Abstract Internal transcribed spacer 2 (ITS2) sequences of the nuclear rDNA in forty‐seven specimens (thirty‐four species) of the leaf beetle genus Timarcha have been studied. Timarcha ITS2 (523 bp on average) share some sequence features with other Chrysomeloidea relatives ( Chrysolina , Diabrotica and Bruchus ) but have no clear similarity with any other arthropod ITS2 sequences. Interspecific divergences are in the range 0.002–0.166, and 0.124–0.206 in the comparisons between subgenera. No evidence of intragenomic divergent ITS2 sequences has been found. Secondary structures are concordant with the four‐domain model proposed for vertebrates and yeast, but differs from those proposed for dipterans. Phylogenetic analysis of the ITS2 data confirms the results of a previous study based on mitochondrial sequences, as the basality of the Metallotimarcha subgenus and the absence of phylogenetic support for the Timarchostoma subgenus.