Centromeres usually contain large amounts of tandem repeat DNA, which often exceeds the megabase-pair level in size. Alpha satellite DNA is a major centromeric repetitive DNA of simian primates. Humans carry two types of alpha satellite DNA: simple repeats of 171-bp repeat units, and those exhibiting higher-order repeat structures (multiple copies of 171-bp units form a larger repeat unit and the larger units appear periodically). The latter type is known to be evolutionarily new and to play more significant roles in centromere formation. We have recently identified two types of alpha satellite DNA in owl monkeys (genus Aotus; New World monkey). The two types were named OwlAlp1 and OwlAlp2. With respect to the structure, the two types of human alpha satellite DNA differ in repeat organization, but the difference between OwlAlp1 and OwlAlp2 is in the size of their repeat units. The repeat units of OwlAlp1 (185 bp) correspond to part of those of OwlAlp2 (344 bp). All other New World monkeys examined thus far (marmosets, squirrel monkeys, capuchins, tamarins etc.) appear to have only alpha satellite DNA close in size and sequence to OwlAlp2. Thus, OwlAlp1 is considered to be evolutionarily new, derived from OwlAlp2 by partial deletion. OwlAlp1 occupies the centromeric constriction region of all chromosomes, whereas OwlAlp2 is present in pericentric regions of most, but not all, chromosomes, suggesting that OwlAlp1 plays more significant roles. These features support the view that OwlAlp1 replaced OwlAlp2 as the principal centromeric repetitive DNA in the owl monkey lineage after its divergence from those leading to other New World monkeys.