The efficiency of the excitation capture by open Photosystem II (PSII) reaction centres was measured by the Fv/Fm ratios in a collection of winter and spring oats in order to assess the effects of hardening and freezing on the functionality of PSII and also the suitability of a chlorophyll fluorescence‐based method to screen oat cultivars for frost tolerance. A significant reversible decrease in Fv/Fm was found in all genotypes during acclimation to low, non‐freezing temperatures. Fv/Fm analysis appears to be an attractive test for the evaluation of frost tolerance in oats, being rapid, non‐invasive and capable of monitoring a trait related to a crucial stage in the acquisition of frost tolerance. It is more sensitive and precise than other standard methods and highly correlated with field‐evaluated frost damage. The measurements made during recovery 1 or 2 days after stress when the visual symptoms are not yet expressed, were especially advantageous because of the large variability in genotype response. The r‐values (close to 0.8) were reduced due to the non‐standard behaviour of the winter cultivar ‘Aintree’. The cold acclimation response of this genotype has been analysed in detail and the limits of artificial freezing tests are discussed.

Two quantitative trait loci (Fr-H1 and Fr-H2) for frost tolerance (FT) have been discovered on the long arm of chromosome 5H in barley. Two tightly linked groups of CBF genes, known to play a key role in the FT regulatory network in A. thaliana, have been found to cosegregate with Fr-H2. Here, we investigate the allelic variations of four barley CBF genes (HvCbf3, HvCbf6, HvCbf9 and HvCbf14) in a panel of European cultivars, landraces and H. spontaneum accessions. In the cultivars a reduction of nucleotide and haplotype diversities in CBFs compared with the landraces and the wild ancestor H. spontaneum, was evident. In particular, in cultivars the loss of HvCbf9 genetic variants was higher compared to other sequences. In order to verify if the pattern of CBF genetic variants correlated with the level of FT, an association procedure was adopted. The pairwise analysis of linkage disequilibrium (LD) among the genetic variants in four CBF genes was computed to evaluate the resolution of the association procedure. The pairwise plotting revealed a low level of LD in cultivated varieties, despite the tight physical linkage of CBF genes analysed. A structured association procedure based on a general liner model was implemented, including the variants in CBFs, of Vrn-H1, and of two reference genes not involved in FT (a-Amy1 and Gapdh) and considering the phenotypic data for FT. Association analysis recovered two nucleotide variants of HvCbf14 and one nucleotide variant of Vrn-H1 as statistically associated to FT.

Barley (Hordeum vulgare L.) production in many regions is constrained by low temperatures. Numerous screening methods have been proposed for studies of freezing tolerance (FT) and winter hardiness (WH). Here, we compared the response to low temperature of 54 barley genotypes released in Europe. Major components of WH were analyzed in fi eld and growth chamber experiments under different hardening and freezing conditions. Phenotype screening of freezing injury (lethal temperature for 50% of the plants [LT50], plant survival, and chlorophyll fluorescence) and indirect evaluations (molecular-marker-based analysis of vernalization requirement) were used. The maximum quantum yield of the photosystem II photochemistry measured as the variable (Fv) to maximal (Fm) fluorescence ratio (Fv:Fm) analysis was confirmed as a reliable method of screening geneticdiversity for FT in plants at early growth stages. Variability for FT was also found after shorter acclimation at optimal (3/1°C day/night) or suboptimal (12/7°C day/night) hardening temperatures. High levels of FT and WH were found in both winter and facultative growth habits. Facultative genotypes were more responsive to early hardening than spring or winter types. Some of them coped best with frost in both laboratory and field experiments. The most tolerant winter genotypes coped best with prolonged exposure to low temperature in the fi eld experiments. A conservative estimation of the role of FT in WH implies that at least 50% of WH was associated with the level of FT. A combined analysis using physiological and molecular tools is proposed for phenotyping WH in large populations

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