Soybean Mosaic Virus Resistance Genes Research Articles

Development of an NLR-ID Toolkit and Identification of Novel Disease-Resistance Genes in Soybean.

The recognition of pathogen effectors through the nucleotide-binding leucine-rich repeat receptor (NLR) family is an important component of plant immunity. In addition to typical domains such as TIR, CC, NBS, and LRR, NLR proteins also contain some atypical integrated domains (IDs), the roles of which are rarely investigated. Here, we carefully screened the soybean (Glycine max) genome and identified the IDs that appeared in the soybean TNL-like proteins. Our results show that multiple IDs (36) are widely present in soybean TNL-like proteins. A total of 27 Gm-TNL-ID genes (soybean TNL-like gene encoding ID) were cloned and their antiviral activity towards the soybean mosaic virus (SMV)/tobacco mosaic virus (TMV) was verified. Two resistance (R) genes, SRA2 (SMV resistance gene contains AAA_22 domain) and SRZ4 (SMV resistance gene contains zf-RVT domain), were identified to possess broad-spectrum resistance characteristics towards six viruses including SMV, TMV, plum pox virus (PPV), cabbage leaf curl virus (CaLCuV), barley stripe mosaic virus (BSMV), and tobacco rattle virus (TRV). The effects of Gm-TNL-IDX (the domain of the Gm-TNL-ID gene after the TN domain) on the antiviral activity of a R protein SRC7TN (we previously reported the TN domain of the soybean broad-spectrum resistance gene SRC7) were validated, and most of Gm-TNL-IDX inhibits antiviral activity mediated by SRC7TN, possibly through intramolecular interactions. Yeast-two-hybrid (Y2H) and bimolecular fluorescence complementation (BiFC) assays showed that seven Gm-TNL-IDX interacted with SMV-component proteins. Truncation analysis on a broad-spectrum antiviral protein SRZ4 indicated that SRZ4TIR is sufficient to mediate antiviral activity against SMV. Soybean cDNA library screening on SRZ4 identified 48 interacting proteins. In summary, our results indicate that the integration of IDs in soybean is widespread and frequent. The NLR-ID toolkit we provide is expected to be valuable for elucidating the functions of atypical NLR proteins in the plant immune system and lay the foundation for the development of engineering NLR for plant-disease control in the future.

Fine Mapping the Soybean Mosaic Virus Resistance Gene in Soybean Cultivar Heinong 84 and Development of CAPS Markers for Rapid Identification.

Heinong 84 is one of the major soybean varieties growing in Northeast China, and is resistant to the infection of all strains of soybean mosaic virus (SMV) in the region including the most prevalent strain, N3. However, the resistance gene(s) in Heinong 84 and the resistant mechanism are still elusive. In this study, genetic and next-generation sequencing (NGS)-based bulk segregation analysis (BSA) were performed to map the resistance gene using a segregation population from the cross of Heinong 84 and a susceptible cultivar to strain N3, Zhonghuang 13. Results show that the resistance of Heinong 84 is controlled by a dominant gene on chromosome 13. Further analyses suggest that the resistance gene in Heinong 84 is probably an allele of Rsv1. Finally, two pairs of single-nucleotide-polymorphism (SNP)-based primers that are tightly cosegregated with the resistance gene were designed for rapidly identifying resistant progenies in breeding via the cleaved amplified polymorphic sequence (CAPS) assay.

Identifying Quantitative Trait Loci and Candidate Genes Conferring Resistance to Soybean Mosaic Virus SC7 by Quantitative Trait Loci-Sequencing in Soybean.

Soybean mosaic virus (SMV) is detrimental to soybean (Glycine max) breeding, seed quality, and yield worldwide. Improving the basic resistance of host plants is the most effective and economical method to reduce damage from SMV. Therefore, it is necessary to identify and clone novel SMV resistance genes. Here, we report the characterization of two soybean cultivars, DN50 and XQD, with different levels of resistance to SMV. Compared with XQD, DN50 exhibits enhanced resistance to the SMV strain SC7. By combining bulked-segregant analysis (BSA)-seq and fine-mapping, we identified a novel resistance locus, RSMV-11, spanning an approximately 207-kb region on chromosome 11 and containing 25 annotated genes in the reference Williams 82 genome. Of these genes, we identified eleven with non-synonymous single-nucleotide polymorphisms (SNPs) or insertion-deletion mutations (InDels) in their coding regions between two parents. One gene, GmMATE68 (Glyma.11G028900), harbored a frameshift mutation. GmMATE68 encodes a multidrug and toxic compound extrusion (MATE) transporter that is expressed in all soybean tissues and is induced by SC7. Given that MATE transporter families have been reported to be linked with plant disease resistance, we suggest that GmMATE68 is responsible for SC7 resistance in DN50. Our results reveal a novel SMV-resistance locus, improving understanding of the genetics of soybean disease resistance and providing a potential new tool for marker-assisted selection breeding in soybean.

