Developing Soybean Seeds Research Articles

Bioinformatics Identification and Expression Analysis of Acetyl-CoA Carboxylase Reveal Its Role in Isoflavone Accumulation during Soybean Seed Development.

Isoflavones belong to the class of flavonoid compounds, which are important secondary metabolites that play a crucial role in plant development and defense. Acetyl-CoA carboxylase (ACCase) is a biotin-dependent enzyme that catalyzes the conversion of Acetyl-CoA into Malonyl-CoA in plants. It is a key enzyme in fatty acid synthesis and also catalyzes the production of various secondary metabolites. However, information on the ACC gene family in the soybean (Glycine max L. Merr.) genome and the specific members involved in isoflavone biosynthesis is still lacking. In this study, we identified 20 ACC family genes (GmACCs) from the soybean genome and further characterized their evolutionary relationships and expression patterns. Phylogenetic analysis showed that the GmACCs could be divided into five groups, and the gene structures within the same groups were highly conserved, indicating that they had similar functions. The GmACCs were randomly distributed across 12 chromosomes, and collinearity analysis suggested that many GmACCs originated from tandem and segmental duplications, with these genes being under purifying selection. In addition, gene expression pattern analysis indicated that there was functional divergence among GmACCs in different tissues. The GmACCs reached their peak expression levels during the early or middle stages of seed development. Based on the transcriptome and isoflavone content data, a weighted gene co-expression network was constructed, and three candidate genes (Glyma.06G105900, Glyma.13G363500, and Glyma.13G057400) that may positively regulate isoflavone content were identified. These results provide valuable information for the further functional characterization and application of GmACCs in isoflavone biosynthesis in soybean.

Drought stress reduces the photosynthetic source of subtending leaves and the transit sink function of podshells, leading to reduced seed weight in soybean plants.

Drought stress is the key factor limiting soybean yield potential. Soybean seed formation involves a coordinated "subtending leaf-podshell-seed" process, but little is known about the assimilation and transport of photoassimilates in subtending leaves, podshells and seeds or their relationships with soybean seed formation under drought stress. To address these research gaps, two-year experiments with two soybean cultivars, Wandou 37 (drought tolerant) and Zhonghuang 13 (drought sensitive), were conducted under three soil water content (SWC) conditions in 2020 and 2021 based on the responses of their yield to drought. We analyzed the photosynthetic assimilation and translocation of photoassimilates in subtending leaves, podshells and seeds by stable isotope labeling. Compared with those under 75% SWC, 60% SWC and 45% SWC significantly decreased the Wandou 37 seed weight by 19.4% and 37.5%, respectively, and that of Zhonghuang 13 by 26.9% and 48.6%, respectively. Compared with those under 75% SWC, drought stress decreased the net photosynthetic rate and the activities of sucrose phosphate synthase (SPS) and sucrose synthase (SuSy), which in turn decreased the photosynthetic capacity of the subtending leaves. The podshells ensure the input of photoassimilates by increasing the SuSy activity, but the weakened source-sink relationship between podshells and seeds under drought stress leads to a decrease in the translocation of assimilates from podshells to seeds. The lack of assimilates under drought stress is an important factor restricting the development of soybean seeds. We conclude that the decrease in seed weight was caused by the decrease in the photosynthetic capacity of the subtending leaves and the decrease in the overall availability of photoassimilates; moreover, by a decrease in the translocation of assimilates from podshells to seeds.

Investigating Variability in Viral Presence and Abundance across Soybean Seed Development Stages Using Transcriptome Analysis.

Plant transcriptomes offer a valuable resource for studying viral communities (viromes). In this study, we explore how plant transcriptome data can be applied to virome research. We analyzed 40 soybean transcriptomes across different growth stages and identified six viruses: broad bean wilt virus 2 (BBWV2), brassica yellow virus (BrYV), beet western yellow virus (BWYV), cucumber mosaic virus (CMV), milk vetch dwarf virus (MDV), and soybean mosaic virus (SMV). SMV was the predominant virus in both Glycine max (GM) and Glycine soja (GS) cultivars. Our analysis confirmed its abundance in both, while BBWV2 and CMV were more prevalent in GS than GM. The viral proportions varied across developmental stages, peaking in open flowers. Comparing viral abundance measured by viral reads and fragments per kilobase of transcript per million (FPKM) values revealed insights. SMV showed similar FPKM values in GM and GS, but BBWV2 and CMV displayed higher FPKM proportions in GS. Notably, the differences in viral abundance between GM and GS were generally insignificant based on the FPKM values across developmental stages, except for the apical bud stage in four GM cultivars. We also detected MDV, a multi-segmented virus, in two GM samples, with variable proportions of its segments. In conclusion, our study demonstrates the potential of plant transcriptomes for virome research, highlighting their strengths and limitations.

