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Cold Spring Harb Protoc. 2016 Dec 1;2016(12). doi: 10.1101/pdb.prot091553.

Preparation of Intracellular Metabolite Extracts from Liquid Schizosaccharomyces pombe Cultures.

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1
G0 Cell Unit, Okinawa Institute of Science and Technology Graduate University (OIST), Onna-son, Kunigami, Okinawa 904-0495, Japan.

Abstract

The success of metabolomic analysis relies heavily on the sample preparation protocol. Here we present a protocol for intracellular metabolite extraction from liquid fission yeast cultures based on rapid quenching in pure methanol at -40°C, bead-beating in 50% methanol for cell disruption, and 10 kDa cutoff ultrafiltration for removal of proteins. Samples are concentrated by vacuum evaporation and resuspended in 50% acetonitrile for mass spectrometric analysis. This protocol is optimal for extraction of polar metabolites such as amino acids, organic acids, nucleotides, sugars, or sugar-phosphates. Its implementation requires <6 h and allows preparation of multiple samples in parallel.

PMID:
27934687
DOI:
10.1101/pdb.prot091553
[Indexed for MEDLINE]

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