Soybean antiviral immunity conferred by dsRNase targets the viral replication complex

Eukaryotic positive-strand RNA viruses replicate their genomes in membranous compartments formed in a host cell, which sequesters the dsRNA replication intermediate from antiviral immune surveillance. Here, we find that soybean has developed a way to overcome this sequestration. We report the positional cloning of the broad-spectrum soybean mosaic virus resistance gene Rsv4, which encodes an RNase H family protein with dsRNA-degrading activity. An active-site mutant of Rsv4 is incapable of inhibiting virus multiplication and is associated with an active viral RNA polymerase complex in infected cells. These results suggest that Rsv4 enters the viral replication compartment and degrades viral dsRNA. Inspired by this model, we design three plant-gene-derived dsRNases that can inhibit the multiplication of the respective target viruses. These findings suggest a method for developing crops resistant to any target positive-strand RNA virus by fusion of endogenous host genes.

Marker-assisted pyramiding of soybean resistance genes RSC4, RSC8, and RSC14Q to soybean mosaic virus

Soybean mosaic virus (SMV) is one of the major viral pathogens affecting soybean crops worldwide. Three SMV resistance genes, RSC4, RSC8, and RSC14Q, have been identified and mapped on soybean chromosomes 14, 2, and 13 from Dabaima, Kefeng 1, and Qihuang 1 cultivars, respectively. Soybean cultivar Nannong 1138-2 is widely grown in the Yangtze River Valley of China. In this study, crosses were made between Qihuang 1×Kefeng 1 and Dabaima×Nannong 1138-2. Ten simple sequence repeat (SSR) markers linked to three resistance loci (RSC4, RSC8, and RSC14Q) were used to assist pyramided breeding. Pyramided families containing three resistance loci (RSC4, RSC8, and RSC14Q) were evaluated by inoculating them with 21 SMV strains from China. Results indicated that the 10 markers can be used effectively to assist the selection of resistant individuals containing RSC4, RSC8, and RSC14Q A total of 53 F6 plants were confirmed to contain three homozygous alleles conferring resistance to SMV. Five F7 homozygous pyramided families exhibited resistance to 21 strains of SMV and showed desirable agronomic traits using dual selection. The strategy of pyramiding resistance gene derived from different varieties has practical breeding value in providing broad-spectrum resistance against the existing strains of SMV in China.

Fine-mapping and identification of a novel locus Rsc15 underlying soybean resistance to Soybean mosaic virus.

Rsc15, a novel locus underlying soybean resistance to SMV, was fine mapped to a 95-kb region on chromosome 6. The Rsc15- mediated resistance is likely attributed to the gene GmPEX14 , the relative expression of which was highly correlated with the accumulation of H 2 O 2 along with the activities of POD and CAT during the early stages of SMV infection in RN-9. Soybean mosaic virus (SMV) causes severe yield losses and seed quality deterioration in soybean [Glycine max (L.) Merr.] worldwide. A series of single dominant SMV resistance genes have been identified on respective soybean chromosomes 2, 13 and 14, while one novel locus, Rsc15, underlying resistance to the virulent SMV strain SC15 from soybean cultivar RN-9 has been recently mapped to a 14.6-cM region on chromosome 6. However, candidate gene has not yet been identified within this region. In the present study, we aimed to fine map the Rsc15 region and identify candidate gene(s) for this invaluable locus. High-resolution fine-mapping revealed that the Rsc15 gene was located in a 95-kb genomic region which was flanked by the two simple sequence repeat (SSR) markers SSR_06_17 and BARCSOYSSR_06_0835. Allelic sequence comparison and expression profile analysis of candidate genes inferred that the gene Glyma.06g182600 (designated as GmPEX14) was the best candidate gene attributing for the resistance of Rsc15, and that genes encoding receptor-like kinase (RLK) (i.e., Glyma.06g175100 and Glyma.06g184400) and serine/threonine kinase (STK) (i.e., Glyma.06g182900 and Glyma.06g183500) were also potential candidates. High correlations were established between the relative expression level of GmPEX14 and the hydrogen peroxide (H2O2) concentration and activities of catalase (CAT) and peroxidase (POD) during the early stages of SMV-SC15 infection in RN-9. The results of the present study will be useful in marker-assisted breeding for SMV resistance and will lead to further understanding of the molecular mechanisms of host resistance against SMV.

