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Pipeline publication

[…] ellets were reextracted with an additional volume of 1 ml ice-cold sterile water and pooled with the previous extraction. Pooled fractions were flash-frozen using liquid nitrogen and stored at −80°C until further analysis., The collected supernatants were lyophilized and reconstructed in NMR buffer (potassium phosphate buffer in D2O, pH 7.4 [uncorrected], with 500 mM trimethylsilyl propanoic acid [TMSP] as an internal standard). 2D 1H-13C HSQC spectra were collected on a Bruker Avance III-HD 700-MHz spectrometer equipped with a quadrapole-resonance QCI-P cryoprobe (1H, 13C, 15N, and 31P), a SampleJet automated sample changer, Bruker ICON-NMR, and an automatic tuning and matching (ATM) unit. NMRPipe () and NMRViewJ () were used to process and analyze the collected spectra. The TMSP internal standard was used for chemical shift referencing and normalization of NMR peak intensities. NMR peaks from the 2D 1H-13C HSQC spectra were annotated by comparing the observed 1H and 13C chemical shifts to the metabolite reference data from the Platform for RIKEN Metabolomics (PRIMe) (), Human Metabolome Database (HMDB) (), Madison metabolomics Consortium Database (), Metabominer (), and BiomagResBank (BMRB) () with error tolerances of 0.08 ppm and 0.25 ppm for 1H and 13C chemical shifts, respectively. The relative intensity (i.e., concentration) of each metabolite was calculated by averaging the intensities of all NMR peaks unambiguously assigned to the metabolite., For growth analysis, S. aureus cultures were grown aerobically at 37°C in 30 ml TSB supplemented with 14 mM glucose (in 250-ml Erlenmeyer flasks) for 24 h. Culture aliquots of 1 ml were collected hourly and used for optical density (OD600), pH, and metabolite analysis., The effect of various compounds on S. aureus growth was determined by automated spectrophotometric analysis of culture densi […]

Pipeline specifications

Software tools NMRPipe, NMRViewJ, MetaboMiner
Databases PRIMe