Dataset: Metabolomics data for two strains of Pelagibateriales

Sample Collection

Two strains for members of the Alphaproteobacteria order Pelagibacteriales (SAR11) were cultured in the laboratory. LSUCC0261 is an isolate from the SAR11 subclade IIIa and was grown in JW2 media[Henson et al. 2016]. LSUCC0530 is a freshwater strain in the LD12 subclade, and was grown in JW5 media [Henson et al. 2018]. Cells were harvested at late exponential phase by filtration onto 0.1 um Omnipore (Millipore) filters. The filters were stored frozen at -80C until extraction in the laboratory.

In the metadata, 'volume' is the volume filtered for each sample. For each sample, 'cells' is the number of cells per milliliter.

Extraction

A protocol adapted from [Rabinowitz et al. 2007] was used for extraction of intracellular metabolites. Filters were removed from the cryovials and cut into small pieces with methanol-rinsed scissors on combusted aluminum foil. The pieces of the filter were placed in an 8 ml combusted amber glass vial and 1 ml of -20 °C 40:40:20 acetonitrile:methanol:water + 0.1 M formic acid was added to each vial. 25 µl of 1 µg/ml deuterated standard mix (d3-glutamic acid, d4-4-hydroxybenzoic acid, and d5-taurocholate) were added to act as extraction recovery standards. The vials were vortexed to shake apart filter pieces and fully expose them to the ice-cold solvent. They were sonicated for 10 minutes and the extract was transferred to micro-centrifuge tubes using Pasteur pipettes. The filter pieces were rinsed with three times with 500 µl of extraction solvent and the rinse was added to the eppendorf tubes. The extracts were centrifuged at 20,000 × g for 5 minutes. The supernatant was transferred to new 8 ml amber glass vials leaving behind any scraps of filter or pieces of cellular detritus. The extracts were neutralized with 51.2 µl of 6 M ammonium hydroxide and dried down in a vacufuge. The samples were reconstituted in 100 µl labeled injection mix (95:5 water:acetonitrile containing D2 biotin, D6 succinic acid, D4 cholic acid, and D7 indole 3 acetic acid). The solution (100 µl) was placed in a glass insert in an autosampler vial for targeted metabolomic analysis.