Asian soybean rust (ASR), caused byPhakopsora pachyrhizi, is one of the most serious soybean (Glycine max) diseases in tropical and subtropical regions. To facilitate the development of resistant varieties using gene pyramiding, DNA markers closely linked to seven resistance genes, namely,Rpp1,Rpp1-b,Rpp2,Rpp3,Rpp4,Rpp5, andRpp6, were identified. Linkage analysis of resistance-related traits and marker genotypes using 13 segregating populations of ASR resistance, including eight previously published by our group and five newly developed populations, identified the resistance loci with markers at intervals of less than 2.0 cM for all seven resistance genes. Inoculation was conducted of the same population with twoP. pachyrhiziisolates of different virulence, and two resistant varieties, ‘Kinoshita’ and ‘Shiranui,’ previously thought to only harborRpp5, was found to also harborRpp3. Markers closely linked to the resistance loci identified in this study will be used for ASR-resistance breeding and the identification of the genes responsible for resistance.