|
| Status |
Public on Sep 27, 2013 |
| Title |
Blood_Athlete_3_time-point T2 |
| Sample type |
RNA |
| |
|
| Source name |
Whole blood, Athlete 3, immediately after exericse
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: Whole blood
gender: Male
age: 60
subject: athlete
time point: immediately after exercise
|
| Treatment protocol |
Whole blood samples for each participant were stored in PAXgene blood RNA tubes (Qiagen GmbH, Germany) at -80 °C from all three time-points until further analysis.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was isolated from blood sample aliquots (2.5 ml) using a PAXgene blood RNA kit (Qiagen GmbH, Germany) according to the manufacturer’s protocol. Residual DNA was removed with DNase I treatment, according to the manufacturer’s instructions (Qiagen GmbH, Hombrechtikon, Germany). The quality of the isolated RNA was checked with Agilent RNA 6000 Nano Assay kit reagents using an Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA, USA). Samples with a RNA Integrity Number (RIN) of 7.0 or higher were used for further analysis. The RNA samples were quantified using a standard Nanodrop ND-1000 spectrophotometer (Nanodrop, Wilmington, DE, USA).
|
| Label |
Cy3
|
| Label protocol |
Total RNA (200 ng) from each sample was spiked with One-Color RNA Spike-In Kit reagents (Agilent Technologies, Santa Clara, CA, USA) according to the manufacturer’s instructions.Subsequent amplification and labeling of the samples to generate fluorescent cRNA (complimentary RNA) were carried out using the One-Color Quick Amp Labeling Kit (Agilent Technologies, Santa Clara, CA, USA) according to the manufacturer’s instructions.Labeled cRNA was purified using an RNeasy Mini kit (Qiagen). Only cRNA samples with a yield >1.65 µg, and a specific activity >9 pmol Cy3/µg cRNA were used for subsequent hybridization onto microarray slides.
|
| |
|
| Hybridization protocol |
Labeled RNA was hybridized to Agilent whole genome expression array (4x44K) at 65°C for 17h in rotisserie in a hybridization oven. Disassembled arrays were washed in GE wash buffer 1&2 for 1 minutes each.
|
| Scan protocol |
Images were scanned with Agilent scanner at 5µm resolution.
|
| Description |
Gene expression in blood of Athlete 3 immediately after exercise
|
| Data processing |
Raw data obtained from the microarrays were imported into Nexus Expression 3.0 (BioDiscovery Inc, Hawthorne, CA, USA) and processed according to the default settings of the software, which included a local background correction, removal of flagged spots and performing quantile normalization.
|
| |
|
| Submission date |
Sep 26, 2013 |
| Last update date |
Sep 27, 2013 |
| Contact name |
Colin D. Funk |
| E-mail(s) |
funkc@queensu.ca
|
| Organization name |
Queen's University
|
| Department |
Biomedical and Molecular Sciences
|
| Lab |
Room # 413, Botterell Hall
|
| Street address |
18 Stuart Street
|
| City |
Kingston |
| State/province |
Ontario |
| ZIP/Postal code |
K7L 5E3 |
| Country |
Canada |
| |
|
| Platform ID |
GPL6480 |
| Series (1) |
| GSE51216 |
Whole blood transcriptomics to define adaptive biochemical pathways of exercise during aging |
|
