BackgroundDNA methylation is one of the most abundant epigenetic modifications, which plays important roles in flower development, sex differentiation, and regulation of flowering time. Its pattern is affected by cytosine-5 DNA methyltransferase (C5-MTase) and DNA demethylase (dMTase). At present, there are no reports onC5-MTaseanddMTasegenes in heterodichogamousCyclocarya paliurus.ResultsIn this study, 6CpC5-MTaseand 3CpdMTasegenes were identified in diploid (2n = 2 ×  = 32)C. paliurus, while 20CpC5-MTaseand 13CpdMTasegenes were identified in autotetraploid (2n = 4 ×  = 64). 80% of identified genes maintained relatively fixed positions on chromosomes during polyploidization. In addition, we found that some DRM subfamily members didn’t contain the UBA domain. The transcript abundance ofCpC5-MTaseandCpdMTasein male and female flowers of two morphs (protandry and protogyny) from diploidy was analyzed. Results showed that all genes were significantly up-regulated at the stage of floral bud break (S2), but significantly down-regulated at the stage of flower maturation (S4). At S2, someCpC5-MTasegenes showed higher expression levels in PG-M than in PG-F, whereas someCpdMTasegenes showed higher expression levels in PA-M than in PA-F. In addition, these genes were significantly associated with gibberellin synthesis-related genes (e.g.DELLAandGID1), suggesting that DNA methylation may play a role in the asynchronous floral development process through gibberellin signal.ConclusionsThese results broaden our understanding of theCpC5-MTaseandCpdMTasegenes in diploid and autotetraploidC. paliurus, and provide a novel insight into regulatory mechanisms of DNA methylation in heterodichogamy.Supplementary InformationThe online version contains supplementary material available at 10.1186/s12864-023-09383-5.