|
| Status |
Public on May 06, 2009 |
| Title |
Dog Normal mammary gland 5 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
Normal mammary gland
|
| Organism |
Canis lupus familiaris |
| Characteristics |
disease state: normal tissue
tissue: mammary gland
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated with RNAzolB and finally dissolved in RNase-free water. Then 25μg of total RNA was treated with DNase using the Qiagen RNase-free DNase kit and RNeasy spin columns. Total RNA was dissolved in RNase-free water to a final concentration of 0.2 μg/μl.
|
| Label |
Cy5
|
| Label protocol |
cRNA was generated by in vitro transcription using T7 RNA polymerase on 5 μg of total RNA and labelled with Cy5 or Cy3 (Cy Dye, Amersham Pharmacia Biotech).
|
| |
|
| Channel 2 |
| Source name |
Normal reference pool
|
| Organism |
Canis lupus familiaris |
| Characteristics |
sample: Pool of normal samples
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated with RNAzolB and finally dissolved in RNase-free water. Then 25μg of total RNA was treated with DNase using the Qiagen RNase-free DNase kit and RNeasy spin columns. Total RNA was dissolved in RNase-free water to a final concentration of 0.2 μg/μl.
|
| Label |
Cy3
|
| Label protocol |
cRNA was generated by in vitro transcription using T7 RNA polymerase on 5 μg of total RNA and labelled with Cy5 or Cy3 (Cy Dye, Amersham Pharmacia Biotech).
|
| |
|
| |
| Hybridization protocol |
5 μg of labelled-RNA from each sample were co-hybridized with 5 μg of a normal reference pool, consisting of an equal amount of cRNA extracted from mammary healthy samples. Labelled cRNAs were fragmented to an average size of 50-100 nucleotides by heating the samples to 60°C with 10mM of zinc chloride and then adding an hybridization buffer containing 1M NaCl, 0.5% sodium sarcosine, 50mM MES, pH6.5, and formamide to a final concentration of 30%. The final volume was 3ml at 40°C.
|
| Scan protocol |
Slides were washed and scanned using a confocal laser scanner (Agilent Technologies). Images were quantified using Agilent Feature Extraction Software (version 9.1).
|
| Description |
Normal mammary gland
|
| Data processing |
Lowess normalized, background subtracted data obtained from log10 of processed Red signal/processed Green signal. Rosetta Resolver gene expression analysis software (version 3.2, Rosetta Biosoftware, Seattle, WA) was used.
|
| |
|
| Submission date |
Feb 25, 2009 |
| Last update date |
May 05, 2010 |
| Contact name |
Paolo Uva |
| Organization name |
IRCCS Istituto Giannina Gaslini
|
| Lab |
Clinical Bioinformatics
|
| Street address |
Via Gerolamo Gaslini, 5
|
| City |
Genoa |
| ZIP/Postal code |
16147 |
| Country |
Italy |
| |
|
| Platform ID |
GPL7198 |
| Series (1) |
| GSE14999 |
Gene expression profiling of human and dog breast cancers |
|