Unfolding the Dynamics of Mineral Nutrients and Major Storage Protein Fractions during Soybean Seed Development

Unfolding the Dynamics of Mineral Nutrients and Major Storage Protein Fractions during Soybean Seed Development

Identification of Candidate Genes for Drought Resistance during Soybean Seed Development

Drought is the main abiotic stress limiting soybean production worldwide and seriously limits the reproductive growth of soybean. To elucidate the molecular mechanisms of drought stress response in soybean seed developmental stages, transcriptome analysis was performed on five seed developmental stages under drought stress and control conditions. Thousands of genes were found to change in expression level under drought stress, GO and KEGG enrichment analysis showed that differentially expressed genes (DEGs) were involved in drought resistance at every stage or tissue, but there were very few DEGs at the primary stage of seed development, which indicated that the seeds at the primary developmental stage were influenced little by drought stress. There was no shared DEG among all five stages or tissues, this indicated that they have different mechanisms to resist drought stress. Furthermore, we investigated DEGs associated with drought response at different stages to identify candidate regulators of drought stress response during soybean seed development. These genes are associated with metabolic and regulatory networks, among others. Taken together, the results will expand our understanding of the adaptation of soybean to drought stress and provide new insights into the molecular mechanisms of soybean drought tolerance development.

Novel Seed Size: A Novel Seed-Developing Gene in Glycine max.

Soybean-seed development is controlled in multiple ways, as in many known regulating genes. Here, we identify a novel gene, Novel Seed Size (NSS), involved in seed development, by analyzing a T-DNA mutant (S006). The S006 mutant is a random mutant of the GmFTL4pro:GUS transgenic line, with phenotypes with small and brown seed coats. An analysis of the metabolomics and transcriptome combined with RT-qPCR in the S006 seeds revealed that the brown coat may result from the increased expression of chalcone synthase 7/8 genes, while the down-regulated expression of NSS leads to small seed size. The seed phenotypes and a microscopic observation of the seed-coat integument cells in a CRISPR/Cas9-edited mutant nss1 confirmed that the NSS gene conferred small phenotypes of the S006 seeds. As mentioned in an annotation on the Phytozome website, NSS encodes a potential DNA helicase RuvA subunit, and no such genes were previously reported to be involved in seed development. Therefore, we identify a novel gene in a new pathway controlling seed development in soybeans.

Distribution analysis of jasmonic acid-related compounds in developing Glycine max L. (soybean) seeds using mass spectrometry imaging and liquid chromatography-mass spectrometry.

Jasmonic acid (JA) and its precursors are oxylipins derived from α-linolenic acid (αLA) and hexadecatrienoic acid, and regulate seed development. However, their spatial distribution in the developing Glycine max L. (soybean) seeds has not been elucidated. To investigate the distribution of JA-related compounds in the developing soybean seeds using desorption electrospray ionisation-mass spectrometry imaging (DESI-MSI) and liquid chromatography-electrospray ionisation-mass spectrometry (LC-ESI-MS) analyses. Cryosections of developing seeds were prepared using adhesive films, and subjected to DESI-MSI analysis. Verification of the DESI-MSI ion images were performed using DESI-tandem MSI (MS/MSI), LC-ESI-MS and tandem MS (MS/MS). In the DESI-MSI mass spectrum, peaks matching the chemical formulae of αLA, 12-oxo-phytodienoic acid (OPDA), and 3-oxo-2-(2-(Z)-pentenyl)-cyclopentane-1-octanoic acid (OPC-8:0) were detected. These compounds were mainly distributed in the seed coat, especially near the hilum. This was consistent with the quantitative results obtained by LC-ESI-MS. While, DESI-MS/MSI and LC-ESI-MS/MS suggested the presence of isomers for OPDA and OPC-8:0. The effect of isomers on the DESI-MSI ion images was small for OPDA, and considerable for OPC-8:0. These results demonstrated that free αLA, OPDA, and OPC-8:0 were the abundant JA-related compounds mainly distributed in the seed coat of the developing soybeans. OPDA and OPC-8:0 might exert a biological role in the seed coat. To the best of my knowledge, this is the first report on the accumulation of OPDA and OPC-8:0 in the seed coat. The combination of DESI-MSI and LC-ESI-MS is a useful tool for distribution analysis of JA-related compounds in the developing seeds.

