Preparation of Intracellular Metabolite Extracts from Liquid Schizosaccharomyces pombe Cultures | Zendy

Tomáš Pluskal | Zendy; Takahiro Nakamura | Zendy; Mitsuhiro Yanagida | Zendy

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Preparation of Intracellular Metabolite Extracts from Liquid Schizosaccharomyces pombe Cultures

Author(s) -

Tomáš Pluskal,

Takahiro Nakamura,

Mitsuhiro Yanagida

Publication year - 2016

Publication title -

cold spring harbor protocols

Language(s) - English

Resource type - Journals

SCImago Journal Rank - 0.674

H-Index - 51

eISSN - 1940-3402

pISSN - 1559-6095

DOI - 10.1101/pdb.prot091553

Subject(s) - schizosaccharomyces pombe , metabolite , intracellular , schizosaccharomyces , liquid culture , chemistry , liquid medium , biochemistry , microbiology and biotechnology , biology , chromatography , yeast , botany , saccharomyces cerevisiae

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.

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