Two Tightly Linked Genes for Soybean Mosaic Virus Resistance in Soybean

Soybean mosaic virus (SMV), a member of the Potyviridae family, is the most common virus negatively affecting yield and seed quality in soybean [Glycine max (L.) Merr.]. Seven SMV strains, G1 through G7, and three independent SMV resistance genes (R‐genes), Rsv1, Rsv3, and Rsv4, have been previously identified. The Rsv1 locus contains at least 10 alleles conferring differential plant reactions to SMV strains, and it was mapped at very complex resistance‐gene‐rich region. In this study, two alleles of the Rsv1 locus were analyzed by crossing PI 96983 (Rsv1) and cultivar York (Rsv1‐y) soybean to evaluate whether Rsv1 and Rsv1‐y belong to the same or different but closely linked loci. To break possible linkage, 3000 F2:3 lines were developed and investigated using infections of the SMV‐G1 strain in a greenhouse. The occurrence of segregating and susceptible lines indicated tight linkage between two genes. The recombination frequency (RF) was estimated using the maximum likelihood (ML) formula concluding that Rsv1 and Rsv1‐y are two tightly linked loci located apart with genetic distance of 2.2 cM on chromosome 13. A new symbol of Rsv5 has been assigned for the resistance gene in York to replace the original Rsv1‐y allele assignment. This research provided the first evidence for existence of two R‐genes on chromosome 13, conferring resistance to different SMV strains. Both loci, Rsv1 and Rsv5, can be easily transferred into susceptible cultivars in a breeding program to provide broad and durable protection against SMV strains.

Identification of candidate genes for resistance to Soybean mosaic virus strain SC3 by using fine mapping and transcriptome analyses

This study is a continuation of our earlier reports on the identification and mapping of the Soybean mosaic virus (SMV) strain SC3 resistance gene present on chromosome 13 (LG F) of soybean (Glycine max (L.) Merr.). We used a combination of fine mapping and transcriptome sequencing to discover the candidate genes for SMV resistance in Qihuang-1. To fine-map the resistance gene, near-isogenic lines (NILs) from a cross between Qihuang-1 (resistant) and Nannong 1138-2 (susceptible) were genotyped with simple sequence repeats (SSR) and insertion and deletion (indel) markers to identify recombination events. Analysis of plants carrying key recombination events placed the resistance gene to a 180-kb region of the ‘Williams 82’ genome sequence with 17 annotated genes. Transcriptome and quantitative real-time PCR (qRT-PCR) analyses revealed that SMV resistance in Qihuang-1 was probably attributable to the four candidate genes (Glyma13g25920, Glyma13g25950, Glyma13g25970 and Glyma13g26000). The four candidate genes identified in this study can be used in further studies to investigate the role of resistance genes in conferring SC3 resistance in Qihuang-1.

Assessment of genetic variation at soybean mosaic virus resistance loci in Indian Soybean (Glycine max L. Merill) genotypes using SSR markers

