Repetitive flanking sequences challenge SSR marker development: a case study in the lepidopteran Melanargia galathea

Microsatellite DNA families (MDF) are stretches of DNA that share similar or identical sequences beside nuclear simple-sequence repeat (nSSR) motifs, potentially causing problems during nSSR marker development. Primers positioned within MDFs can bind several times within the genome and might result in multiple banding patterns. It is therefore common practice to exclude MDF loci in the course of marker development. Here, we propose an approach to deal with multiple primer binding sites by purposefully positioning primers within the detected repetitive element. We developed a new protocol to determine the family type and the primer position in relation to MDFs using the software packages RepARK and RepeatMasker together with an in-house R script. We re-evaluated newly developed nSSR markers for the lepidopteran Marbled White (Melanargia galathea) and explored the implications of our results with regard to published data sets of the butterfly Ephydryas aurinia, the grasshopper Stethophyma...