Downy mildew in cucumber (Cucumis sativus L.) is a disease caused by Psuedoperonospora cubensis (Berk. and Curt.). Genetic resistance is used to control this disease. However, selection for resistance to downy mildew requires considerable effort. Molecular markers that are closely linked to a gene for downy mildew resistance may be useful for increasing selection efficiency. RAPD (randomly amplified polymorphic DNA) markers are molecular markers that have several advantages including economy, abundance and high rate polymorphism. Two populations, WI 1983 × Straight 8 and ZuDM1 × Straight 8, were used to identify RAPD markers linked to a downy mildew resistance gene. The WI 1983 × Straight 8 population contained 63 F3 families and the ZuDM1 × Straight 8 population contained 90. These F3 families were evaluated for disease reactions in four locations—Wisconsin, South Carolina, Spain, and The Netherlands. Four replications were conducted in Wisconsin and two in the remaining locations. The ranking of susceptible and resistant families was consistent over the locations. Spearman rank coefficients between locations ranged from 0.65 to 0.84. DNA samples from individual F2 plants in each population were combined into two bulks, one resistant and one susceptible, based on the family performance. Then, PCR analysis was performed. Over 100 polymorphic primers were screened using this bulked segregant analysis to find the resistance gene to RAPD genetic linkage.