Microbial Techniques: Preparing Minimal Media

Minimal Media contains the essentials for bacterial species to grow. The media is often used to define if a particular microbial species is a heterotroph, namely an organism that does not have any nutritional requirements beyond core sources of carbon (sugars) and nitrogen (to synthesize amino acids and nucleic acid bases). Auxotrophs, or organisms with nutritional requirements will not be able to grow on minimal media.

Important components are:

  1. Erlenmeyer flask of volume at 2X for amount of media to be made (i.e. 1L flask to make 500mL)
  2. M9 minimal salts solution (5X concentrate)
  3. Distilled water
  4. 1M solution of magnesium sulfate (MgSO4).
  5. 20% solution wt/wt of glucose (20g gulcose in 80g of water). Add powdered glucose (also called "dextrose") to water solution slowly while mixing, else the glucose will clump and not easily dissolve. Filter sterilize when finished - do not autoclave as the heat will carmelize the sugar.
  6. 1M solution of calcium chloride (CaCl2)

  7. To make solid media, include agar base and sterile Petrie plates.
Method After autoclaving, swirl to mix evenly and "temper" at room temperature until you can place your hand on the flask for 2 second (an adult should do this!), then add the following: Swirl to mix evenly.

Liquid media should be distributed to sterile bottles or tubes with a pipette and pipettor. Small bottles can hold 250mL of media for use when needed, for instance, or aliquots of 5mL or 10mL can be transferred to sterile, capped test tubes.

If plates are to be poured, ~25mL of media should be added per Petrie dish and allowed to solidify at room temperature. Poured/solid plates can be stacked and replaced in the original plastic sleeve in which they came. Label the sleeve with the type of media and date poured.

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