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MB Sample ID: SA341068
Local Sample ID: | TIF2 |
Subject ID: | SU003271 |
Subject Type: | Mammal |
Subject Species: | Mus musculus |
Taxonomy ID: | 10090 |
Genotype Strain: | C57BL/6J |
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Sample Preparation:
Sampleprep ID: | SP003278 |
Sampleprep Summary: | Standard curves for Glucose (Sigma-Aldrich, G7021), UDP (Sigma-Aldrich, 94330), UTP (Jena Bioscience, NU-1024S), cytidine (Sigma-Aldrich, C4654), uridine (Sigma-Aldrich, U3003) and glutamine (Gibco, 25030-34) were used to calculate the concentration of these metabolites in the samples. The standard curve was prepared with a dilution series starting from 5 mM Glucose and Glutamine, 1 mM Cytidine, Uridine, UDP and UTP. Specific dilutions are indicated in the Study Design table. Metabolites and calibration curves were extracted simultaneously by addition of 800 μl of MS-grade methanol-water buffer (MeOH:H2O, 5:3, v/v) containing the internal standards glutaric acid (5 μg ml-1, Sigma-Aldrich, G3407) and 13C6-Glucose (30 μg ml-1, Cambridge Isotope Laboratories, Inc., CLM-1396), followed by 500 μl of chloroform. Samples were then vortexed and centrifuged (4 °C for 10 min each). The polar (upper) phases were collected, divided into two equal parts for gas chromatography (GC) and liquid chromatography (LC) mass spectrometry analysis, and dried using a vacuum concentrator. The dried metabolite extracts were stored at -80 °C until analysis. |