Intragenomic variation within ITS1 and ITS2 of freshwater crayfishes (Decapoda: Cambaridae): implications for phylogenetic and microsatellite studies.

Intragenomic variation in ITS1 and ITS2 is known to exit but is widely ignored in phylogenetic studies using these gene regions. The amount of variation in seven crayfish species, including three populations of Orconectes luteus and two of Procambarus clarkii, was assessed by sequencing 3, 5, or 10 clones from the same individuals, for a total of 77 sequences. The ITS1 and ITS2 sequences reported here are some of the longest known, with aligned lengths of 760 and 1,300 bp, respectively. They contain multiple microsatellite insertions, all of which show considerable intragenomic variation in the number of repeat elements. This variation is enough to obscure phylogenetic relationships at the population level, although relationships between species can be estimated. Given the hybridization techniques used to locate microsatellites, multiple-copy regions like ITS1 and ITS2 will be preferentially found if they contain microsatellites, and in these cases the microsatellites will not behave as typical Mendelian markers and could give spurious results.