|
| Status |
Public on Oct 01, 2015 |
| Title |
ZA-5_PA-2 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
PA
|
| Organism |
Rhagoletis pomonella |
| Characteristics |
individual: PA-2
dev_time_point: L3 feeding larva
fruit: apple
tissue: whole 3rd instar larvae
|
| Growth protocol |
Infested apple and snowverry fruit were collected from field sitse near Bellingham, WA, USA, and staged in the lab at 21C, 14:10 LD.
Wandering larvae and pupae were collected daily, allowed to sit at 21C for 10 days, then transferred to 4C for 20 weeks of simulated winter
Following 20 weeks at 4C, diapausing pupae were moved to 21C. Pupae breaking diapause were reared to adulthood.
Adults were crossed in several cages to produce F1 offspring that were layed as eggs into agar domes.
Eggs were transplanted into snowberry and apple fruits, then reared to the third larval instar.
Feeding 3rd instar larvae were snap frozen and RNA-extracted.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
larvae were homogenized in tri-reagent and total RNA was extracted from each individual using the Ambion RiboPure® Kit
|
| Label |
Cy3
|
| Label protocol |
Labeled aRNA from four individuals from each time point was generated using Quick-Amp labeling Kit (Agilent Technologies, Inc.) following the manufacturer recommendations.
We used 250ng of total RNA for reverse transcription to synthesize 1st strand cDNA, then generated labeled aRNA according to the manufacturer’s protocol
|
| |
|
| Channel 2 |
| Source name |
ZA
|
| Organism |
Rhagoletis zephyria |
| Characteristics |
individual: ZA-5
dev_time_point: L3 feeding larva
fruit: apple
tissue: whole 3rd instar larvae
|
| Growth protocol |
Infested apple and snowverry fruit were collected from field sitse near Bellingham, WA, USA, and staged in the lab at 21C, 14:10 LD.
Wandering larvae and pupae were collected daily, allowed to sit at 21C for 10 days, then transferred to 4C for 20 weeks of simulated winter
Following 20 weeks at 4C, diapausing pupae were moved to 21C. Pupae breaking diapause were reared to adulthood.
Adults were crossed in several cages to produce F1 offspring that were layed as eggs into agar domes.
Eggs were transplanted into snowberry and apple fruits, then reared to the third larval instar.
Feeding 3rd instar larvae were snap frozen and RNA-extracted.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
larvae were homogenized in tri-reagent and total RNA was extracted from each individual using the Ambion RiboPure® Kit
|
| Label |
Cy5
|
| Label protocol |
Labeled aRNA from four individuals from each time point was generated using Quick-Amp labeling Kit (Agilent Technologies, Inc.) following the manufacturer recommendations.
We used 250ng of total RNA for reverse transcription to synthesize 1st strand cDNA, then generated labeled aRNA according to the manufacturer’s protocol
|
| |
|
| |
| Hybridization protocol |
A total of 825 ng of labeled aRNA per sample was used for the hybridization,
and hybridization and washing were performed according to the manufacturer's protocol using the Gene Expression Hybridization Kit and Gene Expression Wash Buffers (Agilent Technologies, Inc.).
We used a two-color hybridization design, with six biological (individual larva) replicates, and 2-3 technical replicates per biological replicate.
|
| Scan protocol |
The arrays were washed according to the manufacturer's protocol and then scanned on an Agilent G2505C scanner
Images were quantified using Agilent Feature Extraction Software (version 10.1.1.1).
|
| Description |
ZA-R_PA-G
|
| Data processing |
Data were background corrected and loess normalized using LIMMA
Processed data file (linked as supplementary file on Series record) reports Limma-generated log2 fold change values, estimated after normalization and background correction
|
| |
|
| Submission date |
Mar 17, 2015 |
| Last update date |
Oct 01, 2015 |
| Contact name |
Gregory Ragland |
| E-mail(s) |
gragland@ksu.edu
|
| Phone |
7855326139
|
| Organization name |
Kansas State University
|
| Department |
Entomology
|
| Street address |
123 West Waters Hall
|
| City |
Manhattan |
| State/province |
KS |
| ZIP/Postal code |
66506 |
| Country |
USA |
| |
|
| Platform ID |
GPL19895 |
| Series (1) |
| GSE66975 |
Gene expression in Rhagoletis pomonella and R. zephyria transplanted into snowberry and apple fruits |
|
