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Genome-Wide Association Studies (GWAS) identify tagging variants in the genome that are statistically associated with the phenotype because of their linkage disequilibrium (LD) relationship with the causative mutation (CM). When both low-density genotyped accession panels with phenotypes and resequenced data accession panels are available, tagging variants can assist with post-GWAS challenges in CM discovery.Our objective was to identify additional GWAS evaluation criteria to assess correspondence between genomic variants and phenotypes, as well as enable deeper analysis of the localized landscape of association.We used genomic variant positions as Synthetic phenotypes in GWAS that we named "Synthetic phenotype association study" (SPAS). The extreme case of SPAS is what we call an "Inverse GWAS" where we used CM positions of cloned soybean genes. We developed and validated the Accuracy concept as a measure of the correspondence between variant positions and phenotypes.The SPAS approach demonstrated that the genotype status of an associated variant used as a Synthetic phenotype enabled us to explore the relationships between tagging variants and CMs, and further, that utilizing CMs as Synthetic phenotypes in Inverse GWAS illuminated the landscape of association. We implemented the Accuracy calculation for a curated accession panel to an online Accuracy calculation tool (AccuTool) as a resource for gene identification in soybean. We demonstrated our concepts on three examples of soybean cloned genes. As a result of our findings, we devised an enhanced "GWAS to Genes" analysis (Synthetic phenotype to CM strategy, SP2CM). Using SP2CM, we identified a CM for a novel gene.The SP2CM strategy utilizing Synthetic phenotypes and the Accuracy calculation of correspondence provides crucial information to assist researchers in CM discovery. The impact of this work is a more effective evaluation of landscapes of GWAS associations.

The negative association between protein and oil production in soybean (Glycine max) seed is well-documented. However, this inverse relationship is based primarily on the composition of mature seed, which reflects the cumulative result of events over the course of soybean seed development and therefore does not convey information specific to metabolic fluctuations during developmental growth regimes. In this study, we assessed maternal nutrient supply via measurement of seed coat exudates and metabolite levels within the cotyledon throughout development to identify trends in the accumulation of central carbon and nitrogen metabolic intermediates. Active metabolic activity during late seed development was probed through transient labeling with 13C substrates. The results indicated: (1) a drop in lipid contents during seed maturation with a concomitant increase in carbohydrates, (2) a transition from seed filling to maturation phases characterized by quantitatively balanced changes in carbon use and CO2 release, (3) changes in measured carbon and nitrogen resources supplied maternally throughout development, (4) 13C metabolite production through gluconeogenic steps for sustained carbohydrate accumulation as the maternal nutrient supply diminishes, and (5) oligosaccharide biosynthesis within the seed coat during the maturation phase. These results highlight temporal engineering targets for altering final biomass composition to increase the value of soybeans and a path to breaking the inverse correlation between seed protein and oil content.

Background Soybean is an economically important crop which flowers predominantly in response to photoperiod. Several major loci controlling the quantitative trait for reproductive timing have been identified, of which allelic combinations at three of these loci, E1, E2, and E3, are the dominant factors driving time to flower and reproductive period. However, functional genomics studies have identified additional loci which affect reproductive timing, many of which are less understood. A better characterization of these genes will enable fine-tuning of adaptation to various production environments. Two such genes, E1La and E1Lb, have been implicated in flowering by previous studies, but their effects have yet to be assessed under natural photoperiod regimes. Results Natural and induced variants of E1La and E1Lb were identified and introgressed into lines harboring either E1 or its early flowering variant, e1-as. Lines were evaluated for days to flower and maturity in a Maturity Group (MG) III production environment. These results revealed that variation in E1La and E1Lb promoted earlier flowering and maturity, with stronger effects in e1-as background than in an E1 background. The geographic distribution of E1La alleles among wild and cultivated soybean revealed that natural variation in E1La likely contributed to northern expansion of wild soybean, while breeding programs in North America exploited e1-as to develop cultivars adapted to northern latitudes. Conclusion This research identified novel alleles of the E1 paralogues, E1La and E1Lb, which promote flowering and maturity under natural photoperiods. These loci represent sources of genetic variation which have been under-utilized in North American breeding programs to control reproductive timing, and which can be valuable additions to a breeder’s molecular toolbox.