Soybean mosaic virus (SMV) disease is one of the major diseases of soybean globally that hampers soybean productivity. In India, SMV disease is gaining importance and there is a need to breed the soybean cultivars resistant to SMV. In India, soybean cultivars resistant to SMV have been identified but source of resistance has not been characterized. Therefore, the current study was carried out to study the genetic variation at three SMV resistance loci in a set of SMV resistant and susceptible Indian soybean genotypes using the mapped SSR markers. A total of 13 SSR markers reported to be linked to three SMV resistance loci were screened on 23 soybean genotypes showing differential response to SMV disease. A total of 54 alleles were generated at 12 polymorphic SSR loci with an average of 4.5 alleles/locus. The number of alleles ranged from 2 to 7 and gene diversity for the polymorphic SSR markers ranged from 0.38 to 0.77 with an average of 0.60, indicating the presence of high genetic variation in the Indian soybean genotypes at SMV resistance loci. Cluster analysis grouped the 23 soybean genotypes into three major clusters and results of the Principal Coordinate Analysis also congruent well with the cluster analysis. In the clustering, most of the genotypes were grouped as per their pedigree suggesting that they may be carrying the same SMV resistance gene. This study showed that Indian soybean genotypes have sufficient genetic variability at the three SMV resistance loci and it would be helpful in the selection of suitable parents in breeding for SMV resistance.

Identification of haplotypes at the Rsv4 genomic region in soybean associated with durable resistance to soybean mosaic virus.

Discovery of new germplasm sources and identification of haplotypes for the durable Soybean mosaic virus resistance gene, Rsv 4, provide novel resources for map-based cloning and genetic improvement efforts in soybean. The Soybean mosaic virus (SMV) resistance locus Rsv4 is of interest because it provides a durable type of resistance in soybean [Glycine max (L.) Merr.]. To better understand its molecular basis, we used a population of 309 BC3F2 individuals to fine-map Rsv4 to a ~120 kb interval and leveraged this genetic information in a second study to identify accessions 'Haman' and 'Ilpumgeomjeong' as new sources of Rsv4. These two accessions along with three other Rsv4 and 14 rsv4 accessions were used to examine the patterns of nucleotide diversity at the Rsv4 region based on high-depth resequencing data. Through a targeted association analysis of these 19 accessions within the ~120 kb interval, a cluster of four intergenic single-nucleotide polymorphisms (SNPs) was found to perfectly associate with SMV resistance. Interestingly, this ~120 kb interval did not contain any genes similar to previously characterized dominant disease resistance genes. Therefore, a haplotype analysis was used to further resolve the association signal to a ~94 kb region, which also resulted in the identification of at least two Rsv4 haplotypes. A haplotype phylogenetic analysis of this region suggests that the Rsv4 locus in G. max is recently introgressed from G. soja. This integrated study provides a strong foundation for efforts focused on the cloning of this durable virus resistance gene and marker-assisted selection of Rsv4-mediated SMV resistance in soybean breeding programs.

Genetic analysis and identification of two soybean mosaic virus resistance genes in soybean [Glycine max (L.) Merr]

AbstractSoybean mosaic virus (SMV) commonly affects soybean production worldwide, and the SC18 strain has been widespread in China. This study aimed to characterize and map the SC18 resistance genes present in soybean cultivars ‘Kefeng No. 1’ and ‘Qihuang 22’. Inheritance analysis revealed that two independent single dominant genes in Kefeng No. 1 and Qihuang 22 confer resistance to SC18. Using simple sequence repeat (SSR) markers and bulked segregant analysis, the Kefeng No. 1 and Qihuang 22 resistance genes were located on soybean chromosomes 2 and 13, respectively. We further screened two populations of recombinant inbred lines with 32 SSR markers in the target region, where the resistance gene in Kefeng No. 1 was fine mapped to an 80‐kb region containing six putative genes. Sequence and expression analyses of these genes revealed that SMV resistance in Kefeng No. 1 was probably attributable to three of the candidate genes (i.e. Glyma.02G127800, Glyma.02G128200 and Glyma.02G128300). Collectively, the results of this study will greatly facilitate the cloning of SC18 resistance genes and marker‐assisted breeding of SMV‐resistant soybean cultivars.

Multiple genes confer resistance to soybean mosaic virus in the soybean cultivar Hwangkeum

Recombinant inbred lines (RILs) generated from a cross between a cultivated species and its wild progenitor species serve as important germplasm for introgressing valuable genes from a wild species to a cultivated species. During this breeding process, it is equally important to prevent the loss of agronomically important genes in the cultivated species. In an effort to establish an efficient selection system for the single Rsv1 gene conferring durable resistance to soybean mosaic virus (SMV) in the soybean cultivar Hwangkeum (also known as Suweon 97), which is a parent of a RIL population from Hwangkeum (cultivated soybean) × IT182932 (wild soybean), in the present study, we unexpectedly identified an additional necrosis-conditioning gene unmasked by a recombination in the middle of the Rsv1-containing nucleotide-binding leucine-rich repeat gene cluster region and the Rsv3 gene in Hwangkeum. Thus, Hwangkeum contains at least three SMV resistance genes consisting of two tightly linked genes at the Rsv1 locus and the Rsv3 locus. The results of this study provide additional important information for use of the Hwangkeum genome in soybean breeding programmes.

