Major Soybean Maturity Gene Haplotypes Revealed by SNPViz Analysis of 72 Sequenced Soybean Genomes

In this Genomics Era, vast amounts of next-generation sequencing data have become publicly available for multiple genomes across hundreds of species. Analyses of these large-scale datasets can become cumbersome, especially when comparing nucleotide polymorphisms across many samples within a dataset and among different datasets or organisms. To facilitate the exploration of allelic variation and diversity, we have developed and deployed an in-house computer software to categorize and visualize these haplotypes. The SNPViz software enables users to analyze region-specific haplotypes from single nucleotide polymorphism (SNP) datasets for different sequenced genomes. The examination of allelic variation and diversity of important soybean [Glycine max (L.) Merr.] flowering time and maturity genes may provide additional insight into flowering time regulation and enhance researchers'' ability to target soybean breeding for particular environments. For this study, we utilized two available soybean genomic datasets for a total of 72 soybean genotypes encompassing cultivars, landraces, and the wild species Glycine soja. The major soybean maturity genes E1, E2, E3, and E4 along with the Dt1 gene for plant growth architecture were analyzed in an effort to determine the number of major haplotypes for each gene, to evaluate the consistency of the haplotypes with characterized variant alleles, and to identify evidence of artificial selection. The results indicated classification of a small number of predominant haplogroups for each gene and important insights into possible allelic diversity for each gene within the context of known causative mutations. The software has both a stand-alone and web-based version and can be used to analyze other genes, examine additional soybean datasets, and view similar genome sequence and SNP datasets from other species.     

Uncovering signatures of selection in the soybean genome using SSR diversity near QTLs of agronomic importance

The cultivated soybean [Glycine max (L.) Merr.] is widely considered to descend from the wild soybean (G. soja Sieb. & Zucc.). This study was designed to evaluate the genetic variability and differentiation between G. soja and G. max, and to detect signatures of the selection that may have occurred during the domestication process from G. soja to G. max. A total of 192 G. soja accessions and 104 G. max accessions were genotyped using eight selected simple sequence repeat (SSR) markers assigned to three SSR groups. Four SSRs in group A were not located near any known QTL. Three SSRs in group B were associated with seed protein content, and an SSR in group C was associated with resistance to Sclerotinia stem rot. The number of alleles per locus and the level of genetic variability in G. soja were higher than those in G. max. A total of 122 out of 125 alleles were present in G. soja, but only 59 alleles were detected in G. max. The average gene diversity was 0.74 in G. soja and 0.64 in G. max. Four SSRs near QTLs of agronomic importance showed strong genetic differentiation and shift change in high frequency alleles in groups B and C between G. soja and G. max, revealing selection signatures that may reflect the domestication events and recent selective breeding. With reduced diversity in G. max, some undomesticated genes from G. soja should be prime candidates for introgression to increase the pool of diversity in G. max.     

Single nucleotide mutation leading to an amino acid substitution in the variant <Emphasis Type="Italic">Tik</Emphasis> soybean Kunitz trypsin inhibitor (SKTI) identified in Chinese wild soybean (<Emphasis Type="Italic">Glycine soja</Emphasis> Sieb. &; Zucc.)

The wild soybean (Glycine soja), which is the progenitor of cultivated soybean (Glycine max), is expected to offer more information about genetic variability and more useful mutants for evolutionary research and breeding applications. Here, a total of 1,600 wild soybean samples from China were investigated for genetic variation with regard to the soybean Kunitz trypsin inhibitor (SKTI). A new mutant SKTI, Tik, was identified. It was found to be a Tia-derived codominant allele caused by a transversion point mutation from C to G at nucleotide +171, leading to an alteration of one codon (AAC → AAG) and a corresponding amino acid substitution (Asn → Lys) at the ninth residue. Upon examination of this variant and others previously found in wild soybeans, it became clear that SKTI has undergone high-level evolutionary differentiation. There were more abundant polymorphisms in the wild than in the cultivated soybean.     