The uses of vegetable oils are determined by functional properties arising from their chemical composition. Soybean oil was previously used in margarines and baked foods after partial hydrogenation to achieve heat and oxidative stability. This process, however, generates trans fats that are now excluded from food use because of cardiovascular health risks. Also present in soybean oil are the anti-oxidant tocopherols, with α-tocopherol (vitamin E) typically present as a minor component compared to γ-tocopherol. Genetic improvement of the fatty acid profile and tocopherol profile is an attractive solution to increase the functional and health qualities of soybean oil. The objective of this research was to develop resources to directly select with molecular markers for the elevated vitamin E trait in soybean oil and to use a molecular breeding approach to combine elevated vitamin E with the high oleic/low linolenic acid seed oil trait that improves oil functionality and nutrition. New soybean germplasm was developed from the molecular breeding strategy that selected for alleles of six targeted genes. Seed oil from the novel soybean germplasm was confirmed to contain increased vitamin E α-tocopherol along with a high oleic acid/low linolenic acid profile.

Single nucleotide polymorphisms (SNPs) and insertions/deletions (Indels) are widely spread across all chromosomes of the genome and act as biological markers, which aid in identification of genes associated with traits or phenotypes. With the advances in next-generation sequencing (NGS) technology, large amounts of SNPs and Indels data have become available, making it difficult to effectively perform analysis across multiple samples and intuitively integrate, compare and/or visualize them simultaneously. Genome-wide association studies (GWAS) is a widely used method to find genetic variations associated with a trait, but it lacks an efficient way to investigate genomic variant functions. To tackle these issues, we have developed SNPViz v2.0, a web-based tool to visualize large-scale haplotype blocks with detailed SNPs and Indels grouped by their chromosomal coordinates, along with their overlapping gene models, phenotype to genotype accuracies, Gene Ontology (GO) annotations, protein families (Pfam) annotations, genomic variant annotations, and their functional effects. Moreover, SNPViz v2.0 integrates several large scale soybean SNPs and Indels datasets including G. Soja, GWAS, NAM41, USB-15x, USB-40x, MSMC and Zhou302, available from multiple studies. SNPViz v2.0 is deployed for all organisms and available in both SoyKB and KBCommons frameworks. For soybean data only, the SNPViz 2.0 is publicly available at http://soykb.org/SNPViz2/. For other organisms such as Arabidopsis thaliana, Mus musculus and Zea mays, SNPViz 2.0 is publicly available in their respective knowledge bases at https://kbcommons.org.

The inverse correlation between protein and oil production in soybeans is well-documented; however, it has been based primarily on the composition of mature seeds. Though this is the cumulative result of events over the course of soybean seed development, it does not convey information specific to metabolic fluctuations during developmental growth regimes. Maternal nutrient supply via seed coat exudate measurements and metabolite levels within the cotyledon were assessed across development to identify trends in the accumulation of central carbon and nitrogen metabolic intermediates. Active metabolic operation during late seed development was probed through transient labeling with13C substrates. The results indicated: i) a drop in lipid during seed maturation with a concomitant increase in carbohydrates, ii) a transition from seed filling to maturation phase characterized by quantitatively balanced changes in the carbon use and CO2release, iii) changes in measured carbon and nitrogen resources supplied maternally over development, iv)13C metabolites processed through gluconeogenesis towards sustained carbohydrate accumulation as the maternal nutrient supply diminishes, and v) oligosaccharide biosynthetic metabolism during seed coat senescence at maturation. These results highlight temporal engineering targets for altering final biomass composition to increase the value of soybeans and a path to breaking the inverse seed protein and oil correlation.One-sentence summaryAssessment of temporal changes in metabolism during soybean seed development indicated that lipid turnover during maturation contributes carbon for gluconeogenic production of carbohydrates.

