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Epigenetic phenomena have been associated with modifications of chromatin structure. These are achieved, in part, by histone post-translational modifications including acetylations and deacetylations, the later being catalyzed by histone deacetylaces (HDACs). Eukaryotic HDACs are grouped into three major families, RPD3/HDA1, SIR2 and the plant-specific HD2. HDAC genes have been analyzed from model plants such as Arabidopsis, rice and maize and have been shown to be involved in various cellular processes including seed development, vegetative and reproductive growth and responses to abiotic and biotic stress, but reports on HDACs from other crops are limited. In this work two full-length cDNAs (HvHDAC2-1 and HvHDAC2-2) encoding two members of the plant-specific HD2 family, respectively, were isolated and characterized from barley (Hordeum vulgare), an agronomically important cereal crop. HvHDAC2-1 and HvHDAC2-2 were mapped on barley chromosomes 1H and 3H, respectively, which could prove useful in developing markers for marker-assisted selection in breeding programs. Expression analysis of the barley HD2 genes demonstrated that they are expressed in all tissues and seed developmental stages examined. Significant differences were observed among tissues and seed stages, and between cultivars with varying seed size, suggesting an association of these genes with seed development. Furthermore, the HD2 genes from barley were found to respond to treatments with plant stress-related hormones such as jasmonic acid (JA), abscisic acid (ABA) and salicylic acid (SA) implying an association of these genes with plant resistance to biotic and abiotic stress. The expression pattern of HD2 genes suggests a possible role for these genes in the epigenetic regulation of seed development and stress response.