QTL affecting fitness of hybrids between wild and cultivated soybeans in experimental fields

The objective of this study was to identify quantitative trait loci (QTL) affecting fitness of hybrids between wild soybean (Glycine soja) and cultivated soybean (Glycine max). Seed dormancy and seed number, both of which are important for fitness, were evaluated by testing artificial hybrids of G. soja × G. max in a multiple‐site field trial. Generally, the fitness of the F₁ hybrids and hybrid derivatives from self‐pollination was lower than that of G. soja due to loss of seed dormancy, whereas the fitness of hybrid derivatives with higher proportions of G. soja genetic background was comparable with that of G. soja. These differences were genetically dissected into QTL for each population. Three QTLs for seed dormancy and one QTL for total seed number were detected in the F₂ progenies of two diverse cross combinations. At those four QTLs, the G. max alleles reduced seed number and severely reduced seed survival during the winter, suggesting that major genes acquired during soybean adaptation to cultivation have a selective disadvantage in natural habitats. In progenies with a higher proportion of G. soja genetic background, the genetic effects of the G. max alleles were not expressed as phenotypes because the G. soja alleles were dominant over the G. max alleles. Considering the highly inbreeding nature of these species, most hybrid derivatives would disappear quickly in early self‐pollinating generations in natural habitats because of the low fitness of plants carrying G. max alleles.     

Evaluation of the geographical pattern of genetic diversity of Glycine soja and Glycine max based on four single copy nuclear gene loci: For conservation of soybean germplasm

Genetic diversity and its geographical patterns play a very important role in species conservation and exploitation. Here, nucleotide polymorphism patterns of four single copy nuclear gene loci in wild (Glycine soja) and cultivated soybean (Glycine max) populations from different geographical regions as well as their demographic history were analyzed. The results showed that: (1) Southern subpopulation has the highest, while central subpopulation revealed the lowest genetic diversity among three Chinese G. soja subpopulations. (2) Northern Chinese G. max subpopulation depicted higher genetic diversity than other two Chinese, Korean, Japanese and American G. max subpopulations. (3) Significant genetic differentiation (P < 0.001) was observed among Chinese G. soja subpopulations from three ecological zones. There was also a significant genetic differentiation（P < 0.01）between three Chinese and Japanese subpopulations of G. max. (4) The demographic dynamics revealed that effective population size of G. soja is expanding, while it was constant in G. max. G. soja is a useful germplasm resource to widen the genetic diversity of G. max. This study suggests that native populations of G. soja from different geo-ecological regions should be protected to conserve the genetic diversity.     

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.     

Divergence of flowering genes in soybean

Soybean genome sequences were blasted with Arabidopsis thaliana regulatory genes involved in photoperiod-dependent flowering. This approach enabled the identification of 118 genes involved in the flowering pathway. Two genome sequences of cultivated (Williams 82) and wild (IT182932) soybeans were employed to survey functional DNA variations in the flowering-related homologs. Forty genes exhibiting nonsynonymous substitutions between G. max and G. soja were catalogued. In addition, 22 genes were found to co-localize with QTLs for six traits including flowering time, first flower, pod maturity, beginning of pod, reproductive period, and seed filling period. Among the genes overlapping the QTL regions, two LHY/CCA1 genes, GI and SFR6 contained amino acid changes. The recently duplicated sequence regions of the soybean genome were used as additional criteria for the speculation of the putative function of the homologs. Two duplicated regions showed redundancy of both flowering-related genes and QTLs. ID 12398025, which contains the homeologous regions between chr 7 and chr 16, was redundant for the LHY/CCA1 and SPA1 homologs and the QTLs. Retaining of the CRY1 gene and the pod maturity QTLs were observed in the duplicated region of ID 23546507 on chr 4 and chr 6. Functional DNA variation of the LHY/CCA1 gene (Glyma07g05410) was present in a counterpart of the duplicated region on chr 7, while the gene (Glyma16g01980) present in the other portion of the duplicated region on chr 16 did not show a functional sequence change. The gene list catalogued in this study provides primary insight for understanding the regulation of flowering time and maturity in soybean.     