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).

The Requirement of Multiple Defense Genes in SoybeanRsv1–Mediated Extreme Resistance toSoybean mosaic virus

Soybean mosaic virus (SMV) is a major viral pathogen of soybean. Among the three SMV resistance genes, Rsv1 mediates extreme resistance (ER) against most SMV strains, including the β-glucuronidase-tagged G2 isolate that was previously used in studies of Rsv1. Using virus-induced gene silencing (VIGS), we screened 82 VIGS constructs to identify genes that play a role in Rsv1-mediated ER to SMV infection. The target genes included putative Rsv1 candidate genes, soybean orthologs to known defense-signaling genes, and 62 WRKY transcription factors. We identified eight VIGS constructs that compromised Rsv1-mediated resistance when the target genes were silenced, including GmEDR1, GmEDS1, GmHSP90, GmJAR1, GmPAD4, and two WRKY transcription factors. Together, our results provide new insight into the soybean signaling network required for ER against SMV.

Fine mapping of the R SC14Q locus for resistance to soybean mosaic virus in soybean

Soybean mosaic virus (SMV) disease is one of the most destructive soybean (Glycine max (L.) Merr.) diseases worldwide, causing yield losses and significant deterioration in seed quality. ‘Qihuang No.1’ has a wide resistance spectrum and is extensively used in breeding programs in China. The dominant SMV resistance gene R SC14Q from the cultivar Qihuang No.1 was previously mapped to a 4.2 cM region on soybean chromosome 13 (formerly linkage group (LG) F). Fine mapping of R SC14Q is essential for high efficiency of marker-assisted selection (MAS) and its map-based cloning. Residual heterozygous lines (RHLs) derived from a soybean recombinant inbred line (RIL) were used to fine map R SC14Q . The RILs were obtained from a cross between ‘Qihuang No.1’ (resistant) and ‘Nannong1138-2’ (susceptible). The genomic sequence in an approximately 6 cM interval between the markers Satt334 and Sct_033 flanking R SC14Q was used to develop five simple sequence repeats and two InDel markers. Linkage analysis between inoculation phenotype and genetic markers localized R SC14Q to the interval between Satt334 and MY750, with genetic distances of 0.6 and 0.5 cM, respectively, corresponding to a physical distance on the ‘Williams 82’ draft assembly (Glyma1.13) of 1.18 Mb. Based on the genes sequences in the R SC14Q locus region with predicted functions for resistance, one SNP marker, MY525, was developed between Satt334 and MY750. Thus, the genomic region containing the R SC14Q was further narrowed to a 616 kb interval.

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.

Molecular Mapping and Marker Assisted Selection of Soybean Mosaic Virus Resistance Gene RSC12 in Soybean

The P 1 , P 2 , F 1 plants, F 2 population and F 2:3 lines from the cross of Qihuang22×Nannong1138-2 were inoculated with the soybean mosaic virus (SMV) strain SC12 for identification of their resistance in the greenhouse. Qihuang22 and F 1 individuals were resistant(R), and Nannong1138-2 were susceptible(S). The F 2 population segregated in a ratio of 3(R):1(S), and the F 2:3 lines exhibited a segregation pattern of 1(R):2(Segregating):1(S). These results indicated that a single dominant gene controlled the resistance to SC12. A F 2 population of Qihuang22×Nannong 1138-2 with 219 individuals was constructed for molecular mapping of resistance gene RSC12 to soybean mosaic virus in soybean. Linkage analysis with bulk segregant analysis (BSA) demonstrated that the resistance gene RSC12 was located on the linkage group F and linked with seven SSR markers. The order and genetic distance of markers linked with RSC12 were Sat_297 6.4 cM Sat_234 4.9 cM Sat_154 1.1 cM Satt114 0.7 cM SOYHSP176 1.6 cM Satt334 2.4 cM RSC12 6.3cM Sct_033. The marker-assisted selection (MAS) efficiency of SSR markers Satt334 and Sct_033 was evaluated in F 2 , F 3 and F 4 populations. The results showed that the MAS efficiency of Satt334 and Sct_033 was more than 85%, and that the MAS efficiency reached as high as 95% when these two markers were co-used. Therefore, the two SSR markers can be used effectively in selecting for resistance genes RSC12 instead of inoculation identification.

