|
| Status |
Public on Feb 12, 2008 |
| Title |
45GRa_Blood_GlobinClear_HRT |
| Sample type |
RNA |
| |
|
| Source name |
blood samples in PAXgene, globin reduction using GlobinClear kit from ambion, HRT user
|
| Organism |
Homo sapiens |
| Characteristics |
postmenopausal women
|
| Extracted molecule |
total RNA |
| Extraction protocol |
PAXgene Blood RNA Kit (PreAnalytiX, Hombrechtikon, Switzerland)
|
| Label |
digoxigenin
|
| Label protocol |
NanoAmp™ RT-IVT Labeling Kit from Applied Biosystems
|
| |
|
| Hybridization protocol |
Applied Biosystems Chemiluminescence Detection Kit
|
| Scan protocol |
Applied Biosystems Chemiluminescence Detection Kit
|
| Description |
Briefly, 10 μg of labeled cRNA targets from each sample were first fragmented, mixed with internal control target (24-mer oligo labeled with LIZ® fluorescent dye) and hybridized to a pre-hybridized microarray at 55°C for 16 hr. After washing to remove unhybridized DIG-labeled molecules, an alkaline phosphatase-antibody conjugate was added to bind to the DIG-labeled target. The addition of substrate and a chemiluminescence enhancer initiates the chemiluminescent reaction. Eight images were collected for each microarray using the 1700 analyzer including 2 “short” chemiluminescent images (5 seconds exposure length each) and 2 “long” chemiluminescent images (25 seconds exposure length each) for gene expression analysis, 2 fluorescent images for feature finding and spot normalization and 2 QC images for spectrum cross-talk correction. Applied Biosystems Expression System software was used to extract signal intensities, signal to noise ratios (S/N) and flagging values from the microarray images.
|
| Data processing |
Using Applera package in R, we set the filtering criteria so that each gene had flagging value < 2 and a signal to noise ratio (S/N) ≥ 3 in at least 50% of the samples. After filtration, we proceed with quantile normalization of the log-intensities.
|
| |
|
| Submission date |
Feb 08, 2007 |
| Last update date |
Feb 12, 2008 |
| Contact name |
Vanessa Dumeaux |
| E-mail(s) |
vanessad@rr-research.no
|
| Organization name |
"The Norwegian Women and Cancer project"
|
| Street address |
University of Tromsoe
|
| City |
Tromsoe |
| ZIP/Postal code |
9037 |
| Country |
Norway |
| |
|
| Platform ID |
GPL2986 |
| Series (1) |
| GSE7008 |
Comparison of globin RNA processing methods for genome-wide transcriptome analysis from whole-blood |
|