The salt resistance of wild soybean (Glycine soja Sieb. et Zucc. ZYD 03262) under NaCl stress is mainly determined by Na+ distribution in the plant

Understanding the salt resistance mechanism of wild soybean is important in improving salt tolerance of cultivated soybean. Therefore, we comparatively analyzed effects of NaCl on photosynthesis, antioxidant enzyme activity, and ion distribution in a cultivated (Glycine. max) and a wild (Glycine soja) soybean to study the salt resistance mechanism of the G. soja. The results showed that more Na⁺ was accumulated in the G. soja roots than in the G. max roots, but the Na⁺ in the G. soja leaves was much less than that observed in the G. max leaves. The Na⁺ concentrations in the G. soja leaves were not high enough to affect the photosynthetic apparatus, which was demonstrated by less inhibition of photosynthetic activity, stomatal conductance, carboxylation efficiency in the G. soja leaves than in the G. max leaves after treated with different concentrations of NaCl. Meanwhile, there were no significant changes in intercellular CO₂ concentration, maximum PSII quantum yield, and relative water content in the G. soja leaves after NaCl treatment, while they significantly decreased in the G. max leaves. The non-photochemical quenching and the activities of superoxide dismutase (EC 1.15.1.1) and ascorbate peroxidase (EC 1.11.1.11) in the G. soja leaves increased with the increasing of NaCl concentrations, whereas only the activity of superoxide dismutase increased in G. max leaves. Based on these results, we suggested that the G. soja is able to accumulate higher levels of Na⁺ in its roots, and prevent the transportation of Na⁺ to leaves to protect photosynthetic apparatus from salt damage.     

Identification of putative QTL that underlie yield in interspecific soybean backcross populations

Glycine soja, the wild progenitor of soybean, is a potential source of useful genetic variation in soybean improvement. The objective of our study was to map quantitative trait loci (QTL) from G. soja that could improve the crop. Five populations of BC₂F₄-derived lines were developed using the Glycine max cultivar IA2008 as a recurrent parent and the G. soja plant introduction (PI) 468916 as a donor parent. There were between 57 and 112 BC₂F₄-derived lines in each population and a total of 468 lines for the five populations. The lines were evaluated with simple sequence repeat markers and in field tests for yield, maturity, plant height, and lodging. The field testing was done over 2 years and at two locations each year. Marker data were analyzed for linkage and combined with field data to identify QTL. Using an experimentwise significance threshold of P=0.05, four yield QTL were identified across environments on linkage groups C2, E, K, and M. For these yield QTL, the IA2008 marker allele was associated with significantly greater yield than the marker allele from G. soja. In addition, one lodging QTL, four maturity QTL, and five QTL for plant height were identified across environments. Of the 14 QTL identified, eight mapped to regions where QTL with similar effects were previously mapped. Many regions carrying the yield QTL were also significant for other traits, such as plant height and lodging. When the significance threshold was reduced and the data were analyzed with simple linear regression, four QTL with a positive allele for yield from G. soja were mapped. One epistatic interaction between two genetic regions was identified for yield using an experimentwise significance threshold of P=0.05. Additional research is needed to establish whether multiple trait associations are the result of pleiotropy or genetic linkage and to retest QTL with a positive effect from G. soja.Communicated by H.C. Becker     

Study of the biosafety of genetically modified soybean in the center of its origin and diversity in the Far East of the Russian Federation

Wild soybean (Glycine soja Sieb. and Zucc.) is the nearest relative of cultivated soybean (G. max (L.) Merr.). Study of the population genetic structure of wild-growing relatives of genetically modified (GM) plants in the centers of their origin is one of the main procedures before introduction of GM crops in these areas. We studied the genetic variability of nine wild-growing soya populations of Primorskii krai using RAPD analysis. The level of genetic variability of G. soja was considerably higher than that of G. max. We analyzed phylogenetic relationships in the genus Glycine subgenus Soja using RAPD markers. Our data confirm the validity of allocation to G. gracilis of the rank of species.     

