Antifreeze proteins (AFPs) accumulate in the leaves of barley during cold acclimation, where they may inhibit ice recrystallization and produce freezing resistance of the plant. Four parental diallel crosses of the barley varieties were used to determine the heritability of AFPs and the relationship between the accumulation level of AFPs and freezing resistance. The concentration of apoplastic proteins in the cold-acclimated leaves was increased in the mean by four-fold over as compared with that of nonacclimated. The diallel cross analyses revealed that the gene of Sacheon 6 was dominant and those of Reno and Dongbori 1 were recessive. The AFPs had high narrow-sense heritabilities. The general combining ability effects of Reno and Dongbori 1 were much higher than the other parents. The bands of 32-kD for GLP, 35-& 28-kD for CLP and 25-, 22- & 16-kD for TLP were observed in the apoplastic extracts from cold-acclimated plants, but there were no clear differences between the parents and Fl hybrids. The concentrations of AFPs were significantly correlated with the degree of freezing resistance, indicating that the concentration of AFPs in the plant is the very important factor for freezing resistance.

Freezing-resistant plants can survive subzero temperatures by withstanding extracellular ice formation. During cold acclimation, their leaves accumulate antifreeze proteins (AFPs) that are secreted into the apoplast and have the ability to modify the normal growth of ice crystals. Three barley, two wheat and two rye cultivars were grown under two different temperature regimes (20/16~circC and 5/2~circC , day/night). Apoplastic proteins from winter cereals were separated by SDS-PAGE and detected with antisera to AFPs from winter rye. Apoplastic proteins accumulated to much higher levels in cold-acclimated (CA) leaves compared with nonacclimated (NA) ones in winter cereals. After cold acclimation, the protein concentration of apoplastic extracts increased significantly from 0.088 mgmL-1 to 0.448 mgmL-1 , with about 5-fold increment. Also, the apoplastic protein content per gram leaf fresh weight in CA leaves ranged from 31 ~mu~textrmg (gFW)-1 to 120 ~mu~textrmg (gFW)-1 with an averaged value of 77 ~mu~textrmg (gFW)-1 , and coefficients of variation of 54.9%. The CA leaves in Musketeer (a Canadian winter rye cultivar) showed the greatest AFPs and antifreeze activity followed by 'Geurumil' (a Korean winter wheat cultivar), and 'Dongbori l' (Korean facultative barley cultivar). The proteins secreted into the wheat leaf apoplast at CA condition were more numerous than those observed in winter rye, where two β -1,3-glucanase-like proteins (GLPs), two chitinase-like proteins (CLPs) and two thaumatin-like proteins (TLPs) accumulated during cold acclimation. The proteins in barley leaf apoplast at CA conditions were a little different from those in wheat leaves. The AFPs were various among and within species. More freezing-resistant cultivars had more clear and numerous bands than less freezing-resistant ones. The high determination coefficient (R2 =91 %) between freezing resistance and AFPs per gram leaf fresh weight indicated that the amount of AFPs was highly related to freezing resistance in winter cereal crops.