Prediction of eye and hair color from DNA is being increasingly employed in forensics and the studies of ancient populations. HIrisPlex-S is a prediction tool trained on the Dutch dataset and verified using a few other European populations. The accuracy of its predictions for other regions of the world has not been studied yet. Russian populations pose a special interest because, unlike the majority of world populations, their representatives can have not only dark but also light color eyes and hair. The aim of this work was to evaluate the accuracy of eye and hair color prediction in Russian populations. We recruited 144 representatives of indigenous peoples of Russia (Avar, Aleut, Buryat, Itelmen, Karelian, Koryak, Mari, Nanai, Russian, Rutulian, Chuvash, Chukchi, Evenk, and Even populations). All study participants were photographed. Eye and hair colors were identified from the anthropological images by anthropologists. The SNP markers included in the HIrisPlex system were genotyped. Phenotypes were predicted from the obtained genotypes and subsequently compared to the actual phenotypes. Quality metrics were calculated for HIrisPlex prediction accuracy in the populations of European Russia and Siberia. On the whole, HIrisPlex prediction accuracy was satisfactory, although a bit lower than in Western European datasets. Further research could focus on identifying additional markers to improve the accuracy of predictions in Russian populations.