Nutrient Media for Plant Tissue Cultures
One of the first decisions that must be made when developing a tissue culture system is what
medium to use. Nutrient media for plant tissue culture are designed to allow plant tissues to be
maintained in a totally artificial environment. Many different tissue culture media have been
developed, but only a few have found wide-spread use, e.g. MS (Murashige and Skoog, 1962). SH
(Shenck and Hildebrandt), and Gamborg's B5. One of the most successful media, devised by
Murashige and Skoog (Murashige and Skoog, 1962) was formulated by analyzing the inorganic
components in tobacco plants and then adding them to medium in amounts similar to those found
in the plants. Not only did they find that the ions themselves were important, but the forms in
which the ions were supplied were critical as well.
In addition to mineral elements, the macro- and micronutrients that are similar to what is found in
fertilizers, nutrient media also contain organic compounds such as vitamins, plant growth regulators,
and a carbon source.
I. Mineral elements
N, K, P, Ca, Mg
1. Nitrogen (N)
- Nitrogen is essential for plant growth. Most inorganic nitrogen is converted to
amino acids and then to proteins. Nitrogen is typically added to plant nutrient media as the nitrate
, oxidized) and/or the ammonium ion (NH
, reduced), which are added as inorganic salts.
Inorganic nitrogen generally ranges from 25-60 mM in nutrient media. In devising media, both the
total amount of nitrogen as well as the relative amounts of NO
are important. There are
usually lower levels of NH
in medium; nitrate is usually added at concentrations
between 25 and 20 mM and ammonium at concentrations between 2 and 20 mM. For example, the
amount of NH
in MS medium is less than half that of NO
and in other media the NH
concentration is lower still. Cultures of some species can proliferate on medium containing nitrates
alone, and some can grow on a medium with ammonium as the sole inorganic nitrogen source if one
or more of the TCA cycle acids (citrate, succinate, malate) are included in the medium at
concentrations of about 10 mM. In poorly buffered media, the use of both nitrogen forms helps
maintain pH. Also, many plant species appear to respond best if they are given both forms, although
the reason for this is not known.
Nitrogen may also be added to medium in an organic form, as amino acids such as proline or
glutamine, hydrolysates (such as casein hydrolysate), or, as above, as organic acids. Organic nitrogen
is already reduced, i.e. in the form in which most nitrogen exists in the plant, and so may be taken up
more readily than inorganic nitrogen. The organic forms are often added to media that do not
contain ammonium. However, almost always, some inorganic nitrogen is present.
2. Potassium (K)