Major metabolic pathways and genes affected by low-temperature treatment were identified and a thorough picture of the early transcriptional changes in sugar beet plantlets upon cold stress was given. Sugar beet (Beta vulgaris L.) is an important source of sugar and bioethanol production in temperate areas worldwide. In these areas, plantlet survival and sucrose yield of mature plants can be seriously limited by low temperatures, especially when plantlets are exposed to freezing temperatures (below 0 °C) at the early developmental stages. This frequently occurs when the crop is sown in early spring or even in autumn (autumn sowing) to escape drought at maturity and pathogen outbreaks. The knowledge of molecular responses induced in plantlets early upon exposure to low temperature is necessary to understand mechanisms that allow the plant to survive and to identify reactions that can influence other late-appearing traits. In this work, a wide study of sugar beet transcriptome modulation after a short exposure to a cold stress, mimicking what is experienced in vivo by young plantlets when temperature drops in the early spring nights, was carried out by high-throughput sequencing of leaves and root RNAs (RNA-Seq). A significant picture of the earliest events of temperature sensing was achieved for the first time for sugar beet: the retrieval of a great amount of transcription factors and the intensity of modulation of a large number of genes involved in several metabolic pathways suggest a fast and deep rearrangement of sugar beet plantlets metabolism as early response to cold stress, with both similarities and specificities between the two organs.