Archaeological evidence supports the inhabitation of the Lake Baikal region since the Paleolithic. Both metric and nonmetric osteological studies suggest that Neolithic Cis-Baikal populations are the ancestors of contemporary inhabitants of the region. To date, ancient DNA data have not been used to corroborate this biological continuity hypothesis. This study presents a temporal snapshot of the Cis-Baikal Neolithic by examining mtDNA diversity in two cemetery populations situated on the Angara River downstream of Lake Baikal. The 800 years separating the use of the two cemeteries is thought to represent a biocultural hiatus in the Cis-Baikal region, one that ended when a new group migrated into the area. To assess the likelihood that genetic continuity exists between these two Neolithic groups, we examined both mtDNA coding region and hypervariable region I (HVI) polymorphisms from skeletal remains excavated from both cemeteries (Lokomotiv and Ust'-Ida). The mtDNA haplogroup distributions of the two cemetery populations differ significantly, suggesting that they were biologically distinct groups. When the biological distance between these Neolithic groups is compared with modern Siberian and other East Eurasian groups, the posthiatus group (Serovo-Glazkovo) generally aligns with contemporary Siberians, while the prehiatus (Kitoi) individuals are significantly different from all but modern Kets and Shorians living in the Yenisey and Ob River basins to the west of Lake Baikal. These results suggest that the Lake Baikal region experienced a significant depopulation event during the sixth millennium BP, and was reoccupied by a new immigrant population some 800 years later. Am J Phys Anthropol, 2006. © 2005 Wiley-Liss, Inc.