The zebrafish, Danio rerio, has become a major model organism used in biomedical studies. The widespread use of Danio rerio in research laboratories requires a comprehensive understanding of the husbandry of this species to ensure efficient propagation and maintenance of healthy and genetically diverse colonies. Breeding is a key element in zebrafish husbandry. It is a complex process influenced by a number of factors. Mate choice and mating behavior depend, for example, on olfactory cues, visual stimuli, and social interactions. Spawning is affected by the age and size of fish, interval at which fish are used for egg production, light cycle, diet, and fish health status. A number of breeding strategies, based on either single-pair matings or group crosses, are commonly employed in the laboratory to propagate lines and to identify carriers of specific mutations and/or transgenes. Propagation of zebrafish lines, in particular wild-type-derived strains, is closely monitored to ensure that genetic diversity and vigor are maintained. A robust zebrafish line typically carries a large number of polymorphic variations, which may interfere with reproducibility of experiments. To get a better insight into these variations, a wild-type hybrid Sanger AB Tübingen line has been generated from sequenced homozygous founders.