ProtocolsRecipesABMS < Lab

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---+++ ABMS: Acinetobacter Minimal Succinate

Courtesy of the Averhoff lab, with modifications for available reagents.

To make standard ABMS, combine the following *pre-sterilized* ingredients at room temperature:
|  *1L*  |  *5L*  |  *Component*  |
| 880 mL | 4.4 L | diH<sub>2</sub>O |
| 20 mL | 100 mL | 1M Sodium Succinate |
| 50 mL | 250 mL | 20&times; Mineral Solution |
| 50 mL | 250 mL | 20&times; Phosphate Buffer |

To make ABMS agar:

Per liter of media, add 16 g/L agar and 1 mL/L antifoam to diH<sub>2</sub>O, then autoclave in a 2 L flask. When water and agar are cool enough to touch, add component solutions. Optional: warm components at 55°C in a water bath before combining with agar to reduce the temperature difference when combined.

---++++ ABMS Component Recipes

*SL9 (trace minerals)*
|  *1L*  |  *Component*  |
| 800 mL | diH<sub>2</sub>O |
| 12.8 g | Nitrilotriacetic acid |
| 2 g | FeSO<sub>4</sub>&bull;7H<sub>2</sub>O (note: first dissolve in 2M HCl) |
| 104 mg | CoCl<sub>2</sub> (anhydrous) |
| 122 mg | MnCl<sub>2</sub>&bull;4H<sub>2</sub>O |
| 70 mg | ZnCl<sub>2</sub> |
| 36 mg | Na<sub>2</sub>MoO<sub>4</sub>&bull;2H<sub>2</sub>O |
| 13 mg | NiCl<sub>2</sub> |

Add 1 mL of 10 mL 10&times; solution:
| 60 mg | H<sub>3</sub>BO<sub>3</sub> |
| 20 mg | CuCl<sub>2</sub>&bull;2H<sub>2</sub>O |

Adjust pH to 6.5 using 1 M NaOH. Bring volume to 1000 mL with diH<sub>2</sub>O. Note: this requires a lot of NaOH, so leave room when bringing up to volume.

*Mineral Solution (20&times;)*
| *500 mL*  |  *Component*  |
| 10 g | NH<sub>4</sub>Cl |
| 5.8 g | MgSO<sub>4</sub>&bull;7H<sub>2</sub>O |
| 1 g | KNO<sub>3</sub> |
| 0.67 g | CaCl<sub>2</sub>&bull;2H<sub>2</sub>O &dagger; |
| 20 mg | (NH<sub>4</sub>)<sub>6</sub>Mo<sub>7</sub>O<sub>24</sub>&bull;4H<sub>2</sub>O |
| 10 mL | SL 9 |

*&dagger; Substitution:* You may use 465.8 µL of a 1 M CaCl<sub>2</sub> per 500 ml of stock solution for this component. The final concentration of Ca<sup>2+</sup> in the 20&times; stock is 9.12 mM.

Bring to 500mL in a graduated cylinder with deionized water.

*Phosphate Buffer (20&times;)*
|  *500mL*  |  *Components*  |
| 68 g | KH<sub>2</sub>PO<sub>4</sub> |
| 132.5 g | Na<sub>2</sub>HPO<sub>4</sub>&bull;H<sub>2</sub>O |

Bring to 500 mL with diH<sub>2</sub>O; pH should be ~6.9-7.0 (you may add 400 mL diH<sub>2</sub>O first and bring pH up to 6.8 then complete volume to 500mL)

* Source: Averhoff lab
Barrick Lab > ProtocolsMediaRecipes > ProtocolsRecipesABMS
Topic revision: r6 - 2019-06-23 - 12:09:45 - Main.JeffreyBarrick