|
| Status |
Public on Sep 10, 2016 |
| Title |
embryo_WT_d3 |
| Sample type |
RNA |
| |
|
| Source name |
OR
|
| Organism |
Drosophila melanogaster |
| Characteristics |
tissue: embryo
embryonic stage: 11-15
genotype: Oregon R wild-type
|
| Treatment protocol |
Embryos were dechorionated in 50% bleach for three minutes and then thoroughly washed with ddH2O.
|
| Growth protocol |
Embryos were collected from OregonR (OR) or foxL1 null (foxL1del/foxL1cr) flies for eight hours at room temperature and then aged for 11 additional hours at room temperature.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
300ul of Trizol was added to embryos on ice. Samples were homogenized with pestles by hand for 30-60seconds. 200ul of Trizol was then added and sampes were centrifuged at 13000rpm for 10minutes at 4C. The supernatent was tranferred to a new tube and 150ul of chloroform was added. Tubes were vortexed for 15seconds and then incubated at room temperature for 2-3 minutes. Samples were then centrifuged at 13000rpm for 15min at 4C. The upper phase was placed in a new tube and 0.7 volues of isopropanol was added. Samples were incubated at room temperature for 5 minutes and then centrifuged at 13000rpm for 15min at 4C. The pellet was then washed with 70% EtOH/DEPC H2O and centrifuged at 13000rpm for 10min at 4C. The pellet was then dried on a heat block and resuspended in 100ul RNAsefree H2O. An RNeasy kit was then used to further purify the RNA. RNA concentrations were around 2ug/ul. Approximately 200ng was used per experiment.
|
| Label |
biotin
|
| Label protocol |
Biotinylated cRNAs were prepared with the GeneChip HT 3' IVT Express Kit per manufacturer's instructions.
|
| |
|
| Hybridization protocol |
Following fragmentation cRNAs were hybridized 16hes at 45° C with rotation (60rpm) as described by Affymetrix in their GeneChip Expression Analysis Technical Manual.
|
| Scan protocol |
Using Affymetrix' GeneChip Scanner 3000 7G and default parameters described by the manufacturer.
|
| Description |
WT = wildtype embryo (OregonR = OR)
|
| Data processing |
Affymetrix CEL files were extracted and their data normalized with the Partek GS 6.6 platform. RMA normalization was used to create quantile-normalized log2 transcript signal values, which were used in subsequent ANOVA analyses.
|
| |
|
| Submission date |
Sep 09, 2016 |
| Last update date |
Sep 10, 2016 |
| Contact name |
Caitlin D Hanlon |
| E-mail(s) |
chanlon3@jhmi.edu
|
| Organization name |
JHMI
|
| Department |
Cell Biology
|
| Lab |
Deborah J Andrew
|
| Street address |
725 N Wolfe St, Hunterian G10
|
| City |
Baltimore |
| State/province |
MARYLAND |
| ZIP/Postal code |
21205 |
| Country |
USA |
| |
|
| Platform ID |
GPL1322 |
| Series (1) |
| GSE86626 |
Expression data from wildtype and foxL1null Drosophila embryos |
|
