Multiplex single nucleotide polymorphism (SNP) assay for detection of soybean mosaic virus resistance genes in soybean

Soybean mosaic virus (SMV) is one of the most destructive viral diseases in soybean (Glycine max). Three independent loci for SMV resistance have been identified in soybean germplasm. The use of genetic resistance is the most effective method of controlling this disease. Marker assisted selection (MAS) has become very important and useful in the effort of selecting genes for SMV resistance. Single nucleotide polymorphism (SNP), because of its abundance and high-throughput potential, is a powerful tool in genome mapping, association studies, diversity analysis, and tagging of important genes in plant genomics. In this study, a 10 SNPs plus one insert/deletion (InDel) multiplex assay was developed for SMV resistance: two SNPs were developed from the candidate gene 3gG2 at Rsv1 locus, two SNPs selected from the clone N11PF linked to Rsv1, one ‘BARC’ SNP screened from soybean chromosome 13 [linkage group (LG) F] near Rsv1, two ‘BARC’ SNPs from probe A519 linked to Rsv3, one ‘BARC’ SNP from chromosome 14 (LG B2) near Rsv3, and two ‘BARC’ SNPs from chromosome 2 (LG D1b) near Rsv4, plus one InDel marker from expressed sequence tag (EST) AW307114 linked to Rsv4. This 11 SNP/InDel multiplex assay showed polymorphism among 47 diverse soybean germplasm, indicating this assay can be used to investigate the mode of inheritance in a SMV resistant soybean line carrying Rsv1, Rsv3, and/or Rsv4 through a segregating population with phenotypic data, and to select a specific gene or pyramid two or three genes for SMV resistance through MAS in soybean breeding program. The presence of two SMV resistance genes (Rsv1 and Rsv3) in J05 soybean was confirmed by the SNP assay.     

Detection and fine-mapping of SC7 resistance genes via linkage and association analysis in soybean

Soybean mosaic virus (SMV) disease is one of the most serious and broadly distributed soybean (Glycine max (L.) Merr.) diseases. Here, we combine the advantages of association and linkage analysis to i...     

Inheritance and allelism of resistance to soybean mosaic virus in Zao18 soybean from China

Soybean mosaic disease caused by soybean mosaic virus (SMV) occurs wherever soybean [Glycine max (L.) Merr.] is grown and is considered one of the most important soybean diseases in many areas of the world. Use of soybean cultivars with resistance to SMV is a very effective way of controlling the disease. China has rich soybean germplasm, but there is very limited information on genetics of SMV resistance in Chinese soybean germplasm and reaction of the resistance genes to SMV strains G1-G7. There also is no report on allelic relationships of resistance genes in Chinese soybeans with other named genes at the three identified loci Rsv1, Rsv3, and Rsv4. The objectives of this study were to examine reactions of Chinese soybean cultivar Zao18 to SMV strains G1-G3 and G5-G7, to reveal the inheritance of SMV resistance in Zao18 and to determine the allelic relationship of resistance genes in Zao18 with previously reported resistance genes. Zao18 was crossed with the SMV-susceptible cultivar Lee 68 to study the inheritance of resistance. Zao18 was also crossed with the resistant lines PI96983, L29, and V94-5152, which possess Rsv1, Rsv3, and Rsv4, respectively, to examine the allelic relationship between the genes in Zao18 and genes at these three loci. Our research results indicated that Zao18 possesses two independent dominant genes for SMV resistance, one of which is allelic to the Rsv3 locus; the other is allelic with Rsv1. The presence of both genes (Rsv1 and Rsv3) in Zao18 confers resistance to SMV strains G1-G7.     

Pyramiding multiple genes for resistance to soybean mosaic virus in soybean using molecular markers

Seven strains of Soybean mosaic virus (SMV) and three independent resistance loci (Rsv1, Rsv3, and Rsv4) have been identified in soybean. The objective of this research was to pyramid Rsv1, Rsv3, and Rsv4 for SMV resistance using molecular markers. J05 carrying Rsv1 and Rsv3 and V94-5152 carrying Rsv4 were used as the donor parents for gene pyramiding. A series of F_2:3, F_3:4, and F_4:5 lines derived from J05 × V94-5152 were developed for selecting individuals carrying all three genes. Eight PCR-based markers linked to the three SMV resistance genes were used for marker-assisted selection. Two SSR markers (Sat_154 and Satt510) and one gene-specific marker (Rsv1-f/r) were used for selecting plants containing Rsv1; Satt560 and Satt063 for Rsv3; and Satt266, AI856415, and AI856415-g for Rsv4. Five F_4:5 lines were homozygous for all eight marker alleles and presumably carry all three SMV resistance genes that would potentially provide multiple and durable resistance to SMV.     

