Population demarcation of eight horse breeds was investigated using genotype information of 306 horses from 26 microsatellite loci. The breeds include the indigenous Norwegian breeds Fjord Horse, Nordland/Lyngen Horse, Døle Horse and Coldblooded Trotter together with Icelandic Horse, Shetland Pony, Standardbred and Thoroughbred. Both phylogenetic analysis and a maximum likelihood method were applied to examine the potential for breed allocation of individual animals. The phylogenetic analysis utilizing simple allele sharing statistics revealed clear demarcation among the breeds; 95% of the individuals clustered together with animals of the same breed in the phylogenetic tree. Even breeds with a short history of divergence like Døle Horse and Coldblooded Trotter formed distinct clusters. Implementing the maximum likelihood method allocated 96% of the individuals to their source population, applying an assignment stringency of a log of the odds ratio larger than 2. Lower allocation stringency assigned nearly all the horses. Only three individuals were wrongly allocated a breed by both methods. In conclusion, the study demonstrates clear distinction among horse breeds, and by combining the two assignment methods breed allocation could be determined for more than 99% of the individuals.