Allelopathic Potential of Hesperozygis ringens Extracts on Seed Germination of Soybeans and Beggarticks

Hesperozygis ringens L., popularly known as espanta-pulga, is a native shrub found in rocky fields in Rio Grande do Sul. Its leaves produce a lot of essential oil, which contains pulegone as a main component that may be allelopathic. The objectives of this study were to analyze the allelopathic effects of H. ringens leaf extracts on the germination and initial development of soybean seeds and to evaluate the herbicidal potential of these extracts on beggarticks (Bidens pilosa L.). The seeds were placed on paper embedded in an aqueous extract of H. ringens leaves at concentrations of 0, 25, 50, 75 and 100% and maintained in a germination chamber at 25 °C and a photoperiod of 12 h. The tests included the following: germination, first germination count, germination speed index, and length and dry mass of seedlings. The experimental design was completely randomized. Based on the results, the aqueous extracts of Hesperozygis ringens leaves had a detrimental effect on the germination and seedling growth of the soybeans and beggarticks, evidencing their allelopathic action.

Genome-Wide Identification and Characterization of APETALA2/Ethylene-Responsive Element Binding Factor Superfamily Genes in Soybean Seed Development.

Glycine max is one of the most important grain and oil crops, and improvement of seed yield is one of the major objectives in soybean breeding. The AP2/ERF superfamily members are involved in regulating flower and seed development in many species, and therefore play key roles in seed yield. However, it is still unknown that how many AP2/ERF members were presented in the G. max genome and whether these AP2/ERF family members function in flower and seed development in G. max. Here, we identified 380 AP2/ERF superfamily genes in the G. max genome. Phylogenetic analysis showed that 323 members were grouped into the ERF family, and 49 into the AP2 family. Among the AP2 family, 14 members of the euAP2 lineage showed high identity with their orthologs, and eight member of the ANT lineage were expressed highly in the seeds. Furthermore, seven of them (GmAP2-1 to GmAP2-7) were successfully cloned and over-expressed in Arabidopsis thaliana. The transgenic Arabidopsis plants over-expressing these GmAP2 genes flowered earlier relative to the wild type control. The seed length and width, and seed area of these over-expression lines were increased compared with the wild type, and seed weight of over-expression lines of GmAP2-1, GmAP2-4, GmAP2-5, and GmAP2-6 were greater than those of the wild type. Furthermore, the seed number per silique of the over-expression lines for GmAP2 genes were not affected except GmAP2-5. Collectively, GmAP2-1, GmAP2-4, and GmAP2-6 played important roles in regulating seed weight by affecting seed length, width and area, and further controlling seed yield.

Analysis of miRNAs Targeted Storage Regulatory Genes during Soybean Seed Development Based on Transcriptome Sequencing.

Soybeans are an important cash crop and are widely used as a source of vegetable protein and edible oil. MicroRNAs (miRNA) are endogenous small RNA that play an important regulatory role in the evolutionarily conserved system of gene expression. In this study, we selected four lines with extreme phenotypes, as well as high or low protein and oil content, from the chromosome segment substitution line (CSSL) constructed from suinong (SN14) and ZYD00006, and planted and sampled at three stages of grain development for small RNA sequencing and expression analysis. The sequencing results revealed the expression pattern of miRNA in the materials, and predicted miRNA-targeted regulatory genes, including 1967 pairs of corresponding relationships between known-miRNA and their target genes, as well as 597 pairs of corresponding relationships between novel-miRNA and their target genes. After screening and annotating genes that were targeted for regulation, five specific genes were identified to be differentially expressed during seed development and subsequently analyzed for their regulatory relationship with miRNAs. The expression pattern of the targeted gene was verified by Real-time Quantitative PCR (RT-qPCR). Our research provides more information about the miRNA regulatory network in soybeans and further identifies useful genes that regulate storage during soy grain development, providing a theoretical basis for the regulation of soybean quality traits.

