Dendrobium(Orchidaceae, Epidendoideae) plants have flowers with a wide variety of colors that persist for a long period throughout the year. The yellow coloration ofDendrobiumflowers is mainly determined by the flavonol pathway and the flavone pathway, but the relevant biosynthesis mechanisms during vernalization remain unclear. To explore the similarities and differences in flavonoid biosynthesis in different tissues during vernalization, we selected two species ofDendrobiumfor a flower color study:Dendrobium capillipesRchb (which has yellow flowers) andDendrobium nobileLindl (which has white flowers). We collected a total of 36 samples from six tissue types and bothDendrobiumspecies during vernalization and subjected the samples to metabolic profiling and transcriptome sequencing. A total of 31,504 differentially expressed genes (DEGs) were identified between different tissues of the twoDendrobiumspecies by transcriptomic analysis. However, many differentially accumulated metabolites (DAMs) and DEGs were enriched not only in the general pathway of “flavonoid biosynthesis” but also in multiple subpathways of “flavone and flavonol biosynthesis”. According to a combined transcriptome and metabolome analysis, Putrescine hydroxycinnamoyl transferase 1 (LOC110093422) may be the main gene responsible for the differences in flavonoid accumulation during vernalization, which is closely associated with yellow flowers. Taken together, the results of our study preliminarily revealed the metabolites responsible for and the key genes regulating flavonoid biosynthesis during vernalization. These results provide a basis for the further study of the molecular mechanism of flavonoid synthesis during vernalization.