Refer to Supplemental File, \"57Fe Wall Loss Experiment diagram\" for the steps indicated in bold.
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All materials were soaked overnight with heating in 1.5% Citrad Citric Acid liquid cleaner in deionized water, rinsed in RO water, and soaked in 10% HCl in Milli-Q ultrapure water for one week (due to time constraints), then rinsed with MilliQ ultrapure water, let dry in an AirClean 400 work station overnight, and double-bagged in polyethylene bags (Mellett et al., 2017). M9 Minimal Media for bacterial cultures was made using Milli-Q ultrapure water, containing final concentrations of 33.7 mM Na\u2082HPO\u2084\u00b72H\u2082O, 22 mM KH\u2082PO\u2084, 8.56 mM NaCl, 18.7 mM NH\u2084Cl, 0.1 M magnesium chloride, 0.1 M calcium chloride, 2 mg/ml Thiamine HCl in 70% EtOH, and 20% Glucose (Kutter & Sulakvelidze, 2004). Half of the media was chelexed using Chelex-100 resin (Pai et al., 1988) that was not acid-cleaned (C), and half remained un-chelexed (B). Half of each treatment was spiked with 10 \u00b5M labeled \u2075\u2077FeSO\u2084(2) while half remained un-spiked (1). A volume of 25 ml was then filtered through either a 0.2 \u00b5m Sterivex PVDF syringe filter for the dissolved fraction (D), or a 0.02 \u00b5m Whatman Anatop syringe filter for the soluble fraction (S) (Gledhill & Buck, 2012). Samples were directly filtered into trace metal clean polycarbonate Erlenmeyer flasks, placed in a clean bag with a small opening to vent, and left shaking at 37 \u00b0C for one week. Samples of each treatment were taken initially, then after the 1st, 3rd, and 7th days (t= 00, 01, 03, 07). The samples were diluted, acidified, and analyzed using an ELEMENT XR High Resolution Inductively Coupled Plasma Mass Spectrometer (HR-ICP-MS). The limit of detection (LOD) for \u2075\u2076Fe and \u2075\u2077Fe were 0.3 nM and 0.012 nM, respectively (Shrivastava & Gupta, 2011).
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Problem Report:
\n\u200bThe original measurements of the M9 minimal media \u2075\u2076Fe contamination was high, so in an attempt to lower background contamination the media was chelexed. However, the Chelex-100 resin was not rinsed with acid, so was less active for removal of metals. This resulted in higher than expected background concentrations of metals.
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Three samples were contaminated with bacterial growth (as indicated by visual turbidity and confirmed using SYBR nucleic acid stain and epifluorescence microscopy) over the course of the week. On t=3, sample B2 was contaminated. By t=7, sample B1D and B1S were contaminated.
This data was collected as part of a method development study investigating the precipitation and wall loss of labeled \u2075\u2077Fe when added to M9 Minimal Media, which is used to grow the model bacterial species Escherichia coli. The bulk media was prepared with the same components and a portion was treated with Chelex-100 to remove metals, while the other portion remained un-chelexed. Half of each treatment was spiked with labeled \u2075\u2077Fe and either 0.2 \u03bcm or 0.02 \u03bcm filtered for comparison of the dissolved and soluble fractions. The \u2075\u2077Fe content was monitored over four time points for one week in a shaking incubator, under the same conditions used to culture E. coli for labeling experiments.
BCO-DMO Processing:
\n- modified parameter names to conform with BCO-DMO naming conventions\u00a0(replaced spaces with underscores, removed puncutation, placed letters before numbers);
\n- formatted year to yyyymmdd;
\n- replaced blanks with \"nd\";
\n- replaced mu symbol with u in DESCRIPTION column.