Genome size and GC‐percent were determined by means of a special method of DNA flow cytometry in 154 vertebrate species. For the total dataset, a highly significant positive correlation was found between both parameters. The overall distribution of points is not linear but triangular: a wide range of GC‐percent values is observed at the lower end of genome size range, whereas with an increase in genome size the lower limit for GC‐percent is elevated, gradually approaching the upper limit (about 46%). In teleost fishes, which occupy the lower part of genome size range, the negative relationship between both parameters was observed. Two positive linear relationships were found between mean genome size and GC‐percent of the main vertebrate groups (one includes fishes, amphibians, and mammals, the other consists of reptiles and birds, which show the higher GC‐percent for their genome sizes). Distribution of variance between taxonomic levels indicates that GC‐percent is more evolutionarily conservative than genome size in anamniotes. Anuran amphibians show the greatest part of genome size variability at the lower taxonomic levels as compared to other vertebrates (with no additional variance already above the genus level). The data obtained with different methods are compared. It is shown that the proposed method can provide useful data for studies on genome evolution and biodiversity. Cytometry 31:100–109, 1998.