The possible significance of specific biochemical changes in relation to cold acclimation has been discussed extensively (13), but causal relationship between metabolic changes and cold acclimation is difficult to establish. Generally, the development of frost hardiness has been reported to be accompanied with changes in hormones (12,19,24), and in quality and quantity of proteins (14,16). Under a low temperature regime, an increase in leaf soluble protein content was observed only in those potato species which are able to acclimate, i.e. Solanum acaule and Sc4 (4). The net increase in soluble proteins was significantly and positively correlated with the increase of frost hardiness in these species (4). Potato stem cuttings can be easily rooted and propagated in a nutrient agar medium. Such 'stem-cultured plants' can be acclimated to similar frost hardiness levels as with normal tuberpropagated plants grown in pots (5). ABA could substitute for low temperature treatment in inducing frost hardiness in stem-cultured Sc plants, and GA did not affect the cold acclimation (5).The following is a report of experimental results in attempting to determine if there is a relationship between ABA changes and protein synthesis during the development of frost hardiness in the potato during cold acclimation. MATERIALS AND METHODSPlant Materials. Solanum tuberosum L. cv 'Red Pontiac' was propagated from seed tubers, whereas Solanum commersonii (Oka 5040) was propagated from stem cuttings. Plants were planted in 15-cm diameter pots each in a mixture of soil, sand, and peat (3:2:2, v/v). Plants of both species were grown in a uniform environmental chamber for 2 months in a regime of 14-h photoperiod with 450 ,uEm-2 s-' PAR and 20/15°C D/N temperature. Plants of each species were then divided into two groups. One group (controls) was maintained in the same chamber. The second group (treated) was exposed to low temperature of 2°C D/N with a 14-h photoperiod. Frost hardiness of fully expanded leaves was determined after 0, 1, 2, 3, 4, 5, 10, and 15 d of treatments. At the same time, samples were also collected for chemical analyses and moisture determination. The conditions for propagating and growing stem cultures of St and Sc plants were similar to an earlier report (5).Frost Hardiness Evaluation. Excised leaflets or whole shoots (stem-cultured plants) were subjected to controlled freezing (5). After freezing, leaf tissues were placed over ice in a moisturesaturated box in a 5C room and thawed overnight, followed by warming to room temperature. Viability of the tissues after freezing tests was estimated by the conductivity method (20

Plasma membrane alteration In two tuber-bearing potato species during a 20-day cold acclimation period were investigated. Leaf-callu tssues of the frost-resistant Soeaum acaule Hawkes 'Oka 3878 and the frostsusceptible, comly grown SOa_w tuberom 'Red Pontiac,' were used. The former is a species that can be hardened after subjecting to the low temperature, and the latter does not harden. Samples for the electron microscopy were prepared from callus cultures after hardenin at 2 C In the dark for 0, 5, 10, 15, and 20 days. After 20 days acclimation, S. acial incressed in frost hardiness from -6 to -9 C (killing temperature), whereas frost hardiness of S. tubemsun remained unchanged (killed at -3 C). Actually, after 15 days acclimation, a -9 C frost hardiness level In S.acaude callus cultures had been achieved. Studies in our laboratory have shown that the commonly grown potato (Solanum tuberosum L.) possesses no frost tolerance (4), whereas a number of noncultivated species can survive at -4 C or colder without frost injury. Some of these species, for example, Solanum acaule Hawkes and Solanum commersonii Dun., can also be cold acclimated to increase their frost hardiness up to -9 C or to -12 C, respectively (4). Also, similar increases in hardiness could be achieved both for potted and leaf-callus cultures (4). Chen et al. (4) suggested that the callus, a homogeneous mass of undifferentiated parenchyma cells, could serve for cold acclimation studies in place of intact potato plants. The stage of differentiation is easily controlled in callus cultures. They thus provide a desirable material for the study of frost hardiness at cellular level.Studies of ultrastructural alterations in relation to frost hardiness in plants are limited (3, 9, 20, 24). The observations of chloroplasts (26) and mitochondria (3) during cold acclimation treatments indicate some structural alterations in these organelles which may relate to plant adaptability to a given environment.