|
| Status |
Public on May 26, 2016 |
| Title |
ESC |
| Sample type |
RNA |
| |
|
| Source name |
B6J-23 ESC
|
| Organism |
Mus musculus |
| Characteristics |
strain: C57BL/6J
cell type: Embryonic stem cells (ESC)
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was prepared using the Trizol and RNeasy Kit following the manufacturer's manuals. RNA was quantified using a NanoDrop-1000 Spectrophotometer.
|
| Label |
Cy3
|
| Label protocol |
Cyanine-3 (Cy3) labeled cRNA was prepared from 100ng RNA using the Low Input Quick Amp Labeling kit,One-color (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 |
600ng of Cy3-labelled cRNA (specific activity >6.0 pmol Cy3/ug cRNA) was fragmented at 60°C for 30 minutes in a reaction volume of 25ul containing 25x fragmentation and 10x blocking agent following the manufacturers instructions. On completion of the fragmentation reaction,25ul of 2x GE Hybridization Buffer HI-RPM was added to the fragmentation mixture and hybridized to Agilent SurePrint G3 Mouse Gene Expression 8x60K Microarrays (G4852A) 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) and 1 minute with 37°C GE Wash buffer 2 (Agilent).
|
| Scan protocol |
Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner (G2505CA) using one color scan setting for 8x60k array slides (Scan Area 61x21.6 mm, Scan resolution 3um, Dye channel is set to Green. No XDR).
|
| Description |
SAMPLE 1
|
| Data processing |
The scanned images were analyzed with Feature Extraction Software 11.5.1.1 (Agilent) using default parameters (protocol:GE1_1105_Oct12 and Grid: 028005_D_F_20131202) to obtain background subtracted and spatially detrended Processed Signal intensities. Features flagged in Feature Extraction as Feature Non-uniform outliers were excluded.
|
| |
|
| Submission date |
Mar 23, 2016 |
| Last update date |
May 26, 2016 |
| Contact name |
Akira Kunitomi |
| E-mail(s) |
akira.kunitomi@gladstone.ucsf.edu
|
| Organization name |
The J. David Gladstone Institutes
|
| Street address |
1650 Owens Street
|
| City |
San Francisco |
| State/province |
CA |
| ZIP/Postal code |
94158 |
| Country |
USA |
| |
|
| Platform ID |
GPL10787 |
| Series (1) |
| GSE79515 |
Gene expression profiles of OSKH iPSCs, OSK iPSCs and ESC |
|
