The aim of this study was to characterize the fresh and cryopreserved whitefish semen and to verify the usefulness of fresh semen parameters in order to qualify the cryopreserved semen. The relationship between DNA microsatellite genotyping and quality parameters of fresh and cryopreserved semen was also examined. Whitefish semen was characterized by high quality parameters values, mainly sperm motility and viability. After cryopreservation an intense decrease of the percentage of motile cells was noticed whereas viability decreased in a lower degree. Low regressions were found between quality parameters for both fresh and cryopreserved semen, except for the regression between the amplitude of lateral head displacement and curvilinear velocity. A significant but low regression was found between the percentage of heterozygotic microsatellite loci and the percentage of sperm motility of fresh semen and seminal plasma osmolality. Differences in sperm motility and linearity of cryopreserved semen between homozygotes and heterozygotes at locus Sfo-292A (P < 0.05) were noticed. The parameters of fresh semen demonstrated little usefulness for predicting the quality of cryopreserved whitefish semen. Genetic characterization of male whitefish by microsatellite variability seems to be a promising tool for the study of genetic background of male reproduction in fish.