Stearic acid is a relatively minor component of soybean [Glycine max (L.) Merr.] seed oil. Increasing the stearic acid concentration is desired for industrial and food applications, as well as enhanced oxidative stability of the oil. Several ethyl methane sulfonate (EMS)-induced mutants with elevated seed stearic acid concentration have been reported in soybean. Such mutants were often found to carry alterations in the GmSACPD-C gene encoding Δ9-stearoyl-acyl carrier protein-desaturase, the enzyme that catalyzes the conversion of stearic acid to oleic acid in the fatty acid biosynthetic pathway. In the present study, we identified a new GmSACPD-C allele in an EMS-induced high-stearic-acid mutant PE980. The mutant showed a seed stearic acid concentration of ∼160 g kg⁻¹, fourfold that of the wild-type ‘Pungsannamul’. A single base substitution from guanine to adenine at genomic position 703 of first exon of the GmSACPD-C gene was identified through sequencing, and validated by cosegregation analysis of 167 F₂ plants using an allele-specific SimpleProbe marker. The mutation caused a valine to methionine (+70 position) change in the predicted GmSACPD-Cprotein sequence. The sequence alignments and three-dimensional structure modeling of GmSACPD-C showed that the V70M mutation affects substrate binding and dimerization to reduce enzyme activity. Since the mutant showed significant reductions in seed yield compared with the wild type, marker-assisted backcrossing may be needed to reduce the yield gap.

Oil value is determined by the functional qualities imparted from the fatty acid profile. Soybean oil historically had excellent utilization in both food and industrial uses, but the need to increase the stability of the oil without negative health consequences has led to a significant decline in soybean oil use. One solution to oil stability is to alter the fatty acid profile of the oil to have high oleic acid (greater than 70%) and lower linolenic acid. Other fatty acid profile changes intended to meet specific market needs are to lower the total saturated fatty acids, palmitic acid plus stearic acid; in addition, an oil high in stearic acid is desired for some baking applications. Mutant alleles in different soybean genes encoding key enzymes in the fatty acid biosynthesis pathways have been identified to address each of the desired fatty acid profiles in soybean oil. The objective of this research was to determine the interaction of the high oleic acid oil trait in combination with various mutant alleles controlling fatty acid profiles. Soybean lines containing high oleic acid allele combinations plus other fatty acid modifying alleles were created, and seed was produced in field environments in two locations in 2012 and four locations in 2013. Seed samples were analyzed for fatty acid components of the oil fraction and subjected to statistical analyses for the effects on palmitic, stearic, oleic, linoleic, and linolenic acids. The results identified allele combinations that met the target fatty acid profile thresholds and were stable across environments.

A high ω-6/ω-3 fatty acid ratio in the soybean seed adversely affects human health. The objective of the present study was to improve the fatty acid biosynthesis to reduce the ω-6/ω-3 ratio by combining the FAD2-1A and FAD2-1B mutant alleles with α-linolenic acid (ω-3) related alleles from wild soybean. The F2 population comprising 2320 F 2:3 lines developed from S08-14717 × PI 483463 cross exhibited significant variation for fatty acid components. Of these, 114 lines were advanced to the F 5:6 generation and genotyped for FAD2-1A and FAD2-1B alleles. The lines carrying mutant FAD2-1A and FAD2-1B alleles showed ~ 761 g kg−1 oleic, and ~ 50 g kg−1 linoleic acids, which reduced ω-6/ω-3 ratios to ~ 0.6. Conversely, the lines carrying FAD2-1A or FAD2-1B mutant alleles had 267 or 399 g kg−1 oleic, 327 or 471 g kg−1 linoleic, and 120 or 130 g kg−1 α-linolenic acids concentration, respectively. The elevated α-linolenic acid resulted in the reduction of ω-6/ω-3 ratios in the range 2.5–3.9. The present study demonstrated that combining FAD2 mutant alleles with α-linolenic acid-related alleles from wild soybean reduces the seed ω-6/ω-3 ratio.

