|
| Status |
Public on Dec 31, 2014 |
| Title |
KKAyLiver_4week_rep1 |
| Sample type |
RNA |
| |
|
| Source name |
KKAy liver, 4 weeks
|
| Organism |
Mus musculus |
| Characteristics |
strain: KKAy
age: 4 weeks
tissue: liver
disease state: diabetes
|
| Growth protocol |
All mice used in this study were maintained on a 12-hour light-dark cycle and fed a standard rodent chow. Mice were fasted overnight for 13.5 hours.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was prepared using the RNeasy Mini Kit (QIAGEN Inc.) following the manufacturer's recommendations. RNA was quantified using a NanoDrop-1000 spectrophotometer and quality was monitored with the Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA).
|
| Label |
Cy3
|
| Label protocol |
Cyanine-3 (Cy3) labeled cRNA was prepared from 0.5 ug RNA using the Quick Amp Labeling kit, One-Color (Agilent) according to the manufacturer's instructions, followed by RNeasy column purification (QIAGEN, Valencia, CA). Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.
|
| |
|
| Hybridization protocol |
1.65 μg of Cy3-labelled cRNA (specific activity >10.0 pmol Cy3/ug cRNA) was fragmented at 60°C for 30 minutes in a reaction volume of 55 μl containing 10x Blocking Agent and 25x Fragmentation Buffer following the manufacturer's instructions. On completion of the fragmentation reaction, 55 μl of 2x GE Hybridization buffer was added to the fragmentation mixture and hybridized to Agilent Whole Mouse Genome Oligo Microarrays (G4122F) 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 (G2505B) using one color scan setting for 4x44k array slides (Scan Area 61x21.6 mm, Scan resolution 5 μm, Dye channel is set to Green and Green PMT is set to both XDR Hi 100% and XDR Lo 10%).
|
| Description |
4w KKAy liver_1
|
| Data processing |
The scanned images were analyzed with Feature Extraction Software 9.5.1.1 (Agilent) using default parameters (protocol GE1-v5_95_Feb07 and Grid: 014868_D_20070820) to obtain background subtracted and spatially detrended Processed Signal intensities. Features flagged in Feature Extraction as Feature Non-uniform outliers were excluded. The 75th percentile, baseline to median of all samples was used as the normalization method (GeneSpring).
|
| |
|
| Submission date |
Mar 10, 2012 |
| Last update date |
Dec 31, 2014 |
| Contact name |
Hayashi Yasuhiro |
| Organization name |
Hokkaido University
|
| Department |
Faculty of Pharmaceutical Sciences
|
| Street address |
Kita-12, Nishi-6, Kita-ku
|
| City |
Sapporo |
| ZIP/Postal code |
060-0812 |
| Country |
Japan |
| |
|
| Platform ID |
GPL7202 |
| Series (1) |
| GSE36412 |
Liver gene expression signatures for type 2 diabetic mice and normal mice |
|
