Reference: Tempe, 1977, PNAS 74:2848
AT is prepared as 20X buffer and 20X salts solutions that are supplemented with a carbon source.
20X AT Buffer | g/mol | g/L | Final Concentration (1X) |
---|---|---|---|
KH2PO4 | 136.1 | 214 | 0.079 M |
pH to 7.0 w/ NaOH | 35 (solids) | 0.044 M |
20X AT Salts | g/mol | g/L | Final Concentration (1X) |
---|---|---|---|
(NH4)2SO4 | 132.14 | 40 | 0.015 M |
MgSO4-7H2O (anhydrous,1.6) | 246.5 | 3.2 | 0.6 mM |
CaCl2-2H2O (anhydrous, 0.151) | 147.0 | 0.2 | 0.06 mM |
FeSO4-7H2O | 278.0 | 0.1 | 0.027 mM |
MnSO4-H2O | 169 | 0.024 | 0.0071 mM |
1X AT Medium | g/mol | g/L | Final Concentration (1X) |
---|---|---|---|
20X buffer | 50mL/L | 1X | |
20X salts | 50mL/L | 1X | |
50% glucose or 40% succinate | 10mL/L | 0.5% | |
+/- Bacto-agar | 15 | 1.5% |
- Prepare sterile, autoclaved stocks of 20X salts, 20X buffer and the carbon source. Add the appropriate amounts to either sterile deionized water (AT broth) or molten (55 oC) sterile 1.5% agar-water (AT agar); mix gently, but thoroughly.
- Iron will precipitate after autoclaving salts to form red iron oxides. (i.e., rust)
- After 1X broth made, there is no need to autoclave. So the process of 1X preparation should be sterile. DI should be autoclaved separately.
- The sugar (e.g., 50% glucose, 40% succinate) needs to be filtered before storage.