|
| Status |
Public on Dec 01, 2015 |
| Title |
Ucell-ct-2 |
| Sample type |
RNA |
| |
|
| Source name |
Urine sample
|
| Organism |
Homo sapiens |
| Characteristics |
gender: Female
individual: Healthy donor
cell type: Progenitor cells isolated from urines samples
|
| Growth protocol |
Gene expression in atmospheric O2 and 5%CO2 culture condition
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was prepared using the RNeasy Mini Kit (Qiagen) following the manufacturer's recommendations. RNA was quantified using a NanoDrop ND-1000 spectrophotometer and quality was monitored with the Agilent 2100 Bioanalyzer (Agilent).
|
| Label |
Cy5
|
| Label protocol |
Cyanine-3 (Cy3) labeled cRNA and Cyanine-5 (Cy5) labeled cRNA were prepared from 0.1 ug RNA using the Low Input QuickAmp Labeling Kit (Agilent) according to the manufacturer's instructions, followed by RNAeasy column purification (Qiagen). Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.
|
| |
|
| Hybridization protocol |
0.3 ug of Cy3-labelled cRNA (specific activity > 8 pmol Cy3/ug cRNA) pooled with 0.3 µg of Cy5-labelled cRNA (specific activity > 8 pmol Cy5/ug cRNA) were fragmented at 60°C for 30 minutes in a reaction volume of 25 µl containing 1x Agilent fragmentation buffer and 2x Agilent blocking agent following the manufacturers instructions. On completion of the fragmentation reaction, 25 µl of 2x Agilent GEx hybridization buffer Hi-RPM was added to the fragmentation mixture and hybridized to Agilent Sure Print G3 Human 8x 60K Microarrays (G4851A) for 17 hours at 65°C in a rotating Agilent hybridization oven. After hybridization, microarrays were washed 1 minute at room temperature with GE Wash Buffer 1 (Agilent), 1 minute with 37°C GE Wash buffer 2 (Agilent), 10 seconds with Acetonitrile anhydrous 99.8% at room temperature and 1 minute with Stabilization and Drying solution (Agilent) at room temperature. Then, Microarrays were dried immediately.
|
| Scan protocol |
Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner (G2505C) using two colors scan setting for 8x60k array slides (Scan Area HD 61x21.6 mm, Scan resolution 3 µm, Dye channels are set to Green and Red) and PMT is setup on autoscale.
|
| Description |
US82400123_252800418270_S01_GE2_107_Sep09_1_3.txt
|
| Data processing |
The scanned images were analyzed with Feature Extraction Software 10.7.1.1 (Agilent) using default parameters (protocol GE2_107_Sep09 and Grid: 028004_D_F_20110819) to obtain background subtracted and spatially detrended Processed Signal intensities. Features flagged in Feature Extraction as Feature Non-uniform outliers were excluded.
|
| |
|
| Submission date |
Dec 01, 2015 |
| Last update date |
Dec 01, 2015 |
| Contact name |
Karim Si-Tayeb |
| E-mail(s) |
karim.si-tayeb@univ-nantes.fr
|
| Phone |
+33 2 2808 0176
|
| Organization name |
l'institut du thorax, UMR INSERM1087 / CNRS6291, IRS-UN
|
| Street address |
8,quai Moncousu
|
| City |
Nantes |
| ZIP/Postal code |
44007 |
| Country |
France |
| |
|
| Platform ID |
GPL13607 |
| Series (1) |
| GSE75545 |
Urine-sample-derived human induced pluripotent stem cells as a model to study PCSK9-mediated autosomal dominant hypercholesterolemia |
|
