date |
Jul 25, 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) |
| GSE2998 |
Thermoprotection in the Drosophila hsf4 mutant |
|
| Data table header descriptions |
| ID_REF |
|
| CH1_RAW |
Raw fluorescence intensity - Cy3 |
| CH1_BKD_RAW |
Raw background fluorescence intensity - Cy3 |
| CH1_AREA |
Spot area - Cy3 |
| Ch1_Signal_Noise_Ratio |
Signal to noise ratio - Cy3 |
| CH2_RAW |
Raw fluorescence intensity - Cy5 |
| CH2_BKD_RAW |
Raw background fluorescence intensity - Cy5 |
| CH2_AREA |
Spot area - Cy5 |
| Ch2_Signal_Noise_Ratio |
Signal to noise ratio - Cy5 |
| 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 |