Strain-Specific Cylindrical Inclusion Protein of Soybean mosaic virus Elicits Extreme Resistance and a Lethal Systemic Hypersensitive Response in Two Resistant Soybean Cultivars

In the Soybean mosaic virus (SMV)-soybean pathosystem, three independent genes (Rsv1, Rsv3, and Rsv4) conferring resistance to SMV have been identified. Recently, we constructed infectious cDNA clones of SMV G7H and G5H strains and found that these two strains differ in their ability to infect soybean genotypes possessing different SMV resistance genes despite a difference of only 33 amino acids. In particular, pSMV-G7H induced mosaic symptoms systemically in L29 (Rsv3) and provoked a lethal systemic hypersensitive response (LSHR) in Jinpumkong-2, whereas pSMV-G5H could not infect these soybean genotypes. To identify the responsible pathogenic determinants of SMV, we exploited the differential responses of pSMV-G7H- and pSMV-G5H-derived chimeric viruses and amino acid substitution mutant viruses in several soybean genotypes and demonstrated that cylindrical inclusion (CI) protein is the elicitor of Rsv3-mediated extreme resistance and a pathogenic determinant provoking LSHR in Jinpumkong-2. A single amino acid substitution in CI was found to be responsible for gain or loss of elicitor function of CI. Our finding provides a role for CI as a pathogenic determinant in the SMV-soybean pathosystem, and increases the understanding of the basis of the different disease responses of SMV strains.

Molecular variability and genetic structure of the population of Soybean mosaic virus based on the analysis of complete genome sequences

The complete genomes of 30 Soybean mosaic virus (SMV) isolates and strains were sequenced in this study. Together with fourteen previously reported sequences, we analyzed the genetic structure of the SMV population. Analyses of genetic diversity showed that different genomic regions of SMV are under different evolutionary constraints and that there was no significant genetic differentiation between East Asian and North American populations of SMV. Phylogenetic analyses revealed a significant correlation between phylogeny of the cylindrical inclusion (CI) gene of SMV and SMV resistance gene 3 (Rsv3)-relating pathogenicity of SMV, suggesting CI might be a pathogenic determinant in Rsv3-mediated disease response. Interestingly, recombination analyses identified 19 'clear' recombination events in the SMV population. Furthermore, as several resistance-breaking strains were identified as recombinants, it appears that recombination might contribute to overcome host resistance in SMV-soybean pathosystem. Our finding suggests that recombination as well as mutation is an important evolutionary process in the genetic diversification of SMV population.

Marker-assisted identification of resistance genes to soybean mosaic virus in soybean lines

Soybean plants react differentially to soybean mosaic virus (SMV) strains because of interactions among different resistant genes in the soybean genome. Three independent genes resistant to SMV have been identified by inheritance studies and linkage analyses. To develop durable SMV-resistant soybean cultivars, it is necessary to determine which soybean SMV resistance genes can be readily transferred from resistant to susceptible cultivars in a breeding system. Here, we report the type and number of resistance gene(s) in four Korean elite soybean cultivars using a combination of disease reaction symptoms, inheritance studies, and molecular marker mappings. The disease reactions of Sowonkong and Keunolkong soybean varietals in response to infection with SMV strains suggested that both cultivars most likely harbor the Rsv1 gene similar to that in York. Subsequent inheritance studies confirmed that Sowonkong has the Rsv1 gene. The inheritance studies suggested that Sinpaldalkong harbored the Rsv1 gene, which was then confirmed by molecular marker mapping. The inheritance studies also suggested that Jinpumkong 2, which is the most resistant to SMV infection among the four cultivars, contained the Rsv1 and Rsv3 genes; this was confirmed by molecular marker mapping. Our approach, which combined inheritance studies and molecular linkage analyses, allowed the efficient identification of resistance gene(s) in four Korean soybean cultivars.