Fine mapping and analyses of <Emphasis Type="Italic">R</Emphasis><Subscript><Emphasis Type="Italic">SC8</Emphasis></Subscript> resistance candidate genes to soybean mosaic virus in soybean

Soybean mosaic virus (SMV) in soybean [Glycine max (L.) Merr.] is a destructive foliar disease in soybean-producing countries worldwide. In this study, F₂, F_2:3, and F_7:11 recombinant inbred lines populations derived from Kefeng No.1 × Nannong 1138-2 were used to study inheritance and linkage mapping of the SMV strain SC8 resistance gene in Kefeng No.1. Results indicated that a single dominant gene (designated R _SC8 ) controls resistance, which is located on chromosome 2 (MLG D1b). A mixed segregating population was developed by selfing two heterozygous plants (RHL153-1 and RHL153-2) at four markers adjacent to the locus and used in fine mapping R _SC8 . In addition, two genomic-simple sequence repeats (SSR) markers BARCSOYSSR_02_0610 and BARCSOYSSR_02_0616 were identified that flank the two sides of R _SC8 . Sequence analysis of the soybean genome indicated that the interval between the two genomic-SSR markers is 200 kb. QRT-PCR analysis of the candidate genes determined that five genes (Glyma02g13310, 13320, 13400, 13460, and 13470) are likely involved in soybean SMV resistance. These results will have utility in cloning, transferring, and pyramiding of the R _SC8 through marker-assisted selection in soybean breeding programs.     

RSC3K of soybean cv. Kefeng No.1 confers resistance to soybean mosaic virus by interacting with the viral protein P3

Soybean mosaic virus (SMV) is one of the most devastating viral pathogens of soybean (Glycine max (L.) Merr). In total, 22 Chinese SMV strains (SC1–SC22) have been classified based on the responses of 10 soybean cultivars to these pathogens. However, although several SMV-resistance loci in soybean have been identified, no gene conferring SMV resistance in the resistant soybean cultivar (cv.) Kefeng No.1 has been cloned and verified. Here, using F₂-derived F₃ (F_2:3) and recombinant inbred line (RIL) populations from a cross between Kefeng No.1 and susceptible soybean cv. Nannong 1138-2, we localized the gene in Kefeng No.1 that mediated resistance to SMV-SC3 strain to a 90-kb interval on chromosome 2. To study the functions of candidate genes in this interval, we performed Bean pod mottle virus (BPMV)-induced gene silencing (VIGS). We identified a recombinant gene (which we named R_SC3K) harboring an internal deletion of a genomic DNA fragment partially flanking the LOC100526921 and LOC100812666 reference genes as the SMV-SC3 resistance gene. By shuffling genes between infectious SMV DNA clones based on the avirulent isolate SC3 and virulent isolate 1129, we determined that the viral protein P3 is the avirulence determinant mediating SMV-SC3 resistance on Kefeng No.1. P3 interacts with RNase proteins encoded by R_SC3K, LOC100526921, and LOC100812666. The recombinant R_SC3K conveys much higher anti-SMV activity than LOC100526921 and LOC100812666, although those two genes also encode proteins that inhibit SMV accumulation, as revealed by gene silencing in a susceptible cultivar and by overexpression in Nicotiana benthamiana. These findings demonstrate that R_SC3K mediates the resistance of Kefeng No.1 to SMV-SC3 and that SMV resistance of soybean is determined by the antiviral activity of RNase proteins.     

Construction and comparison of three reference‐quality genome assemblies for soybean

We report reference‐quality genome assemblies and annotations for two accessions of soybean (Glycine max) and for one accession of Glycine soja, the closest wild relative of G. max. The G. max assemblies provided are for widely used US cultivars: the northern line Williams 82 (Wm82) and the southern line Lee. The Wm82 assembly improves the prior published assembly, and the Lee and G. soja assemblies are new for these accessions. Comparisons among the three accessions show generally high structural conservation, but nucleotide difference of 1.7 single‐nucleotide polymorphisms (snps) per kb between Wm82 and Lee, and 4.7 snps per kb between these lines and G. soja. snp distributions and comparisons with genotypes of the Lee and Wm82 parents highlight patterns of introgression and haplotype structure. Comparisons against the US germplasm collection show placement of the sequenced accessions relative to global soybean diversity. Analysis of a pan‐gene collection shows generally high conservation, with variation occurring primarily in genomically clustered gene families. We found approximately 40–42 inversions per chromosome between either Lee or Wm82v4 and G. soja, and approximately 32 inversions per chromosome between Wm82 and Lee. We also investigated five domestication loci. For each locus, we found two different alleles with functional differences between G. soja and the two domesticated accessions. The genome assemblies for multiple cultivated accessions and for the closest wild ancestor of soybean provides a valuable set of resources for identifying causal variants that underlie traits for the domestication and improvement of soybean, serving as a basis for future research and crop improvement efforts for this important crop species.     

