|
| Status |
Public on Dec 31, 2016 |
| Title |
Shield Rep 5 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
MTX2-MO shield 13_11_05
|
| Organism |
Danio rerio |
| Characteristics |
cell type: Shield stage embryos
|
| Treatment protocol |
30-50 embryos were collected from clutches at three different time points: 30% epiboly, shield and tailbud stage
|
| Growth protocol |
Outbred wild type zebrafish stocks were reared and embryos obtained. Embryos were incubated at 28.5°C
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Zebrafish embryos were homogenized in RLT buffer (Qiagen) using a 21 gauge needle and 1ml syringe. Total RNA extracted by RNeasy column purification (QIAGEN) was assessed for quantity and integrity on the Bioanalyzer 2100 (Agilent Technologies).
|
| Label |
Cy5
|
| Label protocol |
Total RNA from each sample was linearly amplified from 800ng of total RNA using the Amino Allyl MessageAmp amplification kit (Ambion Inc/GeneWorks), yielding a minimum of 20µg amino-allyl-labeled antisense aRNA per reaction. The quantity and size distribution of the aRNA products was assessed using the Agilent Bioanalyzer 2100. Five micrograms of each aRNA sample was fluorescently labeled by covalent coupling to either Cy5- or Cy3-NHS ester dyes (Amersham).
|
| |
|
| Channel 2 |
| Source name |
STD-MO shield 13_11_05
|
| Organism |
Danio rerio |
| Characteristics |
cell type: Shield stage embryos
|
| Treatment protocol |
30-50 embryos were collected from clutches at three different time points: 30% epiboly, shield and tailbud stage
|
| Growth protocol |
Outbred wild type zebrafish stocks were reared and embryos obtained. Embryos were incubated at 28.5°C
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Zebrafish embryos were homogenized in RLT buffer (Qiagen) using a 21 gauge needle and 1ml syringe. Total RNA extracted by RNeasy column purification (QIAGEN) was assessed for quantity and integrity on the Bioanalyzer 2100 (Agilent Technologies).
|
| Label |
Cy3
|
| Label protocol |
Total RNA from each sample was linearly amplified from 800ng of total RNA using the Amino Allyl MessageAmp amplification kit (Ambion Inc/GeneWorks), yielding a minimum of 20µg amino-allyl-labeled antisense aRNA per reaction. The quantity and size distribution of the aRNA products was assessed using the Agilent Bioanalyzer 2100. Five micrograms of each aRNA sample was fluorescently labeled by covalent coupling to either Cy5- or Cy3-NHS ester dyes (Amersham).
|
| |
|
| |
| Hybridization protocol |
The labeled material was hydrolyzed and used for microarray hybridization with hybridization at 42ºC
|
| Scan protocol |
Hybridized microarrays were scanned on a 600B Scanner (Agilent Technologies Inc.).
|
| Description |
cGENz000486
|
| Data processing |
The images were analyzed using Imagene 5.6.0 (BioDiscovery) to determine mean foreground and background signal intensities for both channels. All primary data, including images, were then imported into the comprehensive microarray relational database, Bio-Array Software Environment (BASE). In the case of all direct comparisons, the raw data for each hybridization was compiled into an experiment and subjected to print tip Loess normalization using R functions from Bioconductor (Linear models for Microarray Data, LIMMA) and Statistics for Microarray Analysis (SMA).
|
| |
|
| Submission date |
Oct 07, 2015 |
| Last update date |
Dec 31, 2016 |
| Contact name |
Charles Cameron Bell |
| E-mail(s) |
charles.bell@mater.uq.edu.au
|
| Phone |
0421483872
|
| Organization name |
Mater Research
|
| Lab |
Cancer Genomics
|
| Street address |
37 Kent Street
|
| City |
Brisbane |
| State/province |
Queensland |
| ZIP/Postal code |
4169 |
| Country |
Australia |
| |
|
| Platform ID |
GPL6590 |
| Series (1) |
| GSE73812 |
Mtx2 dependent genes during Zebrafish gastrulation |
|
