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The question was whether a canyon-shaped reservoir is inhabited by genetically distinct populations of European perch Perca fluviatilis exhibiting different spatial behaviour, or by a single population. The study was conducted in the spring of 2006 at Římov Reservoir, Czech Republic. Epipelagic and bathypelagic perch fry ( EPF and BPF) were captured by trawl net (mesh size 1 × 1.35 mm) controlled by echo sounder, and littoral perch fry and adults ( LPF and LPA) were caught by electrofishing and beach seine (mesh size 1 × 1.35 mm). DNA was isolated in 266 individuals from the reservoir, and phylogenetic analysis based on mt DNA was performed ( n = 18) on BPF, EPF, LPF, and LPA from the reservoir and additional sequences from Central Europe. Six microsatellites were used for differentiating among communities ( n = 266). There was no evidence for genetic divergence in mt DNA sequences. Analyses of microsatellites revealed no statistically significant variations between spatial communities, and confirmed genetic homogeneity among communities. FST values and the number of genetic clusters point to the same conclusion, i.e., the reservoir is inhabited by a single population whose individuals occupy different microhabitats. [ABSTRACT FROM AUTHOR]