eLabFTW2ARC version 2024-12-09. 03/02/2025, 09:35:46   ...

eLabFTW2ARC version 2024-12-09.  03/02/2025, 09:35:46&nbsp; &nbsp; &nbsp;Fetching eLabFTW experiment <b> 269</b>, instance is https://demo.elabftw.net/api/v2/	 <br>03/02/2025, 09:35:46&nbsp; &nbsp; &nbsp;DataHUB URL fetched from elab is "https://git.nfdi4plants.org/usadellab/goxr_characterization.git	 <br>03/02/2025, 09:35:48&nbsp; &nbsp; &nbsp;isa.assay.xlsx has been updated at <b>Ethanol_production_2</b> 	 <br>03/02/2025, 09:35:49&nbsp; &nbsp; &nbsp;Added file ethanol_production_png	 <br>03/02/2025, 09:35:49&nbsp; &nbsp; &nbsp;Finished adding protocol files in ARC	 <br>03/02/2025, 09:35:49&nbsp; &nbsp; &nbsp;All files have been added to ARC, starting to push ARC to	 <br>

assays/Ethanol_production_2/dataset/readme.md

+# Data files
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assays/Ethanol_production_2/protocols/eLabFTW_protocol.md

+**Introduction**
+
+We previously speculated that GOX0313 activity might play a role in the adjustment of the redox balance of NAD+ and NADH under oxygen-limited conditions because expression of the GOX0313 gene was upregulated under oxygen limitation together with the other genes of the pnt operon. Of the various substrates identified for GOX0313, acetaldehyde was reduced with the highest specific activity and is also a likely in vivo substrate, as cytoplasmic carbon catabolism involves decarboxylation of pyruvate to acetaldehyde. Under conditions of a high NADH/NAD+ ratio caused by oxygen limitation or insufficient NADH dehydrogenase activity, acetaldehyde may serve as an electron acceptor for NADH reoxidation by an alcohol dehydrogenase, forming ethanol as product.
+
+**Methods**
+
+Cells of the parental strain and the Δ_goxR_ mutant were cultivated in shake flasks with mannitol medium under vigorous shaking. When the cells had reached the stationary phase, they were harvested by centrifugation, washed and resuspended in 50 mM MOPS buffer adjusted with KOH to pH 6.0. The density of the cell suspension was adjusted to an OD600 of 1. 6 ml of the cell suspension and 6 ml of a 44 mM glucose solution were mixed in a 15 ml Falcon tube. The free space in the tube was flushed with argon gas and the tube immediately closed tightly with a cap. The tube was shaken horizontally at 120 rpm at 30°C for the indicated times. Ethanol in the cell-free supernatant of the cell suspensions was determined by gas chromatography (GC) using an Agilent 7890A gas chromatograph (Agilent Technologies, Waldbronn, Germany) as described previously. Calibration was performed with ethanol solutions from 0.05 to 3.2 mM as an external standard. The detection limit of ethanol was 0.1 mM under the conditions used. Butanol was added to the samples as an internal standard at a concentration of 2 mM. Concentrations were calculated from peak areas using calibration with external ethanol and internal butanol standards.
+
+**Results**
+
+As shown in the figure, the parental strain indeed produced some ethanol in a time-dependent manner. The ΔgoxR mutant also formed ethanol, but at a much lower rate than the parental strain, correlating with a lower expression of the GOX0313 oxidoreductase.
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