new assay Screen for antagonistic interbacterial interactions

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 /_publication/Getzke[[:space:]]et[[:space:]]al.[[:space:]]-[[:space:]]2023[[:space:]]-[[:space:]]Cofunctioning[[:space:]]of[[:space:]]bacterial[[:space:]]exometabolites[[:space:]]drives[[:space:]]r.pdf filter=lfs diff=lfs merge=lfs -text
 _publication/pnas.2221508120.sapp.pdf filter=lfs diff=lfs merge=lfs -text
 _publication/dataverse_files/Table[[:space:]]S2.xlsx filter=lfs diff=lfs merge=lfs -text
+assays/Screen[[:space:]]for[[:space:]]antagonistic[[:space:]]interbacterial[[:space:]]interactions/dataset/Table[[:space:]]S2.xlsx filter=lfs diff=lfs merge=lfs -text
+assays/Screen[[:space:]]for[[:space:]]antagonistic[[:space:]]interbacterial[[:space:]]interactions/dataset/pnas.2221508120fig01.jpg filter=lfs diff=lfs merge=lfs -text

assays/Screen for antagonistic interbacterial interactions/protocols/AntagonisticInterbacterialInteractions.md

+## Screen for antagonistic interbacterial interactions
+
+For the initial mBA experiment (**Fig. 1**), bacterial strains were cultured for seven days in 25% TSB (7.5 g/l Tryptic Soy Broth, Sigma-Aldrich). Briefly, 100 μl of a bacterial solution were resuspended in 50 ml cooled (~38 oC), but still molten, 25% TSA (15 g/l Bacto-Agar, Duchefa Biochemie) and poured into a square petri dish (120x120 mm). After medium solidification, 24 bacterial isolates were spotted on top of the medium using a multi-stamp replicator. The replicator was sterilized by dipping in 70% EtOH (v/v) followed by flaming and cooling. The screen comprising 39,204 binary interactions was conducted once and validated by randomly re-screening 7,470 interaction pairs as described above. All bacterial strains that showed antagonistic activity were rescreened two more times. For the *Ralstonia* inhibition screen, *R. solanacearum* GMI1000 was pre-cultured for two days in CPG medium (1% peptone, 0.5% glucose and 0.1% casamino acids; pH7.0). Before spotting the bacterial cultures, each CPG agar plate (1% Peptone, 0.5% D-glucose, 0.1% casamino acids and 1.5% agar; pH 7.0) was overlaid with 5 ml of *R. solanacearum* suspension (50 μl of pre-cultured *R. solanacearum* in pure sterile water). Excess *R. solanacearum* suspension was removed and the plates were briefly dried, then 5 μl of the bacterial culture were spotted onto the *R. solanacearum*-overlaid plates. All isolates were tested three times. For all subsequent halo assays, strains were cultivated in 50% TSB until turbidity, stored at 4 °C and diluted 1:10 in 50% TSB one day before the experiment. Bacterial cultures were pelleted at 4,000 rpm for 15 min. The resulting bacterial pellets were subsequently washed 3 times and resuspended in 1 ml 10 mM MgCl2. OD600 were measured and set depending on the strain. One hundred microliters bacterial culture were inoculated per 50 ml 25% TSA. After drying, up to nine different 3-μl droplets of bacterial suspensions with 0.4 OD600 were applied with equal distances. For all experiments, plates were incubated at 25 °C for up to 96 hours and photographs were taken thereafter for quantitative image analysis. The size of the halo of inhibition was measured using ImageJ with up to five separate measurements, which were subsequently averaged to reduce variation. Raw data of Fig. 1 are indicated in **Table 2**.