Genome-wide signatures of the geographic expansion and breeding of soybean.

Soybean is a leguminous crop that provides oil and protein. Exploring the genomic signatures of soybean evolution is crucial for breeding varieties with improved adaptability to environmental extremes. We analyzed the genome sequences of 2,214 soybeans and proposed a soybean evolutionary route, i.e., the expansion of annual wild soybean (Glycine soja Sieb. & Zucc.) from southern China and its domestication in central China, followed by the expansion and local breeding selection of its landraces (G. max (L.) Merr.). We observed that the genetic introgression in soybean landraces was mostly derived from sympatric rather than allopatric wild populations during the geographic expansion. Soybean expansion and breeding were accompanied by the positive selection of flowering time genes, including GmSPA3c. Our study sheds light on the evolutionary history of soybean and provides valuable genetic resources for its future breeding.

Salt Tolerance in Soybeans: Focus on Screening Methods and Genetics.

Salinity greatly affects the production of soybeans in arid and semi-arid lands around the world. The responses of soybeans to salt stress at germination, emergence, and other seedling stages have been evaluated in multitudes of studies over the past decades. Considerable salt-tolerant accessions have been identified. The association between salt tolerance responses during early and later growth stages may not be as significant as expected. Genetic analysis has confirmed that salt tolerance is distinctly tied to specific soybean developmental stages. Our understanding of salt tolerance mechanisms in soybeans is increasing due to the identification of key salt tolerance genes. In this review, we focus on the methods of soybean salt tolerance screening, progress in forward genetics, potential mechanisms involved in salt tolerance, and the importance of translating laboratory findings into field experiments via marker-assisted pyramiding or genetic engineering approaches, and ultimately developing salt-tolerant soybean varieties that produce high and stable yields. Progress has been made in the past decades, and new technologies will help mine novel salt tolerance genes and translate the mechanism of salt tolerance into new varieties via effective routes.

Identification of Genomic Regions Associated with Vine Growth and Plant Height of Soybean.

Vining growth (VG) and high plant height (PH) are the physiological traits of wild soybean that preclude their utilization for domesticated soybean breeding and improvement. To identify VG- and PH-related quantitative trait loci (QTLs) in different genetic resources, two populations of recombinant inbred lines (RILs) were developed by crossing a cultivated soybean, Zhonghuang39 (ZH39), with two wild soybean accessions, NY27-38 and NY36-87. Each line from the two crosses was evaluated for VG and PH. Three QTLs for VG and three for PH, detected in the ZH39 × NY27-38 population of the RILs, co-located on chromosomes 2, 17 and 19. The VG- and PH-related QTL in the ZH39 × NY36-87 population co-located on chromosome 19. A common QTL shared by the two populations was located on chromosome 19, suggesting that this major QTL was consistently selected for in different genetic backgrounds. The results suggest that different loci are involved in the domestication or adaptations of soybean of various genetic backgrounds. The molecular markers presented here would benefit the fine mapping and cloning of candidate genes underlying the VG and PH co-localized regions and thus facilitate the utilization of wild resources in breeding by avoiding undesirable traits.

Identification of a Novel Salt Tolerance-Related Locus in Wild Soybean (Glycine soja Sieb. & Zucc.).

Salinity is an important abiotic stress factor that affects growth and yield of soybean. NY36-87 is a wild soybean germplasm with high salt tolerance. In this study, two F2:3 mapping populations derived from NY36-87 and two salt-sensitive soybean cultivars, Zhonghuang39 and Peking, were used to map salt tolerance-related genes. The two populations segregated as 1 (tolerant):2 (heterozygous):1 (sensitive), indicating a Mendelian segregation model. Using simple sequence repeat (SSR) markers together with the bulked segregant analysis (BSA) mapping strategy, we mapped a salt tolerance locus on chromosome 03 in F2:3 population Zhonghuang39×NY36-87 to a 98-kb interval, in which the known gene GmSALT3 co-segregated with the salt tolerance locus. In the F2:3 population of Peking×NY36-87, the dominant salt tolerance-associated gene was detected and mapped on chromosome 18. We named this gene GmSALT18 and fine mapped it to a 241-kb region. Time course analysis and a grafting experiment confirmed that Peking accumulated more Na+ in the shoot via a root-based mechanism. These findings reveal that the tolerant wild soybean line NY36-87 contains salt tolerance-related genes GmSALT3 and GmSALT18, providing genetic material and a novel locus for breeding salt-tolerant soybean.

