Protocol for extracting ppGpp from amino acids or carbon starved cells using AA or Glucose analogs Grow 2-3 ml of cells in minimal T-salts medium supplemented with 0.2 % glucose and 1 mM KH2PO4 At the exponential growth phase (OD600 ≈ 0.5), wash with T-buffer.  Resuspend in the same volume of T-salts supplemented with 0.1% glucose and 0.25 mM Pi Measure OD and dilute cells to a final OD600=0.1- 0.15 T-salts supplemented with 0.1% glucose and 0.25 mM Pi (use the lowest possible volume, usually 200 μl is sufficient). Add 32Pi to a final concentration of 100 μCi/ml and let cells grow for at least one hour Take a 20 μl sample … Continue reading ppGpp assay