Although members of genus Daphnia are the most common water invertebrates and are considered as model organisms for many taxonomic, ecological and evolutionary studies their systematics remains unresolved. Here, morphological differentiation and genetic polymorphism between the geographically distant populations of the sister species Daphnia galeata and Daphnia cucullata from the Curonian Lagoon (Russia, Kaliningrad Oblast) and the Novosibirsk Reservoir (Russia, Novosibirsk Oblast) are presented. The divergence between species and their populations was analyzed based on a large set of morphological and several genetic markers. Phylogenetic relationships between species and populations were constructed based on variation in mitochondrial 16S and 12S rRNA genes and nuclear ITS2 rDNA sequences. As result the differentiation of these population based on morphological and genetic data was estimated. Among the most varying morphological characters are the relative head size, head shape, and spine length. The mitochondrial DNA divergence between species was significant and reflected their monophyletic origin. Intraspecific genetic distances of the geographically distant D. galeata and D. cucullata populations are estimated as insignificant.