Selection of the Salt Tolerance Gene GmSALT3 During Six Decades of Soybean Breeding in China.

Salt tolerance is an important trait that affects the growth and yield of plants growing in saline environments. The salt tolerance gene GmSALT3 was cloned from the Chinese soybean cultivar Tiefeng 8, and its variation evaluated in Chinese wild soybeans and landraces. However, the potential role of GmSALT3 in cultivation, and its genetic variation throughout the history of Chinese soybean breeding, remains unknown. Here we identified five haplotypes of GmSALT3 in 279 Chinese soybean landraces using a whole genome resequencing dataset. Additionally, we developed five PCR-based functional markers: three indels and two cleaved amplified polymorphic sequences (CAPS) markers. A total of 706 Chinese soybean cultivars (released 1956-2012), and 536 modern Chinese breeding lines, were genotyped with these markers. The Chinese landraces exhibited relatively high frequencies of the haplotypes H1, H4, and H5. H1 was the predominant haplotype in both the northern region (NR) and Huanghuai region (HHR), and H5 and H4 were the major haplotypes present within the southern region (SR). In the 706 cultivars, H1, H2, and H5 were the common haplotypes, while H3 and H4 were poorly represented. Historically, H1 gradually decreased in frequency in the NR but increased in the HHR; while the salt-sensitive haplotype, H2, increased in frequency in the NR during six decades of soybean breeding. In the 536 modern breeding lines, H2 has become the most common haplotype in the NR, while H1 has remained the highest frequency haplotype in the HHR, and H5 and H1 were highest in the SR. Frequency changes resulting in geographically favored haplotypes indicates that strong selection has occurred over six decades of soybean breeding. Our molecular markers could precisely identify salt tolerant (98.9%) and sensitive (100%) accessions and could accurately trace the salt tolerance gene in soybean pedigrees. Our study, therefore, not only identified effective molecular markers for use in soybean, but also demonstrated how these markers can distinguish GmSALT3 alleles in targeted breeding strategies for specific ecoregions.

Identification of loci controlling adaptation in Chinese soya bean landraces via a combination of conventional and bioclimatic GWAS.

Landraces often contain genetic diversity that has been lost in modern cultivars, including alleles that confer enhanced local adaptation. To comprehensively identify loci associated with adaptive traits in soya bean landraces, for example flowering time, a population of 1938 diverse landraces and 97 accessions of the wild progenitor of cultivated soya bean, Glycine soja was genotyped using tGBS® . Based on 99 085 high-quality SNPs, landraces were classified into three sub-populations which exhibit geographical genetic differentiation. Clustering was inferred from STRUCTURE, principal component analyses and neighbour-joining tree analyses. Using phenotypic data collected at two locations separated by 10 degrees of latitude, 17 trait-associated SNPs (TASs) for flowering time were identified, including a stable locus Chr12:5914898 and previously undetected candidate QTL/genes for flowering time in the vicinity of the previously cloned flowering genes, E1 and E2. Using passport data associated with the collection sites of the landraces, 27 SNPs associated with adaptation to three bioclimatic variables (temperature, daylength, and precipitation) were identified. A series of candidate flowering genes were detected within linkage disequilibrium (LD) blocks surrounding 12 bioclimatic TASs. Nine of these TASs exhibit significant differences in flowering time between alleles within one or more of the three individual sub-populations. Signals of selection during domestication and/or subsequent landrace diversification and adaptation were detected at 38 of the 44 flowering and bioclimatic TASs. Hence, this study lays the groundwork to begin breeding for novel environments predicted to arise following global climate change.

