|
| Status |
Public on Feb 01, 2013 |
| Title |
larvalCNS-chr2stim-rep1 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
larvalCNS, ChR2, stimulated
|
| Organism |
Drosophila melanogaster |
| Characteristics |
treatment: synaptic stimulation
genotype: UAS-ChR2 x C380-Gal4
developmental stage: third instar larvae
tissue: Ventral ganglia + optic lobes (- eye imaginal discs)
|
| Treatment protocol |
The chr2-stim group was subjected to 5x spaced light stimulation. The chr2-un group was subjected to 0x mock stimulation.
|
| Growth protocol |
The white genotype was grown on standard Bloomington media at 25°C. ChR2-expressing genotypes were grown on standard Bloomington food supplemented with 100mM all-trans retinal at 25°C.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using the miRCURY RNA Isolation kit (Exiqon) yielding ~125 ng of total RNA per CNS. RNA was eluted with 40 ul of RNAse free water (Ambion), flash frozen, and stored at -80°C.
|
| Label |
Hy3
|
| Label protocol |
Total RNA from sample and reference was labelled with Hy3 and Hy5 fluorescent label, respectively, using the miRCURY™ LNA Array power labelling kit (Exiqon, Denmark) following the procedure described by the manufacturer.
|
| |
|
| Channel 2 |
| Source name |
common reference pool
|
| Organism |
Drosophila melanogaster |
| Characteristics |
developmental stage: third instar larvae
tissue: Ventral ganglia + optic lobes (- eye imaginal discs)
treatment: common reference pool
genotype: common reference pool
|
| Treatment protocol |
The chr2-stim group was subjected to 5x spaced light stimulation. The chr2-un group was subjected to 0x mock stimulation.
|
| Growth protocol |
The white genotype was grown on standard Bloomington media at 25°C. ChR2-expressing genotypes were grown on standard Bloomington food supplemented with 100mM all-trans retinal at 25°C.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using the miRCURY RNA Isolation kit (Exiqon) yielding ~125 ng of total RNA per CNS. RNA was eluted with 40 ul of RNAse free water (Ambion), flash frozen, and stored at -80°C.
|
| Label |
Hy5
|
| Label protocol |
Total RNA from sample and reference was labelled with Hy3 and Hy5 fluorescent label, respectively, using the miRCURY™ LNA Array power labelling kit (Exiqon, Denmark) following the procedure described by the manufacturer.
|
| |
|
| |
| Hybridization protocol |
The Hy3-labeled samples and a Hy5-labeled reference RNA sample were mixed pair-wise and hybridized to the miRCURY LNA array version 11.0 Other Species (Exiqon, Denmark), which contains capture probes targeting all miRNAs for all other species than human, mouse or rat registered in the miRBASE version 14.0 at the Sanger Institute. The hybridization was performed according to the miRCURY™ LNA array manual using a Tecan HS4800 hybridization station (Tecan, Austria). After hybridization the microarray slides were scanned and stored in an ozone free environment (ozone level below 2.0 ppb) in order to prevent potential bleaching of the fluorescent dyes.
|
| Scan protocol |
The miRCURY™ LNA array microarray slides were scanned using the Agilent G2565BA Microarray Scanner System (Agilent Technologies, Inc., USA) and the image analysis was carried out using the ImaGene 8.0 software (BioDiscovery, Inc., USA).
|
| Description |
raw files 0_ are Hy3 and1_ are Hy5
|
| Data processing |
The quantified signals were background corrected (Normexp with offset value 10 – Ritchie et al., 2007) and normalized using the global Lowess (LOcally WEighted Scatterplot Smoothing) regression algorithm.
|
| |
|
| Submission date |
Jan 31, 2013 |
| Last update date |
Feb 01, 2013 |
| Contact name |
Scott A Barbee |
| E-mail(s) |
scott.barbee@du.edu
|
| Phone |
303-871-3582
|
| Organization name |
University of Denver
|
| Department |
Biological Sciences
|
| Street address |
2190 E. ILIFF Ave.
|
| City |
Denver |
| State/province |
CO |
| ZIP/Postal code |
80208 |
| Country |
USA |
| |
|
| Platform ID |
GPL16576 |
| Series (2) |
| GSE43943 |
The miRNA pathway controls rapid changes in activity-dependent synaptic structure at the Drosophila melanogaster neuromuscular junction (miRNA array) |
| GSE43945 |
The miRNA pathway controls rapid changes in activity-dependent synaptic structure at the Drosophila melanogaster neuromuscular junction |
|
