Rose is the most economically important ornamental plant. However, cold stress seriously affects the survival and regrowth of garden roses in northern regions. Cold acclimation was studied using two genotypes (Rosa wichurana and R. hybrida ‘Yesterday’) selected from a rose breeding program. During the winter season (November to April), the cold hardiness of stems, soluble sugar content, and expression of dehydrins and the related key genes in the soluble sugar metabolism were analyzed. ‘Yesterday’ is more cold-hardy and acclimated faster, reaching its maximum cold hardiness in December. R. wichurana is relatively less cold-hardy, only reaching its maximum cold hardiness in January after prolonged exposure to freezing temperatures. Dehydrin transcripts accumulated significantly during November–January in both genotypes. Soluble sugars are highly involved in cold acclimation, with sucrose and oligosaccharides significantly correlated with cold hardiness. Sucrose occupied the highest proportion of total soluble sugars in both genotypes. During November–January, downregulation of RhSUS was found in both genotypes, while upregulation of RhSPS was observed in ‘Yesterday’ and upregulation of RhINV2 was found in R. wichurana. Oligosaccharides accumulated from November to February and decreased to a significantly low level in April. RhRS6 had a significant upregulation in December in R. wichurana. This study provides insight into the cold acclimation mechanism of roses by combining transcription patterns with metabolite quantification.