Background Soybean is native to the temperate zones of East Asia. Poor yields of soybean in West African countries may be partially attributed to inadequate adaptation of soybean to tropical environments. Adaptation will require knowledge of the effects of allelic combinations of major maturity genes (E1, E2, and E3) and stem architecture. The long juvenile trait (J) influences soybean flowering time in short, ~ 12 h days, which characterize tropical latitudes. Soybean plant architecture includes determinate or indeterminate stem phenotypes controlled by the Dt1 gene. Understanding the influence of these genetic components on plant development and adaptation is key to optimize phenology and improve soybean yield potential in tropical environments. Results Soybean lines from five recombinant inbred populations were developed that varied in their combinations of targeted genes. The soybean lines were field tested in multiple environments and characterized for days to flowering (DTF), days to maturity (DTM), and plant height in locations throughout northern Ghana, and allelic combinations were determined for each line for associating genotype with phenotype. The results revealed significant differences based on genotype for DTF and DTM and allowed the comparison of different variant alleles of those genes. The mutant alleles of J and E1 had significant impact on DTF and DTM, and alleles of those genes interacted with each other for DTF but not DTM. The Dt1 gene significantly influenced plant height but not DTF or DTM. Conclusions This research identified major and minor effect alleles of soybean genes that can be combined to control DTF, DTM, and plant height in short day tropical environments in Ghana. These phenotypes contribute to adaptation to a low latitude environment that can be optimized in a soybean breeding program with targeted selection of desired allele combinations. The knowledge of the genetic control of these traits will enhance molecular breeding to produce optimally adapted soybean varieties targeted to tropical environments.

Two independent variant raffinose synthase 3 ( RS3 ) alleles produced an equivalent phenotype and implicated the gene as a key contributor to soybean seed carbohydrate phenotype. Soybean is an important crop because the processed seed is utilized as a vegetable oil and a high protein meal typically used in livestock feeds. Raffinose and stachyose, the raffinose family of oligosaccharides (RFO) carbohydrate components of the seed, are synthesized in developing soybean seeds from sucrose and galactinol. Sucrose is considered positive for metabolizable energy, while RFO are anti-nutritional factors in diets of monogastric animals such as humans, poultry, and swine. To increase metabolizable energy available in soybean seed meal, prior research has been successful in deploying variant alleles of key soybean raffinose synthase (RS) genes leading to reductions or near elimination of seed RFO, with significant increases in seed sucrose. The objective of this research was to investigate the specific role of variants of the RS3 gene in a genomic context and improve molecular marker-assisted selection for the ultra-low (UL) RFO phenotype in soybean seeds. The results revealed a new variant of the RS3 allele (rs3 snp5, rs3 snp6) contributed to the UL RFO phenotype when mutant alleles of RS2 were present. The variant RS3 allele identified was present in about 15% of a small set of soybean cultivars released in North America. A missense allele of the RS3 gene (rs3 G75E) also produced the UL RFO phenotype when combined with mutant alleles of RS2. The discoveries reported here enable direct marker-assisted selection for an improved soybean meal trait that has the potential to add value to soybean by improving the metabolizable energy of the meal.

Natural esters (triglycerides) possess desirable attributes such as the higher fire points and better environmental compatibility relative to petroleum-derived insulating mineral oil, as a result, the use of such natural esters is on the rise as insulating and heat transfer fluids in electrical devices such as transformers. However, natural esters differ significantly in chemical composition from mineral oils and are more prone to oxidation than mineral oils. High susceptibility of natural esters can be addressed in part through the use of natural esters with higher monounsaturated fatty acid-high oleic natural esters. Recently several high oleic acid soybeans have been introduced commercially. These include Plenish® (Pioneer / Dupont) Vistive Gold® (Monsanto / Bayer) and Soyleic® (Missouri Soybean Association). Both Plenish® and Vistive Gold® are obtained from GMO soybeans, while Soyleic® has been developed through more traditional plant breeding. The oleic acid content of the three natural esters was found to be 79%, 71%, and 81% respectively. While polyunsaturated fatty acid (linoleic and linolenic) contents of the three natural esters were found to be 10.3%, 22.5%, and 7.8% respectively. High oleic soybean oils along with canola oil with oleic content of 75% and polyunsaturated fatty acid content of 17.2% and generic soybean with oleic content of 24.5% and polyunsaturated fatty acid content of 60.0% were subjected to accelerated aging under conditions outlined in IEC 62770 2013. Test results showed that natural esters with higher oleic and lower polyunsaturated acid contents were more stable than the natural esters with lower oleic and higher polyunsaturated content. This stability was reflected in little or no increase in kinematic viscosity and little increase in dissolved acidity. These natural esters are therefore more suitable for dielectric applications where long-term stability is of paramount concern.