Differential responses of molecular mechanisms and physiochemical characters in wild and cultivated soybeans against invasion by the pathogenic Fusarium oxysporum Schltdl

Cultivated soybean (Glycine max) was derived from the wild soybean (Glycine soja), which has genetic resources that can be critically important for improving plant stress resistance. However, little information is available pertaining to the molecular and physiochemical comparison between the cultivated and wild soybeans in response to the pathogenic Fusarium oxysporum Schltdl. In this study, we first used comparative phenotypic and paraffin section analyses to indicate that wild soybean is indeed more resistant to F. oxysporum than cultivated soybean. Genome‐wide RNA‐sequencing approach was then used to elucidate the genetic mechanisms underlying the differential physiological and biochemical responses of the cultivated soybean, and its relative, to F. oxysporum. A greater number of genes related to cell wall synthesis and hormone metabolism were significantly altered in wild soybean than in cultivated soybean under F. oxysporum infection. Accordingly, a higher accumulation of lignins was observed in wild soybean than cultivated soybean under F. oxysporum infection. Collectively, these results indicated that secondary metabolites and plant hormones may play a vital role in differentiating the response between cultivated and wild soybeans against the pathogen. These important findings may provide future direction to breeding programs to improve resistance to F. oxysporum in the elite soybean cultivars by taking advantage of the genetic resources within wild soybean germplasm.     

Development,validation and genetic analysis of a large soybean SNP genotyping array

Cultivated soybean (Glycine max) suffers from a narrow germplasm relative to other crop species, probably because of under‐use of wild soybean (Glycine soja) as a breeding resource. Use of a single nucleotide polymorphism (SNP) genotyping array is a promising method for dissecting cultivated and wild germplasms to identify important adaptive genes through high‐density genetic mapping and genome‐wide association studies. Here we describe a large soybean SNP array for use in diversity analyses, linkage mapping and genome‐wide association analyses. More than four million high‐quality SNPs identified from high‐depth genome re‐sequencing of 16 soybean accessions and low‐depth genome re‐sequencing of 31 soybean accessions were used to select 180 961 SNPs for creation of the Axiom^® SoyaSNP array. Validation analysis for a set of 222 diverse soybean lines showed that 170 223 markers were of good quality for genotyping. Phylogenetic and allele frequency analyses of the validation set data indicated that accessions showing an intermediate morphology between cultivated and wild soybeans collected in Korea were natural hybrids. More than 90 unanchored scaffolds in the current soybean reference sequence were assigned to chromosomes using this array. Finally, dense average spacing and preferential distribution of the SNPs in gene‐rich chromosomal regions suggest that this array may be suitable for genome‐wide association studies of soybean germplasm. Taken together, these results suggest that use of this array may be a powerful method for soybean genetic analyses relating to many aspects of soybean breeding.     

Environmental versus geographical effects on genomic variation in wild soybean (Glycine soja) across its native range in northeast Asia

A fundamental goal in evolutionary biology is to understand how various evolutionary factors interact to affect the population structure of diverse species, especially those of ecological and/or agricultural importance such as wild soybean (Glycine soja). G. soja, from which domesticated soybeans (Glycine max) were derived, is widely distributed throughout diverse habitats in East Asia (Russia, Japan, Korea, and China). Here, we utilize over 39,000 single nucleotide polymorphisms genotyped in 99 ecotypes of wild soybean sampled across their native geographic range in northeast Asia, to understand population structure and the relative contribution of environment versus geography to population differentiation in this species. A STRUCTURE analysis identified four genetic groups that largely corresponded to the geographic regions of central China, northern China, Korea, and Japan, with high levels of admixture between genetic groups. A canonical correlation and redundancy analysis showed that environmental factors contributed 23.6% to population differentiation, much more than that for geographic factors (6.6%). Precipitation variables largely explained divergence of the groups along longitudinal axes, whereas temperature variables contributed more to latitudinal divergence. This study provides a foundation for further understanding of the genetic basis of climatic adaptation in this ecologically and agriculturally important species.     

