Y-specific 49a/TaqI haplotypes were determined for 831 individuals drawn from 21 different southern African populations. A total of 31 new haplotypes were observed, some of which contained new alleles or allelic variants. Duplication, in addition to CpG mutation, is implicated in the generation of certain allelic variants. Cluster analysis of genetic distances between the populations, calculated using the 49a/TaqI haplotype frequencies, revealed a basic split between African and non-African populations. Hybrid groups cluster with the caucasoid groups, indicating that male gene flow has occurred from the latter into the former. Clustering of the negroid and Khoisan groups is not what might have been expected from the known linguistic affinities. It is suggested that the 49a/TaqI haplotype analysis of these populations is not sufficiently sensitive to distinguish between many of the populations. The Y-specific 49a/PvuII polymorphism was studied in 127 individuals from southern African populations, and 17 polymorphic fragments ranging in size from 3.6 kb to greater than 48 kb were identified. A total of 53 PvuII haplotypes were observed, corresponding to only 30 TaqI haplotypes. There appears to be poor correlation between the two polymorphisms.