Soybean Mosaic Virus (SMV) is a prevalent viral pathogen transmitted by aphids and via seed, causing significant yield loss and quality reduction. There are seven SMV strain groups (G1 to G7) and three resistance loci (Rsv1, Rsv3, and Rsv4) reported in soybean. New sources of SMV resistance would be valuable for breeding cultivars with durable resistance to multiple strains. The objectives of this research were to pyramid three genes from different sources for durable SMV resistance; and to identify and map new genes/alleles for differential reactions to SMV strains. PI96983 (Rsv1) was crossed with PI548317 (Rsv3, Rsv4) to pyramid all three SMV resistance genes with the aid of SSR markers. The molecular screening of a F₂ plant population derived from PI96983 x PI548317 showed good fit to the expected genotypic ratios at each marker locus, and 2 out 68 plants were identified to contain three resistance genes (Rsv1+Rsv3+Rsv4). A new SMV resistant line PI61944 with unknown gene(s) (Rsv?) was crossed with Essex (rsv), PI96983 (Rsv1), L29 (Rsv3), and V94-5152 (Rsv4) to investigate the inheritance and allelism of SMV resistance. F₂ plants were screened with selected SSR markers, and the F_2:3 lines were screened with SMV G1 and G7. SMV screening and molecular analysis revealed that the PI61944 carries a new allele at the Rsv3 locus. This research has discovered a new SMV resistance allele, and combined three genes for durable resistance in soybean. These germplasm will be useful in soybean breeding programs where SMV resistance is an objective. This research also demonstrated that SSR technology is extremely useful in marker assisted selection (MAS).