Sequence variation of the internal transcribed spacer (ITS) region of ribosomal DNA in Cerastoderma species (Bivalvia: Cardiidae)

摘要

The internal transcribed spacer (ITS) region of ribosomal DNA (ITS1, 5.8S rRNA gene and ITS2) constitutes one of the most widely applied molecular markers in phylogenetic studies and species differentiation. Here, the ITS region of the cockles Cerastoderma edule from NW Spain and C. glaucum from the Mediterranean and Baltic Seas was characterized and its variation examined. The length of the ITS was 762–783 bp (ITS1, 226–251 bp; 5.8S rRNA gene, 158–161 bp; ITS2, 305–325 bp) and the GC content ranged between 57% and 60% (ITS1, 52–62%; 5.8S rRNA gene, 56–57%; ITS2, 61–63%). All individuals showed variation among ITS repeats, mostly in the spacers, with nucleotide diversity values of 0.0005–0.07. Among the C. glaucum sequences, two types of ITS (ITSI and ITSII) were distinguished, based on the percentage of differences observed in the spacers, sequence of the 5.8S gene, GC content of the ITS1 and clustering in phylogenetic trees. A PCR assay using specific primers for each ITS type demonstrated that both types coexist in all C. glaucum individuals. While type I seems to contain the bona fide 5.8S rRNA gene, type II could contain a pseudogene. Cerastoderma edule sequences grouped separately on phylogenetic trees, as expected for a differentiated species, but those of C. glaucum from the Mediterranean and Baltic Seas were intermixed in the two clades defining the ITS types, which supports the absence of systematic division previously inferred from other molecular markers.