The rapid accumulation of ancient human genomes from various areas and time periods
potentially enables the expansion of studies of biodiversity, biogeography, forensics, population
history, and epidemiology into past populations. However, most ancient DNA (aDNA) data
were generated through microarrays designed for modern-day populations, which are known to
misrepresent the population structure. Past studies addressed these problems by using ancestry
informative markers (AIMs). It is, however, unclear whether AIMs derived from contemporary
human genomes can capture ancient population structures, and whether AIM-finding methods are
applicable to aDNA. Further the high missingness rates in ancient—and oftentimes haploid—DNA
can also distort the population structure. Here, we define ancient AIMs (aAIMs) and develop
a framework to evaluate established and novel AIM-finding methods in identifying the most
informative markers. We show that aAIMs identified by a novel principal component analysis
(PCA)-based method outperform all of the competing methods in classifying ancient individuals
into populations and identifying admixed individuals. In some cases, predictions made using the
aAIMs were more accurate than those made with a complete marker set. We discuss the features of
the ancient Eurasian population structure and strategies to identify aAIMs. This work informs the
design of single nucleotide polymorphism (SNP) microarrays and the interpretation of aDNA results,
which enables a population-wide testing of primordialist theories