Molecular sequence variations of the lipoxygenase-2 gene in soybean

Soybean lipoxygenase genes comprise a multi-gene family, with the seed lipoxygenase isozymes LOX1, LOX2, and LOX3 present in soybean seeds. Among these, the LOX2 isozyme is primarily responsible for the “beany” flavor of most soybean seeds. The variety, Jinpumkong 2, having null alleles (lx1, lx2, and lx3) lacks the three seed lipoxygenases; so, sequence variations between the lipoxygenase-2 genes of Pureunkong (Lx2) and Jinpumkong 2 (lx2) cultivars were examined. One indel, four single nucleotide polymorphisms (SNPs), a 175-bp fragment in the 5′-flanking sequence, and a missense mutation within the coding region were found in Jinpumkong 2. The distribution of the sequence variations was investigated among 90 recombinant inbred lines (RILs) derived from a cross of Pureunkong × Jinpumkong 2 and in 480 germplasm accessions with various origins and maturity groups. Evidence for a genetic bottleneck was observed: the 175-bp fragment was rare in Glycine max, but present in the majority of the G. soja accessions. Furthermore, the 175-bp fragment was not detected in the 5′ upstream region of the Lx2 gene on chromosome (Chr) 13 in Williams 82; instead, a similar 175-bp fragment was positioned in the homeologous region on Chr 15. The findings indicated that the novel fragment identified was originally present in the Lx2 region prior to the recent genome duplication in soybean, but became rare in the G. max gene pool. The missense mutation of the conserved histidine residue of the lx2 allele was developed into a single nucleotide-amplified polymorphism (SNAP) marker. The missense mutation showed a perfect correlation with the LOX2-lacking phenotype, so the SNAP marker is expected to facilitate breeding of soybean cultivars which lack the LOX2 isozyme.     

Genetic diversity and association mapping in a collection of selected Chinese soybean accessions based on SSR marker analysis

For broadening the narrow genetic base of modern soybean cultivars, 159 accessions were selected from the Chinese soybean collection which contained at least one of seven important agronomic traits: resistance to soybean cyst nematode (SCN) or soybean mosaic virus (SMV), tolerance to salt, cold, or drought, high seed oil content or high protein content. Genetic diversity evaluation using 55 microsatellite loci distributed across the genome indicated that a large amount of genetic diversity (0.806) and allelic variation (781) were conserved in this selected set, which captured 65.6% of the alleles present in Chinese soybean collection (1,863 accessions). On average, 9.4 rare alleles (frequency <5%) per locus were present, which were highly informative. Using model-based Bayesian clustering in STRUCTURE we distinguished four main clusters and a set of accessions with admixed ancestry. The four clusters reflected different geographic regions of origin of the accessions. Since the clusters were also clearly different with respect to the seven agronomic traits, the inferred population structure was introduced when association analysis was conducted. A total of 21 SSR markers on 16 chromosomes were identified as significantly (P < 0.01) associated with high oil content (6), high protein content (1), drought tolerance (5), SCN resistance (6) and SMV resistance (3). Twelve of these markers were located in or near previously identified quantitative trait loci (QTL). The results for both genetic relationship and trait-related markers will be useful for effective conservation and utilization of soybean germplasm.     

