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3a To confirm this assumption, the C-labelled building blocks [1-C]acetate, [1,2-C]acetate and [1-C]propionate were fed to exponentially growing shake-flask cultures of Verrucosispora maris AB-18-032.
Once the trees are grown, shake the tree & pick up the fruit.
In brief, Salmonella overnight cultures grown in LB shaking at 37°C were back diluted 1∶200 in LB and grown shaking until they reached an OD of ∼1.2.
Cultures were grown shaking at 37°C to OD600 0.5, induced with 1 mM IPTG and grown for an additional 3 hours.
For purification of His6-tagged CPD fusion proteins, overnight cultures of the appropriate strain were diluted 1∶500 into 1 L 2YT media and grown shaking at 37°C.
LB kan cultures for the sdiA+ strains or LB kan cam cultures for the sdiA mutant strains were grown shaking at 37°C or 30°C overnight.
The cultures were grown shaking at 37°C, 250 rpm.
Cultures were grown shaking at 30°C to exponential phase (OD600 approx. 0.8) in LB broth containing the appropriate antibiotics.
For the live bacteria treatments, A. machipongonensis liquid cultures were grown shaking in HN media at 30°C for 24 hr.
Cells were grown shaking at room temperature and visualized at times 0 270 min to allow the cells to take up the dye by endocytosis.
Cultures were then grown to OD600 0.2, treated with 5 μg∙ml−1 of C18G peptide (Anaspec, Fremont, CA), and grown shaking at 37°C for 90 min. Following incubation, alkaline phosphatase activity was measured.
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