The cucumber lines, S94 (Northern China open-field type, powdery mildew (PM) susceptible) and S06 (European greenhouse type, PM resistant), and their F(6:7) populations were used to investigate PM resistance under seedling spray inoculation in 2005/Autumn and 2006/Spring. QTL analysis was undertaken based on a constructed molecular linkage map of the corresponding F(6) population using composite interval mapping. A total of four QTLs (pm1.1, pm2.1, pm4.1 and pm6.1) for PM resistance were identified and located on LG 1, 2, 4 and 6, respectively, explaining 5.2%-21.0% of the phenotypic variation. Three consistent QTLs (pm1.1, pm2.1 and pm4.1) were detected under the two test conditions. The QTL pm6.1 was only identified in 2005/Autumn. The total phenotypic variation explained by the QTLs was 52.0% and 42.0% in 2005/Autumn and 2006/Spring, respectively. Anchor markers tightly linked to those loci (<5 cM) could lay a basis for both molecular marker-assisted breeding and map-based gene cloning of the PM-resistance gene in cucumber.