The Potato virus Y (PVY) is distributed worldwide and is one of the most damaging viruses in terms of yield reduction in the potato (Solanum tuberosum L.) crop with losses sometimes reaching 80%. To reduce its impact, there is great interest in obtaining varieties carrying the Ryadg gene that provides extreme resistance to this virus. This could be facilitated with SCAR (sequence characterized amplified region) RYSC3 by molecular marker-assisted selection (MAS). This study compared the effectiveness of the RYSC3 marker in the detection of the Ryadg gene vs. biological tests on populations of the Potato Breeding Program of the Instituto de Investigaciones Agropecuarias (INIA), Chile. Within the group of 71 progenitors, 30 plants had some kind of resistance to the virus, of which 17 were carriers of the RYSC3 marker. These genotypes came from Cornell University, Centro Internacional de la Papa (CIP), Peru, and INIA. The analysis of 460 progenies which came from three different crosses showed that 299 individuals amplified the RYSC3 marker and had the resistant phenotype, with the exception of one plant. Within the group of non-RYSC3 carrier plants, a significant percentage (22.5%) showed a resistant phenotype, indicating that these progenies segregate other R genes (e.g., hypersensitivity) that reduce biological test effectiveness. This high effectiveness (99.7%) in the detection of the Ryadg gene in both parents and segregating progenies, showed that this marker is appropriate in assisting selection of genotypes with extreme PVY-resistance in the potato breeding programs.