|
| Status |
Public on Feb 01, 2014 |
| Title |
Subordinate male urine stimulation, OB brain tissue, replicate 1 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
OB Brain, SUB stimuli
|
| Organism |
Oreochromis mossambicus |
| Characteristics |
brain tissue: OB
olfactory stimuli: subordinate male urine
|
| Treatment protocol |
Dominant males were chemically stimulated with social relevant stimuli while electro-olfactograms measured their electrical activity at the level of the olfactory rosette. After stimulation the fish were killed, their brains dissected and brain samples were microdissected for analysis.
|
| Growth protocol |
Mixed-sex groups which were kept in tanks with gravel substrate, which promotes nest digging by males and the establishment of territories and social hierarchies, at a temperature of 26 ± 2◦C and a 12L:12D photoperiod. Fish were fed twice daily with commercial cichlid sticks.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA extracted using Trizol following manufacturer's instructions
|
| Label |
Cy5
|
| Label protocol |
mRNA (500 ng) from each experimental sample or reference were reverse transcribed using SuperScript II (Invitrogen) and labeled. Following this reverse transcription, RNA was hydrolyzed and purified before being dye-coupled with Cy3 or Cy5 post-labeling Reactive Dye Pack (Amersham).
|
| |
|
| Channel 2 |
| Source name |
OB Brain, blank stimuli
|
| Organism |
Oreochromis mossambicus |
| Characteristics |
brain tissue: OB
olfactory stimuli: none
|
| Treatment protocol |
Dominant males were chemically stimulated with social relevant stimuli while electro-olfactograms measured their electrical activity at the level of the olfactory rosette. After stimulation the fish were killed, their brains dissected and brain samples were microdissected for analysis.
|
| Growth protocol |
Mixed-sex groups which were kept in tanks with gravel substrate, which promotes nest digging by males and the establishment of territories and social hierarchies, at a temperature of 26 ± 2◦C and a 12L:12D photoperiod. Fish were fed twice daily with commercial cichlid sticks.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA extracted using Trizol following manufacturer's instructions
|
| Label |
Cy3
|
| Label protocol |
mRNA (500 ng) from each experimental sample or reference were reverse transcribed using SuperScript II (Invitrogen) and labeled. Following this reverse transcription, RNA was hydrolyzed and purified before being dye-coupled with Cy3 or Cy5 post-labeling Reactive Dye Pack (Amersham).
|
| |
|
| |
| Hybridization protocol |
A reference and experimental sample were competitively hybridized at 65oC overnight to a 19K A. burtoni cDNA microarray (GEO platform GPL6416) constructed from brain-specific and mixed tissue libraries representing a total of 17,712 cichlid-specific features
|
| Scan protocol |
Microarrays were scanned with an Axon 4000B scanner (Axon Instruments) using Genepix 4.0 software (Axon Instruments).
|
| Description |
Samples from blank stimulations (control) collected from eight different individuals (for both areas) were pooled and aliquoted to be used as reference in a reference based array design
|
| Data processing |
Microarray data were processed using the LIMMA software package (v3.12.0; [62] in R (v2.15.0; the R Foundation for Statistical Computing, 2012). Background-subtracted mean intensities were calculated using the minimum method and further normalized using within-array loess normalization. After this normalization step, Bayesian analysis was used to calculate gene expression levels using the ratios of intensities measured. Finally, to compare between expression profiles for the different olfactory stimulations, unsupervised hierarchical clustering analysis were done using the hclust function in R/Bioconductor.
|
| |
|
| Submission date |
Jan 28, 2014 |
| Last update date |
Feb 01, 2014 |
| Contact name |
José Miguel Simões |
| Organization name |
ISPA - Instituto Universitário
|
| Department |
Eco-Ethology Research Unit
|
| Lab |
Integrative Behavioural Biology Group
|
| Street address |
Rua Jardim do Tabaco 34
|
| City |
Lisboa |
| ZIP/Postal code |
1140-041 |
| Country |
Portugal |
| |
|
| Platform ID |
GPL6416 |
| Series (1) |
| GSE54468 |
Social odors conveying dominance and reproductive information induce rapid physiological and neuromolecular changes in a cichlid fish. |
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