Soybean plays an important role in seed oil production for foods and industrial products in the USA. Chemical hydrogenation of commodity soybean oil increased functionality but unavoidably created trans fat which has been linked to many health issues in humans. An alternative to using hydrogenation of the oil to enhance oxidative stability is to genetically increase the level of oleic acid in the seed oil. Our goal was to develop a soybean germplasm as a source for more functional soybean oil with a high oleic acid, increased stearic acid, and low linolenic acid profile utilizing new and existing variant alleles of key fatty acid desaturase genes. Using forward genetics, novel alleles of FAD2-1A were discovered and characterized from a mutant soybean population already containing elevated seed stearic acid content. One of the two new FAD2-1A alleles, when combined with existing mutant alleles of FAD2-1B created a high oleic acid seed oil phenotype in soybean germplasm lines. The other new FAD2-1A allele produced lower and more variable oleic acid levels when combined with existing mutant alleles of FAD2-1B. Seed stearic acid increased to ~ 10–11% in lines containing combinations of FAD2-1A and FAD2-1B mutant alleles plus the SACPD-C missense mutant alleles; however, the increase in stearic acid was associated with a decrease in the oleic acid content and did not meet the target of at least 20% stearic acid in the seed oil. In addition, existing FAD3 mutant alleles were incorporated into the novel high oleic acid and high oleic/elevated stearic acid soybean lines with four or five targeted alleles combined (FAD2-1A, FAD2-1B, FAD3A, FAD3C, and SACPD-C) to create soybean germplasm with more functional soybean oil. This study is beneficial for improving the quality of soybean oil based on nutritional value and oxidative stability.

The mature seeds of soybean (Glycine max L. Merr) are a valuable source of high-quality edible lipids and protein. Despite dramatic breeding gains over the past 80 years, soybean oil continues to be oxidatively unstable, due to a high proportion of polyunsaturated triacylglycerols. Until recently, the majority of soybean oil underwent partial chemical hydrogenation. Mounting health concerns over trans fats, however, has increased breeding efforts to introgress mutant and biotechnological genetic alterations of soybean oil composition into high-yielding lines. As a result, there is an ongoing need to characterize fatty acid composition in a rapid, inexpensive and accurate manner. Gas chromatography is the most commonly used method, but near-infrared reflectance spectroscopy (NIRS) can be calibrated to non-destructively phenotype various seed compositions accurately and at a high throughput. Here we detail development of NIRS calibrations using intact seeds for every major soybean fatty acid breeding goal over an unprecedented range of oil composition. The NIRS calibrations were shown to be equivalent to destructive chemical analysis, and incorporation into a soybean phenotyping operation has the potential to dramatically reduce cost and accelerate phenotypic analysis.

Soybeans (Glycine max (L.) Merr.) with black seed coats and green cotyledons are rich in anthocyanins and chlorophylls known as functional nutrients, antioxidants and compounds with anticarcinogenic properties. Understanding the genetic diversity of germplasm is important to determine effective strategies for improving the economic traits of these soybeans. We aimed to analyze the genetic diversity of 470 soybean accessions by 6K single nucleotide polymorphic loci to determine genetic architecture of the soybeans with black seed coats and green cotyledons. We found soybeans with black seed coats and green cotyledons showed narrow genetic variability in South Korea. The genotypic frequency of the d1d2 and psbM variants for green cotyledon indicated that soybean collections from Korea were intermingled with soybean accessions from Japan and China. Regarding the chlorophyll content, the nuclear gene variant pair d1d2 produced significantly higher chlorophyll a content than that of chloroplast genome psbM variants. Among the soybean accessions in this study, flower color plays an important role in the anthocyanin composition of seed coats. We provide 36 accessions as a core collection representing 99.5% of the genetic diversity from the total accessions used in this study to show potential as useful breeding materials for cultivars with black seed coats and green cotyledons.

