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There is an inverse relationship between cell size and capacity to survive the freeze‐preservation protocol. Pregrowth of cell suspensions in media rendered more negative in water potential by addition of mannitol enhances the survival capacity of Acer pseudoplatanus and Capsicum annuum cells but this effect can only partially be explained in terms of the associated reduction in mean cell size. Studies with cell suspensions of Daucus carota indicate the importance for successful freeze‐preservation of the stage in the growth cycle of suspensions propagated in batch culture; highest survival was recorded for cells taken at lag phase or early exponential phase. Regrowth of recovered cells depends upon the establishment of an appropriate inoculum density of cells which have retained the capacity to divide. The dividing cells only achieve a growth rate equal to that of untreated cells after a number of cell generations. A proportion of the recovered cells giving an initial positive fluorescein diacetate reaction lose this capacity rapidly (within 24 h), others lose the capacity more slowly and others, in which the positive reaction persists, are incapable of division. These observations indicate that different levels of injury are inflicted by the freeze‐preservation protocol and that only in a proportion of the cells is the injury reparable or compatible with growth by cell division.

Summary The composition of the inorganic fraction of White's medium has been modified to permit studies in nitrogen nutrition. A preparation of ferric sodium ethylenediamine tetra‐acetate is described which enables an addition of 1 p.p.m. iron to meet the iron requirement of exised tomato roots in culture media whose pH is below 7.5. The content of calcium and magnesium in the medium has been reduced to one‐tenth the standard concentration in order to prevent precipitation at pH values above 6.0, particularly when using enhanced phosphate concentrations. Techniques whereby the pH of root culture medium can be adjusted aseptically or stabilized by the addition of soluble phosphates or of sparingly soluble salts of calcium (phosphate, carbonate) are described and evaluated. These techniques have made possible a high level of growth of excised tomato roots receiving ammonium as their sole source of nitrogen. The ability of the roots to grow with nitrite as their sole nitrogen source has also been demonstrated. In contrast, hydroxylamine, even at low concentrations, inhibited the extension growth which otherwise occurs on absence of external nitrogen and caused death of the root apices. pH‐growth response curves with ammonium and with nitrate as nitrogen sources indicate pH 4.7–4.9 as optimum for growth with nitrate nitrogen and pH 7.0–7.2 as optimum for growth with ammonium nitrogen. Less complete data indicates pH 5.0–6.0 as the optimum region for growth with nitrite. From studies of the pH drifts occurring as a result of root growth in media containing various ammonium: nitrate ratios and from direct determinations of nitrate and ammonium uptake at high and low pH, it is concluded that the differing pH optima for these two nitrogen sources are not explicable in terms of the direct effects of pH upon the uptake of these ions.

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