The cross-utility of 12 microsatellite loci (including nine newly developed loci) amongst the viviparous subfamily, the Mexican Goodeinae, was assessed, examining both the probability of amplification and the potential incidence of null alleles from tests of FIS. Genetic diversity was relatively high in comparison to other freshwater species. Amplification success was not correlated with genetic distance between microsatellite source and target species, but taxa that were more distantly related were less likely to be cross-polymorphic for microsatellite loci. On average, species that were cross-polymorphic were separated by a genetic distance of 15% at the cytochrome oxidase I locus, while those that were monomorphic were separated by 19%. There was no evidence that null alleles become more frequent at greater source-target genetic distance.