Dynamic Transcriptome Changes Related to Oil Accumulation in Developing Soybean Seeds.

Soybean is one of the most important oil crops in the world. Revealing the molecular basis and exploring key candidate genes for seed oil synthesis has great significance for soybean improvement. In this study, we found that oil accumulation rates and gene expression levels changed dynamically during soybean seed development. The expression levels of genes in metabolic pathways such as carbon fixation, photosynthesis, glycolysis, and fatty acid biosynthesis were significantly up-regulated during the rapid accumulation of oil in developing soybean seeds. Through weighted correlation network analysis, we identified six co-expression modules associated with soybean seed oil content and the pink module was the most positively correlated (r = 0.83, p = 7 × 10−4) network. Through the integration of differential expression and co-expression analysis, we predicted 124 candidate genes potentially affecting soybean seed oil content, including seven genes in lipid metabolism pathway, two genes involved in glycolysis, one gene in sucrose metabolism, and 12 genes belonged to transcription factors as well as other categories. Among these, three genes (GmABI3b, GmNFYA and GmFAD2-1B) have been shown to control oil and fatty acid content in soybean seeds, and other newly identified candidate genes would broaden our knowledge to understand the molecular basis for oil accumulation in soybean seeds.

Conditional and unconditional QTL analyses of seed hardness in vegetable soybean (Glycine max L. Merr.)

The seed hardness of vegetable soybean (VSB) is generally regarded as an important quality characteristic that contributes largely to its processing and mouthfeel. However, the genetic control of it is still a gray area. To identify the dynamic quantitative trait loci (QTLs) and detect the genetic relationship between seed hardness and growth period traits at the QTL level, conditional and unconditional QTL analyses were performed using a population of 184 recombinant inbred lines in 2015, 2016 and 2017. For seed hardness at stage 1 to stage 4 (W1–W4), 12 and 8 QTLs were identified by unconditional and conditional mapping, respectively. The comparative analyses between unconditional and conditional mapping uncovered three types of dynamic QTLs underlying seed hardness in the development of VSB. These results demonstrated that genes underlying seed hardness were actively expressed in the interval between stage 2 and stage 3, which was in parallel with the mass filling period in the development of soybean seeds. For seed hardness at R6 stage (W), 11 unconditional QTLs were detected. When W was conditioned on the number of days after sowing to R6 stage (DAS), the number of days after flowering to R6 stage (DAF) and flower time (FT), 11, 2, and 2 conditional QTLs were identified, respectively. Four types of unique QTLs distinguished the contributions of DAS, DAF and FT to the genetic effects of W. Of these, uqW.D1b-1 and uqW.A1-2 control W with independent of DAS or FT. Hence, they would be valuable for the selection softer seed lines, without the simultaneous depression of growth period traits even though there exist significant positive correlation between them. The loci identified by conditional and unconditional QTL analyses may be useful for the improvements of decreased seed hardness with the best balance of growth period through marker assisted selection (MAS).

Dynamic Changes of Genome-Wide DNA Methylation during Soybean Seed Development

Seed development is programmed by expression of many genes in plants. Seed maturation is an important developmental process to soybean seed quality and yield. DNA methylation is a major epigenetic modification regulating gene expression. However, little is known about the dynamic nature of DNA methylation and its effects on gene expression during plant development. Through whole-genome bisulfite sequencing, we showed that DNA methylation went through dynamic changes during seed maturation. An average of 66% CG, 45% CHG and 9% CHH contexts was methylated in cotyledons. CHH methylation levels in cotyledons changed greatly from 6% at the early stage to 11% at the late stage. Transcribed genes were approximately two-fold more likely to be differentially methylated than non-transcribed genes. We identified 40, 66 and 2136 genes containing differentially methylated regions (DMRs) with negative correlation between their expression and methylation in the CG, CHG and CHH contexts, respectively. The majority of the DMR genes in the CHH context were transcriptionally down-regulated as seeds mature: 99% of them during early maturation were down-regulated, and preferentially associated with DNA replication and cell division. The results provide novel insights into the dynamic nature of DNA methylation and its relationship with gene regulation in seed development.

