Mini-Tn5ScreenPyoverdineLackR569 assay and protocol

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 assays/BGC[[:space:]]prediction[[:space:]]using[[:space:]]antiSMASH/dataset/Table[[:space:]]S2.xlsx filter=lfs diff=lfs merge=lfs -text
 assays/Mini-Tn5TransposonMutantInR401andR569/dataset/.gitkeep filter=lfs diff=lfs merge=lfs -text
 assays/Mini-Tn5ScreenLossOfR401sGrowthInhibitionRsGMI1600/dataset/.gitkeep filter=lfs diff=lfs merge=lfs -text
+assays/Mini-Tn5ForPyoverdineLackofR569/dataset/.gitkeep filter=lfs diff=lfs merge=lfs -text

assays/Mini-Tn5ForPyoverdineLackofR569/protocols/Mini-Tn5TransposonScreenPyoverdineLackofR569.md

+## Mini-Tn5 transposon mutant screen for lack of pyoverdine fluorescence of R569
+
+R569 mini-Tn5 mutants were cultured in 96 well plates at 25 °C and 180 rpm for five days in 50% TSB medium. For the initial mutant screen, fluorescence was acquired at 𝜆~excitation~=395 nm and 𝜆~emission~=470 nm in a microplate reader (Infinite M200 PRO, Tecan). Out of ~2,000 mutants analysed, the fluorescence-based screening identified twelve R569 mutants that showed severely reduced pyoverdine-specific fluorescence but retained median-like growth behaviour (Thresholds: -6x MAD fluorescence, >-1x MAD Abs~600~). For validation, cultures were pre-grown for five days in 50% TSB before sub-culturing into fresh siderophore medium (see **Table S5**) and growth for five additional days. Finally, bacterial culture density (Abs~600~) was determined, and pyoverdine-specific fluorescence of bacterial culture supernatants was captured at 𝜆~excitation~=410 nm and 𝜆~emission~=500 nm. For  comparison between genotypes, fluorescence capacity was calculated by dividing the pyoverdine-specific fluorescence by culture density.
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