Phenotypic characterization and genetic dissection of nine agronomic traits in Tokachi nagaha and its derived cultivars in soybean (Glycine max (L.) Merr.).

By using the soybean founder parent Tokachi nagaha and its 137 derived cultivars as materials, a genome-wide association analysis was performed to identify the single nucleotide polymorphisms (SNPs) underlying soybean yield and quality related traits at two planting densities. Results of ANOVA showed that genotype, environment, and genotype by environment interaction effects were all significant for each trait. The Tokachi nagaha-derived soybean population could be divided into two subpopulations based on molecular data, and accessions in each subpopulation were almost all from the same Chinese province. Relatedness was detected between pair-wise accessions within the population. Linkage disequilibrium was obvious and the level of intra-chromosome linkage disequilibrium was about 8370kb. A total of 40 SNPs with significant signal were detected and distributed across 18 chromosomes. Some SNP markers were located in or near regions where QTLs have been previously mapped by linkage analysis. Nineteen SNPs were identified both in low- and high- density planting treatments, indicating those loci were common and sTable Sixteen SNPs were co-associated with two or more different traits, suggesting that some of the QTLs/genes underlying those identified SNPs were likely to be pleiotropic.

Development and utilization of a new chemically-induced soybean library with a high mutation density .

Mutagenized populations have provided important materials for introducing variation and identifying gene function in plants. In this study, an ethyl methanesulfonate (EMS)-induced soybean (Glycine max) population, consisting of 21,600 independent M2 lines, was developed. Over 1,000 M4 (5) families, with diverse abnormal phenotypes for seed composition, seed shape, plant morphology and maturity that are stably expressed across different environments and generations were identified. Phenotypic analysis of the population led to the identification of a yellow pigmentation mutant, gyl, that displayed significantly decreased chlorophyll (Chl) content and abnormal chloroplast development. Sequence analysis showed that gyl is allelic to MinnGold, where a different single nucleotide polymorphism variation in the Mg-chelatase subunit gene (ChlI1a) results in golden yellow leaves. A cleaved amplified polymorphic sequence marker was developed and may be applied to marker-assisted selection for the golden yellow phenotype in soybean breeding. We show that the newly developed soybean EMS mutant population has potential for functional genomics research and genetic improvement in soybean.

GmSALT3, Which Confers Improved Soybean Salt Tolerance in the Field, Increases Leaf Cl- Exclusion Prior to Na+ Exclusion But Does Not Improve Early Vigor under Salinity.

Soil salinity reduces soybean growth and yield. The recently identified GmSALT3 (Glycine max salt Tolerance-associated gene on chromosome 3) has the potential to improve soybean yields in salinized conditions. Here we evaluate the impact of GmSALT3 on soybean performance under saline or non-saline conditions. Three sets of near isogenic lines (NILs), with genetic similarity of 95.6-99.3% between each pair of NIL-T and NIL-S, were generated from a cross between two varieties 85-140 (salt-sensitive, S) and Tiefeng 8 (salt-tolerant, T) by using marker-assisted selection. Each NIL-T; 782-T, 820-T and 860-T, contained a common ~1000 kb fragment on chromosome 3 where GmSALT3 was located. We show that GmSALT3 does not contribute to an improvement in seedling emergence rate or early vigor under salt stress. However, when 12-day-old seedlings were exposed to NaCl stress, the NIL-T lines accumulated significantly less leaf Na+ compared with their corresponding NIL-S, while no significant difference of K+ concentration was observed between NIL-T and NIL-S; the magnitude of Na+ accumulation within each NIL-T set was influenced by the different genetic backgrounds. In addition, NIL-T lines accumulated less Cl- in the leaf and more in the root prior to any difference in Na+; in the field they accumulated less pod wall Cl- than the corresponding NIL-S lines. Under non-saline field conditions, no significant differences were observed for yield related traits within each pair of NIL-T and NIL-S lines, indicating there was no yield penalty for having the GmSALT3 gene. In contrast, under saline field conditions the NIL-T lines had significantly greater plant seed weight and 100-seed weight than the corresponding NIL-S lines, meaning GmSALT3 conferred a yield advantage to soybean plants in salinized fields. Our results indicated that GmSALT3 mediated regulation of both Na+ and Cl- accumulation in soybean, and contributes to improved soybean yield through maintaining a higher seed weight under saline stress.

