Preparation of Plant Tissue Culture Media

Since the pioneer plant tissue culture studies, one of the main objectives was to design a proper medium that supports sufficient growth of the explants in a totally artificial environment. The first medium formulations used for plant culture work were based on experience of microorganisms. These media contained several distinct classes of compounds:

• Carbon source
• Organic supplements
• Inorganic salts
• Trace elements.

As works on plant cell cultures progressed, attention was paid in order to specifically define media for an optimal growth of the variable plant cells, tissue, and organs. Today, the most commonly used culture media are based on following components:

• Macroelements—N, P, K, Ca, Mg, and S
• Microelements—Fe, Cu, Mn, Co, Mo, B, I, Zn, Cl and sometimes Al, Ni, and Si
• Carbon source—Sucrose, glucose, fructose, or sorbitol
• Organic compounds (Vitamins)—thiamine, niacin, pyridoxine, biotin, folic acid, ascorbic acid, tocopherol
• Myo-inositol or casein hydrolysate
• Complex organics—Coconut milk and water, yeast extract, fruit juices, or pulps
• Plant growth regulators—Auxins, cytokinins, Gibberellins, abscisic acid
• Gelling agents—Agar, agarose, gellan gum
• Other components—MES, activated charcoal, antibiotics, fungicides.

The number of plant species that have been cultured and the different media used are extensive. Once the plant species to be cultured is decided upon, a suitable medium formulation should be selected. One of the most successful media, is the MS medium, was devised by Murashige and Skoog (1962), based on the constitution of tobacco plants. The choice or formulation of the media basically depends on the requirements of the cultivated explant and species.

Media Formulation
As mentioned above, MS medium was developed for culture of tobacco explants and the formulation was based on the analysis of the mineral compounds present in the tobacco tissue itself (Table 1). It is noted for its high salt levels, particularly K and N salts. LM medium (Linsmaier and Skoog, 1965) is a revision of the MS medium, in which the organic constituents have been modified and only thiamine and myo-inositol retained. White’s medium (White, 1963) was devised for the culture of tomato roots and has lower concentrations of salts than MS medium. Gamborg’s B5 medium (Gamborg et al., 1968) was devised for soybean callus culture and contains a much greater proportion of nitrate compared to the ammonium. SH medium (Schenk and Hildebrandt, 1972) was developed for callus culture of both monocotyledons and dicotyledons. Nitsch’s medium (Nitsch and Nitsch, 1969) was developed for another culture. It contains lower salt concentrations than that of MS, but not as low as that of White’s medium. KM medium (Kao and Michayluck, 1975) was designed to grow cells and protoplasts of Vicia at a very low cell density in liquid media. The medium of Chu (N6) is defined to improve the formation, growth, and differentiation of pollen in rice. The concentration of ammonium proved to be crucial for the development of callus. Recently other media have been developed for specific explants and species.