This note is for those who use or intend to use maize organ cultures for physiological or genetical work.
Nitrogen and carbon are the two major macronutrients required by living cells. In the majority of media formulations commonly used for plant tissue culture, the main nitrogen sources are N03- and NH4+ and also a wide range of amino acids and casein amino acids. During the course of experiments on amino acid metabolism we observed that primary root tips were able to grow in a medium devoid of nitrogen. This unexpected result induced us to find out if the presence of nitrogen in the medium is an absolute nutritional requirement for short term cultures.
For this purpose we cultured embryos, shoot and primary root tips in the presence or absence of nitrogen. Seeds of W22 inbred line were surface sterilized and germinated as previously described (Gavazzi, G. et al., Theor. Appl. Genet. 46:339-345, 1975). Embryos dissected from the endosperms were cultured on a medium consisting of the major and minor salts of Murashige and Skoog (Physiol. Plant. 15:473-497, 1962) supplemented with thiamine (0.4mg/l), sucrose (30g/l) and solidified with agar. For shoot and root cultures, tips were excised shortly after their emergence (10-15 mm long), transferred aseptically in a flask and cultured in a modified White (Growth 7:53-56, 1943) and F (Racchi et al., Plant Sci. Lett. 13:357-364, 1978) media, respectively. The flasks were kept on a rotary shaker (100 rpm) at 25 ± 2 C under 14 hours of daily light.
The results obtained (See table) indicate that embryos and shoots show an absolute requirement for a nitrogen source; in fact, after a few days of culture (7 for shoots and 12 for embryos) in the absence of nitrogen, they show a visible growth inhibition with necrosis of the leaves. In contrast, root tips have normal growth, at least for the time period tested, in media devoid of the commonly used nitrogen sources. Root tips seem to grow in these conditions even better than in the presence of nitrate, and the ratio dry weight/fresh weight is higher than that of the control.
This result could be very useful in view of the fact that the lack of a nitrogen source in a culture medium reduces bacteria and fungal growth, thus preventing culture contaminations. Further analyses are under way to evaluate N03- pool, and protein and amino acid content of root tips grown in these conditions.
Chiara Tonelli and Isabella Viani
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