Roses (Rosa hybrida) are the most important ornamental cut-flowers and breeders' main focus is to develop new desirable modern cultivars. Rose breeding programs center on the introduction of new flower colors, thornless stems, higher production and good post-harvest performance. The study of the main pollen traits, such as pollen quantity and quality, viability, longevity, morphological homogeneity, germination and tube growth, is important for building suitable breeding programs. Recently, a number of studies have shown that reactive oxygen species, like hydrogen peroxide, and nitric oxide, are involved in a wide range of signaling processes including pollen tube growth and pollen-pistil recognition. Pollen viability after anther dehiscence is crucial for successful crossbreeding. In the present work, pollen grains from 5 hybrid tea rose cultivars were stored at -20 °C up to 12 months. Pollen viability and germination rate was monitored in order to provide useful information about pollen storage length. Additionally, pollen grains were tested for their content in hydrogen peroxide and nitric oxide by using a novel approach where the fluorescence is read in a quantitative RealTime PCR (qRT-PCR) machine. Pollen viability and in vitro pollen germination capacity varied among the rose genotypes, while a progressive decrease was evidenced during 12 months of storage at low temperature. Both hydrogen peroxide and nitric oxide production were found to be genotypedependent, whilst accumulation of the two molecules was observed during the storage period. A putative detrimental effect of these molecules during pollen conservation is hypothesized.