Phenotypic Characterization and Genetic Dissection of Growth Period Traits in Soybean (Glycine max) Using Association Mapping.

The growth period traits are important traits that affect soybean yield. The insights into the genetic basis of growth period traits can provide theoretical basis for cultivated area division, rational distribution, and molecular breeding for soybean varieties. In this study, genome-wide association analysis (GWAS) was exploited to detect the quantitative trait loci (QTL) for number of days to flowering (ETF), number of days from flowering to maturity (FTM), and number of days to maturity (ETM) using 4032 single nucleotide polymorphism (SNP) markers with 146 cultivars mainly from Northeast China. Results showed that abundant phenotypic variation was presented in the population, and variation explained by genotype, environment, and genotype by environment interaction were all significant for each trait. The whole accessions could be clearly clustered into two subpopulations based on their genetic relatedness, and accessions in the same group were almost from the same province. GWAS based on the unified mixed model identified 19 significant SNPs distributed on 11 soybean chromosomes, 12 of which can be consistently detected in both planting densities, and 5 of which were pleotropic QTL. Of 19 SNPs, 7 SNPs located in or close to the previously reported QTL or genes controlling growth period traits. The QTL identified with high resolution in this study will enrich our genomic understanding of growth period traits and could then be explored as genetic markers to be used in genomic applications in soybean breeding.

Molecular and geographic evolutionary support for the essential role of GIGANTEAa in soybean domestication of flowering time.

BACKGROUND: Flowering time is a domestication trait of Glycine max and varies in soybeans, yet, a gene for flowering time variation has not been associated with soybean domestication. GIGANTEA (GI) is a major gene involved in the control of flowering time in Arabidopsis, although three GI homologs complicate this model in the soybean genome. RESULTS: In the present work, we revealed that the geographic evolution of the GIGANTEAa (GIa) haplotypes in G. max (GmGIa) and Glycine soja (GsGIa). Three GIa haplotypes (H1, H2, and H3) were found among cultivated soybeans and their wild relatives, yet an additional 44 diverse haplotypes were observed in wild soybeans. H1 had a premature stop codon in the 10(th) exon, whereas the other haplotypes encoded full-length GIa protein isoforms. In both wild-type and cultivated soybeans, H2 was present in the Southern region of China, and H3 was restricted to areas near the Northeast region of China. H1 was genetically derived from H2, and it was dominant and widely distributed among cultivated soybeans, whereas in wild populations, the ortholog of this domesticated haplotype H1 was only found in Yellow River basin with a low frequency. Moreover, this mutated GIa haplotype significantly correlated with early flowering. We further determined that the differences in gene expression of the three GmGIa haplotypes were not correlated to flowering time variations in cultivated soybeans. However, only the truncated GmGIa H1 could partially rescue gi-2 Arabidopsis from delayed flowering in transgenic plants, whereas both GmGIa H2 and H3 haplotypes could significantly repress flowering in transgenic Arabidopsis with a wild-type background. CONCLUSIONS: Thus, GmGIa haplotype diversification may have contributed to flowering time adaptation that facilitated the radiation of domesticated soybeans. In light of the evolution of the GIa gene, soybean domestication history for an early flowering phenotype is discussed.

