Abstract Background The identification and use of Ancestry-Sensitive Markers (ASMs), i.e. genetic polymorphisms facilitating the genetic reconstruction of geographical origins of individuals, is far from straightforward. Results Here we describe the ascertainment and application of five different sets of 47 single nucleotide polymorphisms (SNPs) allowing the inference of major human groups of different continental origin. For this, we first used 74 cell lines, representing human males from six different geographical areas and screened them with the Affymetrix Mapping 10K assay. In addition to using summary statistics estimating the genetic diversity among multiple groups of individuals defined by geography or language, we also used the program STRUCTURE to detect genetically distinct subgroups. Subsequently, we used a pairwise F_ST ranking procedure among all pairs of genetic subgroups in order to identify a single best performing set of ASMs. Our initial results were independently confirmed by genotyping this set of ASMs in 22 individuals from Somalia, Afghanistan and Sudan and in 919 samples from the CEPH Human Genome Diversity Panel (HGDP-CEPH) Conclusion By means of our pairwise population F_ST ranking approach we identified a set of 47 SNPs that could serve as a panel of ASMs at a continental level.