Development of novel microsatellite markers for a specialist species of Lepidoptera, Boloria aquilonaris (Nymphalidae), based on 454 sequences

Microsatellites are the most common markers used in population and conservation genetic studies. However, their isola- tion is laborious and expensive. In some taxa, such as Lepidoptera, it is particularly difficult to isolate microsatellite markers due to the high similarity of the flanking regions of different loci and the presence of null alleles. Here we isolated microsatellites of the endangered butterfly Boloria aquilonaris using 454 GS-FLX Titanium pyro-sequences of biotin enriched DNA libraries and tested the success of cross-amplification on the sister-species B. eunomia. Fifteen polymorphic microsatellite loci were isolated in B. aquilonaris using initially 101 stringently designed primer pairs. Unlike in many other studies of microsatellite isolation in Lepidoptera, few null alleles were detected and only at very low frequencies. Additionally, the raw data set can still be used for the isolation of other microsatellite loci. None of the selected polymorphic loci for B. aquilonaris gave clear banding patterns for B. eunomia, although about 15 other loci gave promising banding patterns for the latter species. Low intra- and inter-specific transferability of developed markers in this study also lends support to the hypothesis that the evolution of the genome of Lepidoptera is dissimilar from that of other organisms.