Mapping of SMV resistance gene Rsc-7 by SSR markers in soybean

Soybean mosaic virus (SMV) is one of the most prevalent pathogens that limit soybean production. In this study, segregation ratios of resistant plants to susceptible plants in P₁, P₂, F₁, F₂ populations of Kefeng No. 1 (P₁)×Nannong 1138-2 (P₂) and derived RIL populations, were used to study the inheritance of resistance to the SMV strain SC-7. Populations Kefeng No. 1 and F₁ were found to be completely resistant to this SMV strain while Nannong 1138-2 was susceptible to it. The F₂ and RIL populations segregated to fit a ratio of 3:1 and 1:1for resistant plants to susceptible ones, respectively. These results indicated that a single dominant gene, designated as Rsc-7, controlled resistance to the SMV strain SC-7 in Kefeng No.1. SSR markers were used to analyze the RIL population and MAPMAKER/EXP 3.0b was employed to establish linkage between markers and this resistance gene. Combining the data of SSRs and resistance identification, a soybean genetic map was constructed. This map, covering 2625.9 cM of the genome, converged into 24 linkage groups, consisted of 221 SSR markers and the resistance gene Rsc-7. The Rsc-7 gene was mapped to the molecular linkage group G8-D1b+W. SSR markers Satt266, Satt634, Satt558, Satt157, and Satt698 were found linked to Rsc-7 with distances of 43.7, 18.1, 26.6, 36.4 and 37.9 cM, respectively.     

Fitness penalty in susceptible host is associated with virulence of Soybean mosaic virus on Rsv1‐genotype soybean: a consequence of perturbation of HC‐Pro and not P3

The multigenic Rsv1 locus in the soybean plant introduction (PI) ‘PI96983’ confers extreme resistance against the majority of Soybean mosaic virus (SMV) strains, including SMV‐N, but not SMV‐G7 and SMV‐G7d. In contrast, in susceptible soybean cultivars lacking a functional Rsv1 locus, such as ‘Williams82’ (rsv1), SMV‐N induces severe disease symptoms and accumulates to a high level, whereas both SMV‐G7 and SMV‐G7d induce mild symptoms and accumulate to a significantly lower level. Gain of virulence by SMV‐N on Rsv1‐genotype soybean requires concurrent mutations in both the helper‐component proteinase (HC‐Pro) and P3 cistrons. This is because of the presence of at least two resistance (R) genes, probably belonging to the nucleotide‐binding leucine‐rich repeat (NB‐LRR) class, within the Rsv1 locus, independently mediating the recognition of HC‐Pro or P3. In this study, we show that the majority of experimentally evolved mutational pathways that disrupt the avirulence functions of SMV‐N on Rsv1‐genotype soybean also result in mild symptoms and reduced accumulation, relative to parental SMV‐N, in Williams82 (rsv1). Furthermore, the evaluation of SMV‐N‐derived HC‐Pro and P3 chimeras, containing homologous sequences from virulent SMV‐G7 or SMV‐G7d strains, as well as SMV‐N‐derived variants containing HC‐Pro or P3 point mutation(s) associated with gain of virulence, reveals a direct correlation between the perturbation of HC‐Pro and a fitness penalty in Williams82 (rsv1). Collectively, these data demonstrate that gain of virulence by SMV on Rsv1‐genotype soybean results in fitness loss in a previously susceptible soybean genotype, this being a consequence of mutations in HC‐Pro, but not in P3.     

Development of SNP markers and haplotype analysis of the candidate gene for rhg1, which confers resistance to soybean cyst nematode in soybean

Soybean cyst nematode (SCN; Heterodera glycines Ichinohe) is one of the most destructive pests in the cultivation of soybean (Glycine max (L.) Merr.) worldwide. Markers based on the SCN resistance gene will enable efficient marker-assisted selection (MAS). We sequenced the candidate gene rhg1 in six resistant and two susceptible soybean genotypes and identified 37 SNPs (single nucleotide polymorphisms) among the sequences, of which 11 were in the coding region. Seven of these 11 SNPs led to changes in the amino acid sequence of the gene. The amino acid sequence we obtained differs from the previously published one by a stretch of 26–27 amino acids. Six codominant allele-specific SNP markers based on agarose gel detection were developed and tested in 70 genotypes, among which occurred only nine different haplotypes. Two neutrality tests (Tajima’s D and Fu and Li’s F) were significant for the six SNP loci in the 70 genotypes, which is consistent with intensive directional selection. A strong LD pattern was detected among five SNPs except 2868T > C. Two SNPs (689C > A and 757C > T) formed one haplotype (689C-757C) that was perfectly associated with SCN resistance. The new allele-specific PCR markers located in the alleged sequence of the rhg1 candidate gene, combined with the microsatellite marker BACR-Satt309, will significantly improve the efficiency of MAS during the development of SCN-resistant cultivars.     

