High somatic instability of a microsatellite locus in a clonal tree, Robinia pseudoacacia

Robinia pseudoacacia L. is a clonal tree species. To investigate a mutation within eight microsatellite loci of R. pseudoacacia, we analyzed DNA samples obtained from different leaf samples within each ramet, leaves from ramets within the genet, and seeds. Of the eight loci, locus Rops15 (AG motif) displayed hypermutability. The mutation rates of Rops15 within each ramet, among ramets within the genet, and offspring were 6.27% (ranging from 0 to 31.1%), 6.11% (from 0 to 25.0%) and 3.78% (from 0 to 10.9%), respectively. The mutation rate increased with allele size (13-71 repeat units). The mutation patterns observed in Rops15 were distinctive in two ways. First, there was a significant bias toward additions over deletions, and both addition and deletion of single repeats were dominant at alleles with lengths less than 232 bp (63 repeats). Second, for the longest allele of 248 bp (71 repeats), the number of losses was higher than the number of gains. These observations suggest that the mutation patterns of microsatellites in R. pseudoacacia may follow a generalized stepwise mutation model, and that the tendency of long alleles to mutate to shorter lengths acts to prevent infinite growth. Finally, the observation of somatic hypermutability at locus Rops15 highlights the need for caution when using highly polymorphic microsatellites for population genetic structure and paternity analysis in tree species.