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Methods:Fully 13C-labeled growth medium (Nakanishi et al. 2011)

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Contents

Purpose

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This medium is used for steady-state labeling of cells with 13C.

Materials Needed

Protocol

E. coli OD2 (13C, 98%; 15N, 98%; Silantes # 106202; Spectra, Andover, MA), supplemented with: 0.5% (w/v) 13C6-d-glucose (13C >98%; Spectra), 0.5% (w/v) 13C,15N Algal Amino Acid Mixture (10% (w/w) Ala, 2.8% (w/w)Arg, 7.8% (w/w) Asp, 0.2% (w/w) Cys, 11% (w/w) Glu, 8.2% (w/w) Gly, 1.9% (w/w) His, 5.9% (w/w) Ile, 11.6% (w/w) Lys, 2.8% (w/w) Met, 2.1% (w/w) Phe, 6.3% (w/w) Pro, 4.4% (w/w) Ser, 4.9% (w/w) Thr, 0.9% (w/w) Tyr, 7.9% (w/w) Val, 13C >97.4%, 15N > 97.9%; Chlorella Industry Co., Ltd., Tokyo), 10% (v/v) deuterium oxide (D2O) (Sigma-Aldrich, St. Louis, MO), 10% inoculum medium containing 0.5% (w/v) 13C6-d-glucose, 0.1% (w/v) cysteine hydrochloride, and 0.05% (w/v) resazurin with 5 mM sodium 2,2-dimethyl-2-silapentane-5-sulfonate (DSS) (Sigma-Aldrich) as an internal standard.[1]

Usage Examples

Nakanishi, Y., S. Fukuda, E. Chikayama, Y. Kimura, H. Ohno, and J. Kikuchi (2011) Dynamic omics approach identifies nutrition-mediated microbial interactions. J. Proteome Res. 10 (2):824–836.PMID:21058740

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References

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  1. Nakanishi, Y et al. (2011) Dynamic omics approach identifies nutrition-mediated microbial interactions. J. Proteome Res. 10 824-36 PubMed EcoliWiki page