Soybean Seed Development: Fatty Acid and Phytohormone Metabolism and Their Interactions.

Vegetable oil utilization is determined by its fatty acid composition. In soybean and other grain crops, during the seed development oil accumulation is important trait for value in food or industrial applications. Seed development is relatively short and sensitive to unfavorable abiotic conditions. These stresses can lead to a numerous undesirable qualitative as well as quantitative changes in fatty acid production. Fatty acid manipulation which targets a higher content of a specific single fatty acid for food or industrial application has gained more attention. Despite several successes in modifying the ratio of endogenous fatty acids in most domesticated oilseed crops, numerous obstacles in FA manipulation of seed maturation are yet to be overcome. Remarkably, connections with plant hormones have not been well studied despite their critical roles in the regulation and promotion of a plethora of processes in plant growth and development. While activities of phytohormones during the reproductive phase have been partially clarified in seed physiology, the biological role of plant hormones in oil accumulation during seed development has not been investigated. In this review seed development and numerous effects of abiotic stresses are discussed. After describing fatty acid and phytohormone metabolism and their interactions, we postulate that the endogenous plant hormones play important roles in fatty acid production in soybean seeds.

Establishment and Analysis of the mRNA Expression Patterns of ABI3-like and Storage Protein Genes during Soybean Seed Development

Abscisic acid insensitive 3 (ABI3) has a regulatory effect on the accumulation of storage proteins, as well as on the dormancy, quality, and drought tolerance of Arabidopsis seeds during development. The storage protein genes (SPGs) and relevant proteomics are already well understood; however, the mRNA expression patterns of the ABI3-like gene in soybean remain unknown. In this paper, the expression of ABI3-like and SPGs in soybean seeds was evaluated at the transcriptional level, which can provide a theoretical basis for soybean molecular breeding. Soybeans of the type ‘NC111-1’ were used as the qRT-PCR test material, in order to study the mRNA expression patterns of ABI3-like factors (GmABI3-8 and GmABI3-18), soybean SPGs (Gy1, Gy2, Gy3, Gy4, Gy5, Gy7, 7sα, 7sα', and 7sβ), and their correlation. The results demonstrate that the mRNA expression patterns of GmABI3-8 and GmABI3-18 include bimodal curves, with the two peak periods corresponding to the filling and dehydration stages of soybean, respectively. The mRNA expression patterns of all SPGs exhibited single peak curves. 7sα, Gy1, and 7sβ play major roles, and the expression level of 7sα was significantly higher than that of other SPGs (P 0.8) in the filling stage, while GmABI3-18 mRNA was highly expressed during the dehydration stage. These new details about the specific transcription factors and regulatory roles of each storage protein, as well as the specific metabolic pathway of ABI3, show that GmABI3-8 and GmABI3-18 play an important role in the development of soybean seeds. This new data will help facilitate the development of molecular breeding methods for the production of new high-protein soybean varieties.

Systems Analysis of Soybean Seed Development; Lipid Body Ontogeny

Systems Analysis of Soybean Seed Development; Lipid Body Ontogeny

Expression of VINV1, SCB1 and SCS1 genes in the seed coats of contrasting soybean seeds1

The seed coat is vital for the proper development of soybean seeds and also, after their physiological maturity, for providing protection against various biotic and abiotic factors. The aim of this work was to evaluate the relative expression of three candidate genes, VINV1, SCB1 and SCS1, which are possibly involved in seed quality, in the contrasting seed coats of four soybean genotypes. Two yellow seed coat genotypes, BMX Potencia RR and CD 202, and two black seed coat genotypes, TP and IAC, were used. Relative gene expression was evaluated by the qPCR technique for seven phases of seed coat development in the four genotypes, at 25, 30, 35, 40, 45, 50 and 55 days after anthesis. The design was completely randomised, with three replications. Data were subjected to variance analysis and the means were compared by Tukey test at 5% probability. The VINV1 and SCS1 genes exhibit greater expression in the early stages of seed coat development in the cultivar BMX Potencia RR. The SCB1 gene displays greater expression in the cultivar CD 202, also in the early stages of seed coat development. The three genes under study show greater expression in yellow seed coat genotypes.