Pod shattering is an ancestral trait that promotes seed dispersal; however, shattering can have substantial yield losses in cultivated soybean. During the improvement process, American soybean breeders virtually eliminated the shatter phenotype for released varieties, but in other countries, such as Ghana, shatter persists. The objective of our research was to find a molecular tool to implicate genetic shatter susceptibility, validate its usefulness, and apply this knowledge to identify shattering potential in parental lines. Previous research revealed the gene Pdh1 on chromosome 16 plays a crucial role in determining the shatter phenotype. A perfect molecular marker assay was developed to detect alleles of the Pdh1 gene. A genome-wide association study (GWAS) was performed using the Pdh1 allele status as a phenotype and identified a highly associated marker in the SoySNP50K array. Soybean accessions from the National Plant Germplasm System (GRIN-NPGS) with shatter score and SoySNP50K data were evaluated to determine the impact of the predicted Pdh1 alleles on early and late pod shattering. An online tool was developed to enable researchers to query the GRIN collection for the predicted Pdh1 allele status. Lines from an African soybean germplasm collection were analyzed, and it was determined that 22.5% of lines had the shatter-susceptible alleles of Pdh1; two of seven Ghanaian released soybean varieties had the shatter-susceptible alleles of Pdh1. Soybean breeding programs that access germplasm from the GRIN or the African collection can utilize these resources to eliminate the Pdh1 effects on pod shatter and thus improve yield potential.

The soybean eu3-a mutant (formerly, eu3-e1) lacks all ureolytic activity. Eu3 encodes urease accessory (Ni insertion) protein, UreG. Eu3 (Glycine max v1.1 Glyma08g08970.1) is the only UreG-encoding gene in the soybean genome. Here we show that the eu3-a lesion is a 2.4 kb deletion, beginning 50 bp upstream of the transcription start, and covering 94 % of the deduced amino acid sequence of UreG, explaining the urease activity-null phenotype of eu3-a. We established near-isogenic lines (NILs), eu3-a versus Eu3, and confirmed a striking accumulation of endogenous urea in eu3-a and none in Eu3, and the greater resistance of eu3-a leaf discs to external urea. However, incubation of discs in light alleviated urea sensitivity in Eu3, apparently by providing C skeletons for urea-released NH3. Hence, urea, endogenous and exogenous, is apparently less toxic than urea-derived NH3. eu3-a and Eu3 plants, however, did not differ in seed Ni content, nor in pool sizes of metabolites that generate urea via arginine. The NILs differed in patterns of nodulation by Bradyrhizobium japonicum and by Bradyrhizobium elkanii. Ureases have been reported to be fungitoxic. However, eu3-a and Eu3 NIL seedlings were equally sensitive to the pathogenic Basidiomycete fungus, Rhizoctonia solani. Since eu3-a accumulates at least some apo-urease, we examined a specific anti-fungal role of the seed urease in seedlings of a second NIL pair, differing in the presence (Eu1) and absence (eu1-a) of the seed urease. Again, this second NIL pair exhibited equal sensitivity to R. solani. To our knowledge, we report only the second test of fungal resistance influenced by urease levels in intact plants.

Background Achieving appropriate maturity in a target environment is essential to maximizing crop yield potential. In soybean [Glycine max (L.) Merr.], the time to maturity is largely dependent on developmental response to dark periods. Once the critical photoperiod is reached, flowering is initiated and reproductive development proceeds. Therefore, soybean adaptation has been attributed to genetic changes and natural or artificial selection to optimize plant development in specific, narrow latitudinal ranges. In North America, these regions have been classified into twelve maturity groups (MG), with lower MG being shorter season than higher MG. Growing soybean lines not adapted to a particular environment typically results in poor growth and significant yield reductions. The objective of this study was to develop a molecular model for soybean maturity based on the alleles underlying the major maturity loci: E1, E2, and E3. Results We determined the allelic variation and diversity of the E maturity genes in a large collection of soybean landraces, North American ancestors, Chinese cultivars, North American cultivars or expired Plant Variety Protection lines, and private-company lines. The E gene status of accessions in the USDA Soybean Germplasm Collection with SoySNP50K Beadchip data was also predicted. We determined the E allelic combinations needed to adapt soybean to different MGs in the United States (US) and discovered a strong signal of selection for E genotypes released in North America, particularly the US and Canada. Conclusions The E gene maturity model proposed will enable plant breeders to more effectively transfer traits into different MGs and increase the overall efficiency of soybean breeding in the US and Canada. The powerful yet simple selection strategy for increasing soybean breeding efficiency can be used alone or to directly enhance genomic prediction/selection schemes. The results also revealed previously unrecognized aspects of artificial selection in soybean imposed by soybean breeders based on geography that highlights the need for plant breeding that is optimized for specific environments. Electronic supplementary material The online version of this article (doi:10.1186/s12870-017-1040-4) contains supplementary material, which is available to authorized users.

