Mitochondrial DNA (mtDNA) D-loop sequences (381 bp) from 100 thoroughbreds in 19 of the most common matrilineal female families were used to reconstruct a founder female population for the thoroughbred ( approximately 1650-1750 AD). Seventeen haplotypes were found to have contributed to the 19 female lineages. In order to place the reconstructed founder population in wider historical context, we examined, using both single strand conformation polymorphism and direct sequence analysis, variation in a 343 bp mtDNA fragment in that population and 13 other horse populations of disparate provenance. Interpopulation diversity analyses revealed no significant difference in variation between the thoroughbred founder population and the 13 other diverse horse populations and suggested a non-random partitioning of diversity among geographically diverse horse populations. Within thoroughbreds, almost half of the female families, which are each considered from pedigrees to have matrilines converging to one of >30 historically recognized female ancestors, contained sequences which were inconsistent with common descent. Examination of the anomalies in the context of pedigree records suggests the majority might be best explained as confusion of mares at the foundation stages, although some could have some bearing on more recent (19th century - 1980) lineages. We have used this combination of molecular and historical information to identify some of the founder dams and to make new interpretations about the early history of the thoroughbred.