Uso de bioativador, bioestimulante e complexo de nutrientes em sementes de soja

Novas descobertas têm estimulado a utilização de diferentes substâncias com efeitos fisiológicos, com vistas a maior expressão no desenvolvimento de culturas agrícolas. Assim, objetivou-se avaliar o efeito do tratamento de sementes com bioestimulante, bioativador e nutrientes, no desenvolvimento inicial de sementes de soja. Foram utilizados dois lotes de sementes (alto e baixo vigor) da cultivar BMX Potência RR. Os produtos testados foram inseticida com efeito bioativador, regulador de crescimento vegetal com efeito bioestimulante, complexo de nutrientes e testemunha. Em laboratório, foram avaliados os parâmetros teor de água, germinação, primeira contagem de germinação, envelhecimento acelerado, teste de frio, comprimento e massa de matéria seca de plântulas. Em casa-de-vegetação, as avaliações incluíram a emergência, índice de velocidade de emergência, comprimento e massa de matéria seca de plântulas. A eficiência dos produtos testados sofreu influência da qualidade fisiológica da semente, sendo observado efeito mais pronunciado dos produtos em lotes de alto vigor. De forma geral, o tratamento que apresentou melhor resultado no desempenho inicial foi o complexo de nutrientes, seguido pelo regulador de crescimento vegetal com efeito bioestimulante. O bioativador teve efeito negativo sobre a germinação das sementes e sobre o desenvolvimento das plântulas.

Gene expression in the lignin biosynthesis pathway during soybean seed development

The study of gene expression in plants is fundamental, and understanding the molecular mechanisms involved in important biological processes, such as biochemical pathways or signaling that are used or manipulated in improvement programs, are key for the production of high-quality soybean seeds. Reports related to gene expression of lignin in seeds are scarce in the literature. We studied the expression of the phenylalanine ammonia-lyase (PAL), cinnamate 4-hydroxylase, 4-hydroxycinnamate 3-hydroxylase, and cinnamyl alcohol dehydrogenase genes involved in lignin biosynthesis during the development of soybean (Glycine max L. Merrill) seeds. As the endogenous control, the eukaryotic elongation factor 1-beta gene was used in two biological replicates performed in triplicate. Relative quantitative expression of these genes during the R4, R5, R6, and R7 development stages was analyzed. Real-time polymerase chain reaction was used for the gene expression study. The analyses were carried out in an ABI PRISM 7500 thermocycler using the comparative Ct method and SYBR Green to detect amplification. The seed samples at the R4 stage were chosen as calibrators. Increased expression of the cinnamate-4-hydroxylase and PAL genes occurred in soybean seeds at the R5 and R6 development stages. The cinnamyl alcohol dehydrogenase gene was expressed during the final development phases of soybean seeds. In low-lignin soybean cultivars, the higher expression of the PAL gene occurs at development stages R6 and R7. Activation of the genes involved in the lignin biosynthesis pathway occurs at the beginning of soybean seed development.

The Circadian Clock‐controlled Transcriptome of Developing Soybean Seeds

A number of metabolic and physiological processes in plants are controlled by the circadian clock, which enables a plant to anticipate daily changes in the environment. Relatively little is known about circadian rhythms in developing seeds, which may be important for determining the extent and timing of nutrient storage in grain. Microarray expression profiling was used to identify genes expressed in developing soybean (Glycine max) seeds that are controlled by the circadian clock. Genes with predicted functions in protein synthesis, fatty acid metabolism, and photosynthesis totaling 1.8% of the mRNAs detected in seed were found to be expressed in a circadian rhythm. Known circadian and light‐controlled promoter elements were identified as over‐represented in the promoters of clock‐controlled seed genes, with the over‐represented elements varying according to the phase of circadian expression. A subset of circadian‐regulated genes were found to be expressed in different phases in developing seeds with respect to leaves from the same plants, many of which have roles in photosynthesis and carbon metabolism. These results help to characterize the genes and processes in seeds that may be regulated by the circadian clock, and provide some insight into organ‐specific phasing of clock controlled gene expression.