Validated microsatellite markers for Gyrodactylus salaris: a toolkit for individual identification and genetic studies

Abstract

Microsatellite markers remain essential for individual-level genetic work in taxa where genome-wide methods are not yet routinely feasible due to extremely low DNA yields per specimen. In Gyrodactylus, even the most recent reference genomes have required pooling thousands of individuals, leaving a practical gap between genome-scale resources and individual-level analyses. Here we present a genome-informed microsatellite panel, developed by selecting single-copy loci with non-repetitive flanking regions and assembling all markers into a single multiplex PCR. Marker identity and performance were verified via amplification tests, Sanger sequencing, and cross-laboratory genotyping, confirming that the same samples generated identical fragment-size profiles in both laboratories. Long tandem repeats occasionally prevented exact repeat-count determination, yet allele-size classes were discrete and reproducible across replicates. The panel enables rapid individual identification and reliable strain and lineage assignment. It also offers a practical starting point for population-genetic and evolutionary studies that require individual-level data.