In this study, we examined for the first time the distribution of the 28S ribosomal genes in beetles of the subfamily Cassidinae. More than 55% of the species in this subfamily have a similar karyotype, 2n = 16 + Xyp. For this work, we selected species belonging to the tribes Cassidini and Mesomphaliini, which have, respectively, the most conserved and diversified karyotype characteristics within the Cassidinae. An analysis of 11 species revealed that rDNA sites on one pair of autosomes is the most frequent pattern, occurring in 10 species. This condition occurs in the seven genera examined and in species of both of the tribes, Cassidini and Mesomphaliini. Nevertheless, the differences in the locations of 28S rDNA were more pronounced in the tribe Cassidini and among species with similar karyotype characteristics. On the other hand, in Mesomphaliini, the increase in the diploid number was not accompanied by an increase in the number of ribosomal sites. Moreover, the comparison of the number and localization of major rDNA sites with the distribution of constitutive heterochromatin indicates that there is no direct correlation between the dispersion of constitutive heterochromatin and 28S rDNA genes in Cassidinae.