Co-expression of G2-EPSPS and glyphosate acetyltransferase GAT genes conferring high tolerance to glyphosate in soybean.

Glyphosate is a widely used non-selective herbicide with broad spectrum of weed control around the world. At present, most of the commercial glyphosate tolerant soybeans utilize glyphosate tolerant gene CP4-EPSPS or glyphosate acetyltransferase gene GAT separately. In this study, both glyphosate tolerant gene G2-EPSPS and glyphosate degraded gene GAT were co-transferred into soybean and transgenic plants showed high tolerance to glyphosate. Molecular analysis including PCR, Sothern blot, qRT-PCR, and Western blot revealed that target genes have been integrated into genome and expressed effectively at both mRNA and protein levels. Furthermore, the glyphosate tolerance analysis showed that no typical symptom was observed when compared with a glyphosate tolerant line HJ06-698 derived from GR1 transgenic soybean even at fourfold labeled rate of Roundup. Chlorophyll and shikimic acid content analysis of transgenic plant also revealed that these two indexes were not significantly altered after glyphosate application. These results indicated that co-expression of G2-EPSPS and GAT conferred high tolerance to the herbicide glyphosate in soybean. Therefore, combination of tolerant and degraded genes provides a new strategy for developing glyphosate tolerant transgenic crops.

Targeted association mapping demonstrating the complex molecular genetics of fatty acid formation in soybean.

BACKGROUND: The relative abundance of five dominant fatty acids (FAs) (palmitic, stearic, oleic, linoleic and linolenic acids) is a major factor determining seed quality in soybean. METHODS: To clarify the currently poorly understood genetic architecture of FAs in soybean, targeted association analysis was conducted in 421 diverse accessions phenotyped in three environments and genotyped using 1536 pre-selected SNPs. RESULTS: The population of 421 soybean accessions displayed significant genetic variation for each FA. Analysis of the molecular data revealed three subpopulations, which reflected a trend depending on latitude of cultivation. A total of 37 significant (p < 0.01) associations with FAs were identified by association mapping analysis. These associations were represented by 33 SNPs (occurring in 32 annotated genes); another four SNPs had a significant association with two different FAs due to pleiotropic interactions. The most significant associations were cross-verified by known genes/QTL or consistency across cultivation year and subpopulations. CONCLUSION: The detected marker-trait associations represent a first important step towards the implementation of molecular-marker-based selection of FA composition with the potential to substantially improve the seed quality of soybean with benefits for human health and for food processing.

Salinity tolerance in soybean is modulated by natural variation in GmSALT3.

The identification of genes that improve the salt tolerance of crops is essential for the effective utilization of saline soils for agriculture. Here, we use fine mapping in a soybean (Glycine max (L.) Merr.) population derived from the commercial cultivars Tiefeng 8 and 85-140 to identify GmSALT3 (salt tolerance-associated gene on chromosome 3), a dominant gene associated with limiting the accumulation of sodium ions (Na+) in shoots and a substantial enhancement in salt tolerance in soybean. GmSALT3 encodes a protein from the cation/H+ exchanger family that we localized to the endoplasmic reticulum and which is preferentially expressed in the salt-tolerant parent Tiefeng 8 within root cells associated with phloem and xylem. We identified in the salt-sensitive parent, 85-140, a 3.78-kb copia retrotransposon insertion in exon 3 of Gmsalt3 that truncates the transcript. By sequencing 31 soybean landraces and 22 wild soybean (Glycine soja) a total of nine haplotypes including two salt-tolerant haplotypes and seven salt-sensitive haplotypes were identified. By analysing the distribution of haplotypes among 172 Chinese soybean landraces and 57 wild soybean we found that haplotype 1 (H1, found in Tiefeng 8) was strongly associated with salt tolerance and is likely to be the ancestral allele. Alleles H2-H6, H8 and H9, which do not confer salinity tolerance, were acquired more recently. H1, unlike other alleles, has a wide geographical range including saline areas, which indicates it is maintained when required but its potent stress tolerance can be lost during natural selection and domestication. GmSALT3 is a gene associated with salt tolerance with great potential for soybean improvement.

