|
| Status |
Public on Jan 23, 2015 |
| Title |
RP_66 |
| Sample type |
RNA |
| |
|
| Source name |
Breast tissue samples from breast reduction operations
|
| Organism |
Homo sapiens |
| Characteristics |
diagnosis: normal patient
sample: RP_66
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Trizol extraction of total RNA was performed according to the manufacturer's instructions.
|
| Label |
Cy3
|
| Label protocol |
1µg RNA were analyzed on nONCOchip 1.0 array according to the manufacturer's protocol (Agilent One-Color Quick Amp Labeling kit protocol) with the adaptation of using an N6 − T 7 primer instead of a polyT-T7 primer.
|
| |
|
| Hybridization protocol |
nONCOchip 1.0 arrays were proceed using the Agilent Hybridization Kit, Agilent Hybrization-Chambers and an commercial Hyb-Oven according to the manufacturer's protocol (Agilent One-Color Quick Amp Labeling Kit).
|
| Scan protocol |
Arrays were scanned using a GenePix 4200 Scanner and the GenePix Pro 6.1 Software (Molecular Devices) with following settings: laser power: 2.32-2.43, PMT: 400, resolution: 5µm, Focus: 1µm, Lines to Average: 2, Wavelength: 532nm, Filter: Standard Green.
|
| Description |
Expression analysis of 5 normal patient samples. Breast tissue samples from breast reduction operations were provided from the Colosseum Clinic, Oslo in co-operation with Akershus University Hospital, Lørenskog and are referred to as normal tissue. Expression was assessed by using an Agilent custom microarray (244K, nONCOchip). The custom array contains probes for genomic regions that have been found to be differentially expressed (i) throughout cell cycle progression, (ii) in response to the anti-proliferative and pro-apoptotic p53 pathway, and (iii) the anti-apoptotic and pro-proliferative STAT-3 pathway by employing TAS (Kampa et al. "Novel RNAs identified from an in-depth analysis of the transcriptome of human chromosomes 21 and 22". Genome Research, 14:331-42, 2004). In addition, the Agilent custom array interrogates probes for genomic regions predicted to contain a conserved secondary structure identified by RNAz (Washietl et al. "Fast and reliable prediction of noncoding RNAs." Proc Natl Acad Sci USA. 102:2454-9, 2005.) or Evofold (Pedersen et al. "Identification and classification of conserved RNA secondary structures in the human genome." PLoS Comput Biol. 2:e33, 2006.), as well as known non-coding RNAs from public databases, and the Agilent mRNA probe set 014850.
|
| Data processing |
The data were analyzed with R (version 2.14.2) and BioConductor. Expression levels were quantile normalized and a linear model was fitted using the R package limma in order to identify differentially expressed probes. Reliable variance estimations were obtained by empirical Bayes moderated t-statistics. False discovery rate was controlled by Benjamini-Hochberg adjustment.
|
| |
|
| Submission date |
Aug 29, 2013 |
| Last update date |
Jan 23, 2015 |
| Contact name |
Kristin Reiche |
| E-mail(s) |
kristin.reiche@izi.fraunhofer.de
|
| Organization name |
Fraunhofer Institute for Cell Therapy and Immunology
|
| Department |
Diagnostics
|
| Street address |
Perlickstr. 1
|
| City |
Leipzig |
| ZIP/Postal code |
04103 |
| Country |
Germany |
| |
|
| Platform ID |
GPL13648 |
| Series (1) |
| GSE50428 |
Expression and differential expression analysis of breast cancer patient samples and normal samples |
|
