This section is not about your regular LB, MHB, BHI, TSB...etc media.
Media Recipes
Some short notes about physiology of media making:
Rich Media: For running physiological experiments, note that they lack buffering capacity. They
also have lower levels of certain cations such as Mg or Fe.
Minimal media: Make sure all essential components are present. Read below.
Following should be considered when making media.
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Nitrogen source: Use Ammonia and a rich nitrogen source such as a defined aminoacid mix
and/or casamino acids, Tryptone, BSA.
Carbon source: Not necessary if you are adding amino acids or other nitrogen source. But
necessary if only NH4 is present as the N source. Succinate or other TCA cycle intermediates are
preferred for Pseudomonads and EM or ED pathway intermediates are preferred for
enterobacteria (preferably Glc or Gly).
Essential cations: K+, Mg++ and Fe++ (Na+ is needed by certain bacteria). Other cations are trace
elements and no need to add.
Essential anions: (i) Sulfur: best given as fresh Cys but commonly given as SO4 ion. (NH4SO4 is
the common chemical to use). (ii) PO4: normally supplied as either Na or K salts.
Vitamins: wildtype bacteria can make all their vitamins, unless you are using a minimal media
with a auxotroph (eg: DH5alpha needs Thiamine)
Nucleic acid bases: Bacteria will make them in minimal media, and they will salvage it (from
Yeast extract or from BHI) in rich media. If needed, best to give as nucleoSIDES (Adenosine or
Guanosine, cytidine).
Trace elements: Mn, Mo, Bo, Cu and other elements are usually present as chelates in the KPO4
salts. Calcium can be added to minimal media <250uM if needed.
Buffers to use: Potassium or Sodium Phosphate buffers (50-150mM Phosphate ion) are popular
because they supply essential ions as well. Other buffers are MOPS for pH 7 and MES for low pH
work (at 50mM). Never use TRIS buffers for biological work.
Making media for live fluorescent microscopy. Avoid any protein digests or yeast extract. Use
BSA and ammonia.
Single factor that will increase the growth rate is the quality of Nitrogen source. More
protenacious, better the growth, specially if rich in Gln and Glu. Combining N and C sources will
increase it more. Don't use a catabolic repressing sugar with another sugar (such as Glc and Gly
combination).
Trace element mix and the vitamine mix became popular after Neidhardt published his seminal
paper about bacterial growth media. Unless you are doing some thing funky, no need to use
them.
Neidhardt also published these two handbook which are quite good.
Neidhardt, F. C., J. Ingraham, and M. Schaechter. 1990. Physiology of the Bacterial Cell: A
Molecular Approach. Sinauer Associates, Sunderland, MA. 520 pagess.
Neidhardt, F. C. (Ed. in Chief), R. Curtiss III, J. L. Ingraham, E. C. C. Lin, K. B. Low, B. Magasanik,
W. S. Reznikoff, M. Riley, M. Schaechter, and H. E. Umbarger (eds). 1996.Escherichia coli
and Salmonella: Cellular and Molecular Biology. American Society for Microbiology. 2 vols.
2898 pages.
HK 2012.