Detecting SNPs underlying domestication-related traits in soybean.

BACKGROUND: Cultivated soybean (Glycine max) experienced a severe genetic bottleneck during its domestication and a further loss in diversity during its subsequent selection. Here, a panel of 65 wild (G. soja) and 353 cultivated accessions was genotyped at 552 single-nucleotide polymorphism loci to search for signals of selection during and after domestication. RESULTS: The wild and cultivated populations were well differentiated from one another. Application of the Fst outlier test revealed 64 loci showing evidence for selection. Of these, 35 related to selection during domestication, while the other 29 likely gradually became monomorphic as a result of prolonged selection during post domestication. Two of the SNP locus outliers were associated with testa color. CONCLUSIONS: Identifying genes controlling domestication-related traits is important for maintaining the diversity of crops. SNP locus outliers detected by a combined forward genetics and population genetics approach can provide markers with utility for the conservation of wild accessions and for trait improvement in the cultivated genepool.

De novo assembly of soybean wild relatives for pan-genome analysis of diversity and agronomic traits.

Wild relatives of crops are an important source of genetic diversity for agriculture, but their gene repertoire remains largely unexplored. We report the establishment and analysis of a pan-genome of Glycine soja, the wild relative of cultivated soybean Glycine max, by sequencing and de novo assembly of seven phylogenetically and geographically representative accessions. Intergenomic comparisons identified lineage-specific genes and genes with copy number variation or large-effect mutations, some of which show evidence of positive selection and may contribute to variation of agronomic traits such as biotic resistance, seed composition, flowering and maturity time, organ size and final biomass. Approximately 80% of the pan-genome was present in all seven accessions (core), whereas the rest was dispensable and exhibited greater variation than the core genome, perhaps reflecting a role in adaptation to diverse environments. This work will facilitate the harnessing of untapped genetic diversity from wild soybean for enhancement of elite cultivars.

Biparental resequencing coupled with SNP genotyping of a segregating population offers insights into the landscape of recombination and fixed genomic regions in elite soybean.

Identification of genes underlying agronomic traits is dependent on the segregation of quantitative trait loci (QTL). A popular hypothesis is that elite lines are becoming increasingly similar to each other, resulting in large genomic regions with fixed genes. Here, we resequenced two parental modern elite soybean lines [ZhongHuang13 (ZH) and ZhongPin03-5373 (ZP)] and discovered 794,876 SNPs with reference to the published Williams82 genome. SNPs were distributed unevenly across the chromosomes, with 87.1% of SNPs clustering in 4.9% of the soybean reference genome. Most of the regions with a high density of SNP polymorphisms were located in the chromosome arms. Moreover, seven large regions that were highly similar between parental lines were identified. A GoldenGate SNP genotyping array was designed using 384 SNPs and the 254 recombinant inbred lines (F8) derived from the cross of ZP × ZH were genotyped. We constructed a genetic linkage map using a total of 485 molecular markers, including 313 SNPs from the array, 167 simple sequence repeats (SSRs), 4 expressed sequence tag-derived SSRs, and 1 insertion/deletion marker. The total length of the genetic map was 2594.34 cM, with an average marker spacing of 5.58 cM. Comparing physical and genetic distances, we found 20 hotspot and 14 coldspot regions of recombination. Our results suggest that the technology of resequencing of parental lines coupled with high-throughput SNP genotyping could efficiently bridge the genotyping gap and provide deep insights into the landscape of recombination and fixed genomic regions in biparental segregating populations of soybean with implications for fine mapping of QTL.

Molecular footprints of domestication and improvement in soybean revealed by whole genome re-sequencing.