Divergence and allelomorphic relationship of a soybean virus resistance gene based on tightly linked DNA microsatellite and RFLP markers

The use of genetically diverse resistance sources is important in breeding for durable disease resistance. Detection and evaluation of resistance genes by conventional inheritance experiments, however, often require laborious screening and genetic testing. In the present study, a marker-assisted screening for resistance sources was initiated in soybean [Glycine max (L.) Merr] using one DNA microsatellite and two RFLP markers tightly linked to a soybean mosaic virus (SMV) resistance gene (Rsv1). The three marker loci were used to screen 67 diverse soybean cultivars, breeding lines, and plant introductions. Five variants were found at the microsatellite locus (HSP176L), and the two RFLP loci (pA186 and pK644a) near Rsv1 show a remarkably higher level of restriction polymorphism than Rsv1-independent RFLP loci. Several specific variants at the three marker loci were found to be correlated with virus resistance, among which HSP176L-2 can be detected by PCR, thus may be useful for germplasm screening. The grouping of the 67 accessions according to their multilocus marker variants agrees with the available pedigree information. When all, or most, of the cultivars within a given group with the same Rsv1-linked marker variant are resistant, their SMV resistance is most likely conferred by Rsv1. These putatively Rsv1-carrying groups contain a total of 38 SMV-resistant lines including six differential cultivars that are known to carry Rsv1. The remaining seven resistant accessions (Columbia, Holladay, Peking, Virginia, FFR-471, PI 507403, and PI 556949) do not carry resistance marker variants, and at least some of them could be sources of resistance genes independent of Rsv1.     

Single nucleotide polymorphism markers for rapid detection of the <Emphasis Type="Italic">Rsv4</Emphasis> locus for soybean mosaic virus resistance in diverse germplasm

Soybean mosaic virus (SMV) causes a substantial decrease in soybean yield and reduction of seed quality. The most effective management strategy to control the virus is the deployment of host resistance. Seven SMV strains and three independent multi-allelic loci for SMV resistance have been identified previously. The goal of this research was to detect single nucleotide polymorphisms (SNPs) associated with SMV resistance at the Rsv4 locus. Ten soybean accessions, with confirmed resistance genes, were used for sequencing the candidate gene Glyma.02g121400. Alignment of these sequences revealed three SNPs displaying 100% consistency for genotypes carrying the Rsv4 gene. These SNPs were applied for a rapid screen of diverse soybean germplasm using the Sequenom iPLEX Gold platform, phenotyped with SMV-G1 and G7 strains to determine phenotype and classified into several groups carrying the proposed R-gene. The population of V94-5152 (Rsv4) × Lee 68 (rsv) was screened using novel SNPs to create a genetic map with improved resolution to determine the location of the Rsv4. To observe the recombination frequencies within the population, three additional SNPs on both sides of the Glyma.02g121400 gene were added. A linkage map revealed a distance of 3.6 cM between the Rsv4 locus and the closest SNP, thus shifting the putative Rsv4 region downstream on chromosome 2. With this region, five candidate genes have been proposed. The genomic position of the discovered SNPs, linked to the Rsv4, could increase screening precision and accelerate breeding efforts to develop multi-strain-resistant crops.     

Soybean RNA interference lines silenced for eIF4E show broad potyvirus resistance

Soybean mosaic virus (SMV), a potyvirus, is the most prevalent and destructive viral pathogen in soybean-planting regions of China. Moreover, other potyviruses, including bean common mosaic virus (BCMV) and watermelon mosaic virus (WMV), also threaten soybean farming. The eukaryotic translation initiation factor 4E (eIF4E) plays a critical role in controlling resistance/susceptibility to potyviruses in plants. In the present study, much higher SMV-induced eIF4E1 expression levels were detected in a susceptible soybean cultivar when compared with a resistant cultivar, suggesting the involvement of eIF4E1 in the response to SMV by the susceptible cultivar. Yeast two-hybrid and bimolecular fluorescence complementation assays showed that soybean eIF4E1 interacted with SMV VPg in the nucleus and with SMV NIa-Pro/NIb in the cytoplasm, revealing the involvement of VPg, NIa-Pro, and NIb in SMV infection and multiplication. Furthermore, transgenic soybeans silenced for eIF4E were produced using an RNA interference approach. Through monitoring for viral symptoms and viral titers, robust and broad-spectrum resistance was confirmed against five SMV strains (SC3/7/15/18 and SMV-R), BCMV, and WMV in the transgenic plants. Our findings represent fresh insights for investigating the mechanism underlying eIF4E-mediated resistance in soybean and also suggest an effective alternative for breeding soybean with broad-spectrum viral resistance.     

