An A in equilibrium with G transition in exon III is known to differentiate alleles A and B of the cattle beta-lactoglobulin (BLG) gene. A BLG exon III fragment containing the transition site was amplified by the polymerase chain reaction. Temperature gradient gel electrophoresis (TGGE) was then used to detect this transition and hence to genotype cattle: the AT base-pair in allele A was readily distinguished from the GC base-pair of allele B. TGGE can be used to detect any single base-pair substitution, and thus is a powerful method of detecting genetic variability.