Intragenomic variability and pseudogenes of ribosomal DNA in stone flounder Kareius bicoloratus.

Molecular study of Stone flounder Kareius bicoloratus revealed a high level of intra-individual polymorphism of ribosomal DNA (18S rDNA, ITS1 and 5.8S rDNA). This polymorphism includes not only multiple functional genes but also putative pseudogenes and recombinants. Three major types (Type I, II and III) of 18S rDNA differing in lengths coexisted in the genome of each studied individual. Type II and Type III have a 15-bp-deletion and Type III has an additional 7-bp-deletion relative to Type I. Expression analyses gave clear evidence that Type I 18S rDNA was abundantly expressed. In contrast, transcription of Type II and III 18S rDNA were not detected. Further sequence analysis revealed that Type I was the expected and functional 18S rDNA in K. bicoloratus, while Type II and Type III were pseudogenes. Phylogeny of the ITS1 sequences formed three groups (Group I, II and III), of which Group I linking with Type I 18S rDNA were functional, whereas the remaining two Groups (II and III) linking with Type II and Type III 18S rDNA were pseudogenes. Groups II and III ITS1 sequences were shorter (685-717bp in length) than Group I ITS1 (762-776bp), but had much higher sequence divergence (9.8%) than the later ones did (4.9%). Group II sequences were recombinants between Group I and Group III sequences, providing the possibility that the rDNA of K. bicoloratus is in a transition stage of concerted evolution. In addition, by comparing standard PCR and high-stringency PCR, we concluded that DMSO might prevent functional ITS1 sequences from being amplified instead of enhancing its amplification efficiency. The analysis suggests a non-concerted evolution of rDNA in K. bicoloratus. Features of rDNA functional genes and pseudogenes are described. Possible causes for the lack of homogenization of rDNA paralogues within individuals are discussed.