Amino acid substitution in P3 of Soybean mosaic virus to convert avirulence to virulence on Rsv4‐genotype soybean is influenced by the genetic composition of P3

The modification of avirulence factors of plant viruses by one or more amino acid substitutions converts avirulence to virulence on hosts containing resistance genes. Limited experimental studies have been conducted on avirulence/virulence factors of plant viruses, in particular those of potyviruses, to determine whether avirulence/virulence sites are conserved among strains. In this study, the Soybean mosaic virus (SMV)–Rsv4 pathosystem was exploited to determine whether: (i) avirulence/virulence determinants of SMV reside exclusively on P3 regardless of virus strain; and (ii) the sites residing on P3 and crucial for avirulence/virulence of isolates belonging to strain G2 are also involved in virulence of avirulent isolates belonging to strain G7. The results confirm that avirulence/virulence determinants of SMV on Rsv4‐genotype soybean reside exclusively on P3. Furthermore, the data show that sites involved in the virulence of SMV on Rsv4‐genotype soybean vary among strains, with the genetic composition of P3 playing a crucial role.     

Genetics of resistance to two strains of soybean mosaic virus in differential soybean genotypes

There are seven pathotypes of soybean mosaic virus (SMV) representing seven strain groups (G1-G7) in the United States. Soybean genotypes [Glycine max (L.) Merr.] may exhibit resistant (R), susceptible (S), or necrotic (N) reactions upon interacting with different SMV strains. This research was conducted to investigate whether reactions to two SMV strains are controlled by the same gene or by separate genes. Two SMV-resistant soybean lines, LR1 and LR2, were crossed with the susceptible cultivar Lee 68. LR1 contains a resistance gene Rsv1-s and is resistant to strains G1-G4 and G7. LR2 contains the Rsv4 gene and is resistant to strains G1-G7. Two hundred F(2:3) lines from LR1 x Lee 68 and 262 F(2:3) lines from LR2 x Lee 68 were screened for SMV reaction. Seeds from each F2 plant were randomly divided into two subsamples. A minimum of 20 seeds from each subsample were planted in the greenhouse and plants were inoculated with either G1 or G7. G1 is the least virulent, whereas G7 is the most virulent strain of SMV. The results showed that all the F(2:3) lines from both crosses exhibited the same reaction to G1 and G7. No recombinants were found in all the progenies for reactions to G1 and G7 in either cross. The results indicate that reactions to both G1 and G7 are controlled by either the same gene or very closely linked genes. This research finding is valuable for studying the resistance mechanism and interactions of soybean genotypes and SMV strains and for breeding SMV resistance to multiple strains.     

Genetic analysis and mapping of genes for resistance to multiple strains of Soybean mosaic virus in a single resistant soybean accession PI 96983

Soybean mosaic virus (SMV) is one of the most broadly distributed soybean (Glycine max (L.) Merr.) diseases and causes severe yield loss and seed quality deficiency. Multiple studies have proved that a single dominant gene can confer resistance to several SMV strains. Plant introduction (PI) 96983 has been reported to contain SMV resistance genes (e.g., Rsv1 and Rsc14) on chromosome 13. The objective of this study was to delineate the genetics of resistance to SMV in PI 96983 and determine whether one gene can control resistance to more than one Chinese SMV strain. In this study, PI 96983 was identified as resistant and Nannong 1138-2 was identified as susceptible to four SMV strains SC3, SC6, SC7, and SC17. Genetic maps based on 783 F₂ individuals from the cross of PI 96983 × Nannong 1138-2 showed that the gene(s) conferring resistance to SC3, SC6, and SC17 were between SSR markers BARCSOYSSR_13_1114 and BARCSOYSSR_13_1136, whereas SC7 was between markers BARCSOYSSR_13_1140 and BARCSOYSSR_13_1185. The physical map based on 58 recombinant lines confirmed these results. The resistance gene for SC7 was positioned between BARCSOYSSR_13_1140 and BARCSOYSSR_13_1155, while the resistance gene(s) for SC3, SC6, and SC17 were between BARCSOYSSR_13_1128 and BARCSOYSSR_13_1136. We concluded that, there were two dominant resistance genes flanking Rsv1 or one of them at the reported genomic location of Rsv1. One of them (designated as “Rsc-pm”) conditions resistance for SC3, SC6, and SC17 and another (designated as “Rsc-ps”) confers resistance for SC7. The two tightly linked genes identified in this study would be helpful to cloning of resistance genes and breeding of multiple resistances soybean cultivars to SMV through marker-assisted selection (MAS).