Athaliana seedlings

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+assays/Mono-association[[:space:]]experiment[[:space:]]of[[:space:]]R401[[:space:]]on[[:space:]]Athaliana[[:space:]]seedlings[[:space:]]in[[:space:]]agar[[:space:]]plates/dataset/.gitkeep filter=lfs diff=lfs merge=lfs -text

assays/Mono-association experiment of R401 on Athaliana seedlings in agar plates/protocols/R401onAthalianaSeedlingsAgarPlates.md

+## Mono-association experiment of R401 on A. thaliana seedlings in agar plates
+
+This protocol is adapted from (25). In brief, *A. thaliana* seeds were sterilized, germinated, and transferred to 1⁄2 MS agar plates without sucrose, as described before. After transfer of seedlings, plants were grown for another 14 days under the same conditions. R401 wild-type and mutants were grown in 50% TSB overnight as described before. Bacterial cells were pelleted by centrifugation, washed 3x in 10 mM MgCl2 and OD~600~ was measured and adjusted to 0.0001. Agar plates were flushed with 15 ml of bacterial suspension for 5 min. The bacterial suspension were removed, and plants were carefully transferred to new 1⁄2 MS agar plates. After 24 h, roots were cut using a sterile scalpel and collected in pre-weighed, sterile 2 ml tubes containing 1 steel bead (3 mm diameter). Tubes were weighed again to assess the root fresh weight. Subsequently, roots were ground in a Precellys 24 TissueLyser (Bertin Technologies) for 2 x 30 s at 6,200 rpm at 15 s intervals. Then, 250 μl of sterile 10 mM MgCl2 were added to each tube and roots were ground again under the same conditions. Each sample was subsequently 5x 1:10 diluted in sterile 10 mM MgCl2. Undiluted samples and each dilution were plated on 50% TSA square plates, dried and left to grow at 25 °C until single colonies appeared. Pictures were taken and single colonies were counted blinded.
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