sample: pooled rna gender: female cyclus phase: follicular time: 1h after run mean age: 29.68yrs mean body mass index: 20.9 mean training sessions per week: 4.4 mean training distance per week: 38.9 mean viat: 11.8
Treatment protocol
Probands were on no medication and exercised regularly. Female probands did not use oral contraceptives.
Extracted molecule
total RNA
Extraction protocol
EDTA anti-coagulated venous blood samples were drawn one hour before and immediately after the running exercise. Peripheral blood mononuclear cells (PBMC) were isolated using the Ficoll-hypaque density gradient technique. After gathering the cells in RLT-buffer, total RNA was extracted using an RNeasy minikit (Qiagen, Hilden Germany) in accordance with the manufacturer’s protocol. The RNA was pooled using equal amounts of RNA. The integrity of extracted RNA was assessed using an Agilent 2100 Bioanalyzer (Agilent Technologies, Palo Alto, California, USA).
Label
Cy5
Label protocol
Amplification of sample RNA was performed using Ambion´s Amino Allyl Message Amp II aRNA Amplification Kit (Ambion Inc., Austin, Texas, USA) together with Amersham CyDye Post-labeling Reactive Pack (GE Healthcare, Buckinghamshire, UK) following the manufacturer`s protocols, and assessing dye incorporation using a Nano Drop ND-1000 spectrophotometer (NanoDrop Technologies, Wilmington, Delaware, USA).
reference: UniRNA provider: Stratagene, La Jolla, California, USA
Treatment protocol
Probands were on no medication and exercised regularly. Female probands did not use oral contraceptives.
Extracted molecule
total RNA
Extraction protocol
EDTA anti-coagulated venous blood samples were drawn one hour before and immediately after the running exercise. Peripheral blood mononuclear cells (PBMC) were isolated using the Ficoll-hypaque density gradient technique. After gathering the cells in RLT-buffer, total RNA was extracted using an RNeasy minikit (Qiagen, Hilden Germany) in accordance with the manufacturer’s protocol. The RNA was pooled using equal amounts of RNA. The integrity of extracted RNA was assessed using an Agilent 2100 Bioanalyzer (Agilent Technologies, Palo Alto, California, USA).
Label
Cy3
Label protocol
Amplification of sample RNA was performed using Ambion´s Amino Allyl Message Amp II aRNA Amplification Kit (Ambion Inc., Austin, Texas, USA) together with Amersham CyDye Post-labeling Reactive Pack (GE Healthcare, Buckinghamshire, UK) following the manufacturer`s protocols, and assessing dye incorporation using a Nano Drop ND-1000 spectrophotometer (NanoDrop Technologies, Wilmington, Delaware, USA).
Hybridization protocol
Hybridization was performed for 14h at 48°C.
Scan protocol
Scanned using scanner from Affymetrix (Santa Clara, California, USA). The photomultiplier tube voltage was set to 100% for both green and red channels. The resulting green and red images were overlaid using ImaGene 5.0 (Biodiscovery Inc. El Segundo, California, USA) as well as for raw data collection.
Description
Female (follicular phase) after run vs. reference.
Data processing
Background correction: normexp (default parameters), intra-array normalization:printtip loess (default parameters), inter-array normalization: quantile normalization (default parameters), using limma package for R.