Novel major quantitative trait loci regulating the content of isoflavone in soybean seeds

Despite their medicinal, pharmaceutical, and nutritional importance of isoflavones, the genetic basis controlling the amounts of isoflavones in soybean seeds is still not well understood. The main obstacle is the great variability in the content of isoflavone in seeds harvested from different environments. In this study, quantitative trait loci (QTL) for the content of different isoflavones including daidzein, genistein, and glycitein were investigated in a population of recombinant inbred lines derived from the cross of “Hwangkeum” (Glycine max) by “IT182932” (Glycine soja). Seeds analyzed were harvested in three different experimental environments. QTL analyses for isoflavone content were conducted by composite interval mapping across a genomewide genetic map. Two major QTL were mapped to soybean chromosomes 5 and 8, which were designated QDZGT1 and QDZGT2, respectively. Both loci have not been previously reported in other isoflavone sources. The results from this study will be useful in cloning genes that can control the contents of isoflavones in soybean and for the development of soybean lines containing a high or low isoflavone content.     

The genetic diversity and population structure of wild soybean evaluated by chloroplast and nuclear gene sequences

Glycine soja, also called wild soybean, is the wild ancestor of domesticated soybean (Glycine max), and one of the world's major cultivated crops. Wild soybean is a valuable resource for the breeding of cultivated soybean and harbors useful genes or agronomic traits. To use and conserve this valuable resource, we conducted a study to evaluate the genetic diversity and population structure of wild soybean using the sequencing data of two nuclear loci (AF105221 and PhyB) and one chloroplast locus (trnQ-rps16) of more than 600 individuals representing 53 populations throughout the natural distribution range. The results showed that most of the variation was found within the populations and groups, but significant genetic differentiation was also detected among different eco-geographical groups. Correlations between genetic and geographical distance at all the loci were consistent with the isolation by distance gene flow model. G. soja exhibited the highest genetic diversity in middle and downstream of Yangzi River (MDYR) region, followed by North East China (NEC), and was the lowest in North West China (NWC). We concluded that both in situ and ex situ conservation strategies required for wild soybean populations, especially which are native to MDYR and NEC regions.     

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.

     

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.     

Amplified fragment length polymorphism (AFLP) in soybean: species diversity, inheritance, and near-isogenic line analysis

Amplified fragment length polymorphism (AFLP) analysis is a PCR-based technique capable of detecting more than 50 independent loci in a single PCR reaction. The objectives of the present study were to: (1) assess the extent of AFLP variation in cultivated (Gycine max L. Merr.) and wild soybean (G. soja Siebold & Zucc.), (2) determine genetic relationships among soybean accessions using AFLP data, and (3) evaluate the usefulness of AFLPs as genetic markers. Fifteen AFLP primer pairs detected a total of 759 AFLP fragments in a sample of 23 accessions of wild and cultivated soybean, with an average of 51 fragments produced per primer pair per accession. Two-hundred and seventy four fragments (36% of the total observed) were polymorphic, among which 127 (17%) were polymorphic in G. max and 237 (31%) were polymorphic in G. soja. F₂ segregation analysis of six AFLP fragments indicated that they segregate as stable Mendelian loci. The number of polymorphic loci detected per AFLP primer pair in a sample of 23 accessions ranged from 9 to 27. The AFLP phenotypic diversity values were greater in wild than in cultivated soybean. Cluster and principal component analyses using AFLP data clearly separated G. max and G. soja accessions. Within the G. max group, adapted soybean cultivars were tightly clustered, illustrating the relatively low genetic diversity present in cultivated soybean. AFLP analysis of four soybean near-isogenic lines (NILs) identified three AFLP markers putatively linked to a virus resistance gene from two sources. The capacity of AFLP analysis to detect thousands of independent genetic loci with minimal cost and time requirements makes them an ideal marker for a wide array of genetic investigations.