|
| Status |
Public on Mar 16, 2011 |
| Title |
Lung EC, experimental, rep2 |
| Sample type |
RNA |
| |
|
| Source name |
Lung endothelial cells isolated from MxCre+ Fk1 L/L Fk2 L/L Afx L/L mouse
|
| Organism |
Mus musculus |
| Characteristics |
cell type: Lung endothelial cells
genotype: MxCre+ Fk1 L/L Fk2 L/L Afx L/L
genetic background: FVBn
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was isolated using Trizol (Invitrogen) and the RNeasy mini kit (Qiagen).
|
| Label |
biotin
|
| Label protocol |
1. RNA is converted into cDNA using a T7 promoter-tailed oligo-dT primer in the synthesis of the first cDNA strand; second strand cDNA synthesis is then carried out. 2. The double-stranded cDNA is used as the template in an in vitro transcription (IVT) reaction catalyzed by T7 polymerase and containing biotinylated CTP and UTP in addition to the four unmodified ribonucleoside triphosphates. 3. The biotinylated complementary RNA (cRNA) is purified from the IVT reaction mixture using magnetic particles. The cRNA is quantified spectrophotometrically and purity of the cRNA is also assessed by spectrophometric measurements. 4. The purified cRNA is fragmented in order to facilitate the subsequent hybridization step.
|
| |
|
| Hybridization protocol |
The fragmented cRNA is added to a hybridization solution containing several biotinylated control oligonucleotides (for quality control), and hybridized to a microarray chip overnight at 45°C. The hybridization solution is then removed. The chips are transferred to a fluidics instrument that performs washes to remove cRNA that has not hybridized to its complementary oligonucleotide probe. The bound cRNA is then fluorescently labeled using phycoerythrin-conjugated streptavidin (SAPE); additional fluors are then added using biotinylated anti-streptavidin antibody and additional SAPE.
|
| Scan protocol |
Each cRNA bound at its complementary oligonucleotide is excited using a confocal laser scanner, and the positions and intensities of the fluorescent emissions are captured. These measures provide the basis of subsequent biostatistical analysis.
|
| Description |
Gene expression data from lung endothelial cells isolated from MxCre+ Fk1 L/L Fk2 L/L Afx L/L mouse
|
| Data processing |
Cel files were processed with dChip (http://biosun1.harvard.edu/complab/dchip/).
|
| |
|
| Submission date |
Mar 14, 2011 |
| Last update date |
Mar 16, 2011 |
| Contact name |
Yonghong Xiao |
| E-mail(s) |
yonghong_xiao@yahoo.com
|
| Organization name |
Dana-Farber Cancer Institute
|
| Street address |
450 Brookline Ave., HIM218A
|
| City |
Boston |
| State/province |
MA |
| ZIP/Postal code |
02115 |
| Country |
USA |
| |
|
| Platform ID |
GPL1261 |
| Series (1) |
| GSE27932 |
FoxOs are lineage-restricted redundant tumor suppressors and regulate endothelial cell homeostasis. |
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