BACKGROUND: Artificial selection played an important role in the origin of modern Glycine max cultivars from the wild soybean Glycine soja. To elucidate the consequences of artificial selection accompanying the domestication and modern improvement of soybean, 25 new and 30 published whole-genome re-sequencing accessions, which represent wild, domesticated landrace, and Chinese elite soybean populations were analyzed. RESULTS: A total of 5,102,244 single nucleotide polymorphisms (SNPs) and 707,969 insertion/deletions were identified. Among the SNPs detected, 25.5% were not described previously. We found that artificial selection during domestication led to more pronounced reduction in the genetic diversity of soybean than the switch from landraces to elite cultivars. Only a small proportion (2.99%) of the whole genomic regions appear to be affected by artificial selection for preferred agricultural traits. The selection regions were not distributed randomly or uniformly throughout the genome. Instead, clusters of selection hotspots in certain genomic regions were observed. Moreover, a set of candidate genes (4.38% of the total annotated genes) significantly affected by selection underlying soybean domestication and genetic improvement were identified. CONCLUSIONS: Given the uniqueness of the soybean germplasm sequenced, this study drew a clear picture of human-mediated evolution of the soybean genomes. The genomic resources and information provided by this study would also facilitate the discovery of genes/loci underlying agronomically important traits.

A platform for soybean molecular breeding: the utilization of core collections for food security.

Soybean is an important crop not only for human consumption but also for its addition of nitrogen to the soil during crop rotation. China has the largest collection of cultivated soybeans (Glycine max) and wild soybeans (Glycine soja) all over the world. The platform of soybean core, mini core and integrated applied core collections has been developed in the past decade based on systematic researches which included the sampling strategies, statistical methods, phenotypic data and SSR markers. Meanwhile, intergrated applied core collections including accessions with single or integrated favorite traits are being developed in order to meet the demand of soybean breeding. These kinds of core collections provide powerful materials for evaluation of germplasm, identification of trait-specific accessions, gene discovery, allele mining, genomic study, maker development, and molecular breeding. Some successful cases have proved the usefulness and efficiency of this platform. The platform is helpful for enhancing utilization of soybean genetic resources in sustainable crop improvement for food security. The efficient utilization of this platform in the future is relying on accurate phenotyping methods, abundant functional markers, high-throughput genotyping platforms, and effective breeding programs.

DNA sequence polymorphism of the Rhg4 candidate gene conferring resistance to soybean cyst nematode in Chinese domesticated and wild soybeans.

Rhg4 is one of the major resistant genes conferring resistance to soybean cyst nematode races 1, 3 and 4. In order to better understand its sequence diversity among different Chinese soybean populations and the impact of human activities on it, we designed 5 primer sets based on its sequence deposited in Genbank (Genbank accession No. AF506518) to obtain the Rhg4 sequence from 104 Chinese cultivated and wild soybean genotypes, and then analyzed the DNA sequence polymorphism in different Chinese soybean populations. The alignment of Rhg4 sequence included 5,216 nucleotide base pairs. A total of 67 single nucleotide polymorphisms (SNPs) including 59 single base changes and 8 DNA insertion-deletions (InDels) were identified with a SNP frequency of 1/78. Except for a 14-base InDel, there were 29 SNPs in coding regions, and among them, 13 were non-synonymous (9 in functional domains with 1 in a leucine-rich repeats region, 2 in a transmembrane region and 6 in a Ser/Thr kinase domain). The probability of substitution at each site was not the same, there were two hot spots, one was in the 5'-untranslated region between positions 124 and 804, and the other was in the region between positions 2520 and 3733. Sequence diversity analysis among 104 soybean genotypes showed π = 0.00102 and θ = 0.00218 for Rhg4. A domestication bottleneck was found because of lower sequence diversity and 58% unique SNPs loss in landraces compared with Glycine soja. Intensive selection increased the sequence diversity of cultivars, which had higher diversity and more unique SNPs than landraces. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s11032-012-9703-1) contains supplementary material, which is available to authorized users.