The Illumina® ForenSeq™ kit amplifies in one multiplex PCR reaction 59 STRs and 172 SNPs relevant to forensic genetic case work. Typed markers include markers for identification, ancestry identification (AIMs) and pigmentary traits. The products were sequenced on the MiSeq FGx Forensic Genomics System and analysed with the ForenSeq™ Universal Analysis Software. Using this approach, we sequenced 10 Danish, 10 Italian, and 10 individuals from other populations, mostly non-europeans. The eye and hair colours as well as the bio-geographical ancestry of the individuals were known.Blue and brown eye colours were predicted accurately by the ForenSeq™ assay, whereas intermediate eye colours were not. The success of the hair colour prediction tool seemed population-dependent. The predictions matched the hair colours reported by the individuals themselves in twice as many Danes as Italians. The geographic ancestries of 22 out of 23 individuals of European or East Asian ancestry were correctly predicted. However, the ForenSeq™ Universal Analysis Software was unable to differentiate between individuals from Middle East/South Asia, South America and North Africa.We found no difference in STR typing results obtained with the ForenSeq™ kit and capillary electrophoresis.