The fatty acid composition of vegetable oil is becoming increasingly critical for its ultimate functionality and utilization in foods and industrial products. Partial chemical hydrogenation of soybean [Glycine max (L.) Merr.] oil increases oxidative stability and shelf life but also results in the introduction of trans fats as an unavoidable byproduct. Due to mandatory labeling of consumer products containing trans fats, conventional soybean oil has lost the ability to deliver the most appropriate economical functionality and oxidative stability, particularly for baking applications. Genetic improvement of the fatty acid profile of soybean oil is one method of meeting these new requirements for oil feedstocks. In this report, we characterized three mutant genetic loci controlling the saturated fatty acid content of soybean oil: two genes additively reduce palmitic acid content (fap1 and fap3-ug), and one gene independently elevates stearic acid content (fas). We identified a new null allele of fap3-ug/GmFATB1A (derived from line ELLP2) present in line RG3. The splicing defect mutation in a beta-ketoacyl-[acyl-carrier-protein] synthase III candidate gene located in the region mapped to fap1, derived originally from ethyl methane sulphonate mutant line C1726 (Cardinal et al. in Theor Appl Genet 127:97–111, 2014), was also present in line RG3. We also utilized the elevated stearic acid line RG7, which has previously been shown to contain novel mutant fas/SACPD-C alleles encoding stearoyl-acyl carrier protein desaturase (Boersma et al. in Crop Sci 52:1736–1742, 2012). Molecular marker assays have been developed to track these causative mutations and understand their contributions to seed oil fatty acid profiles in a recombinant inbred line population segregating for fap1, fap3-ug, and fas alleles.

Soybean is one of the most important oil crops worldwide, and reducing the linolenic acid content of soybean oil will provide increased stability of the oil to consumers and limit the amount of trans fat in processed foods. The linolenic content in soybean seed is controlled by three fatty acid desaturase (FAD) three enzymes, FAD3A, B, and C. The soybean lines with 1 % linolenic acid content which are widely used in breeding for reduced linolenic acid in the USA have mutations in each of the three FAD genes derived from lines A5 (deletion of FAD3A), A26, and A23 (missense mutations in FAD3B and C, respectively). Although soybean line A5 has been released for 30 years, the extent and definition of the deletion of the FAD3A gene has not been characterized, which has prevented researchers from designing robust molecular markers for effective marker-assisted selection (MAS). Using a PCR-based genomic strategy, we have identified a 6.4-kbp deletion of the FAD3A gene in A5 and developed a TaqMan detection assay by targeting the deletion junction in A5, which could be used to distinguish the homozygotes and heterozygotes of the gene. In addition, based on mutant single nucleotide polymorphisms in FAD3B and FAD3C identified in A26 and A23, respectively, we have also developed TaqMan assays for high-throughput MAS. The TaqMan assays have proven to be a very effective platform for detecting the mutant FAD3 alleles and thus will greatly facilitate high-throughput MAS for development of soybean lines with reduced linolenic acid content.

Soybean [Glycine max (L.) Merr] with increased oleic acid is desirable to improve oxidative stability and functionality of soybean seed oil. Recently, soybean genotypes with high oleic acid (≥70 %) were developed by breeding programs. Efficient and effective identification of high oleic acid soybean genotypes using non-destructive near infrared reflectance (NIR) on whole seeds would greatly enhance progress in breeding programs. The objective of this study was to develop a calibration equation for NIR determination of high oleic acid from single soybean seeds. A total of 600 intact, single F2 seeds were scanned by NIR. Spectral data were collected between 400 and 2,500 nm at 2 nm intervals. The relationship between NIR spectral patterns of each soybean seed and its oleic acid content was examined. The best predicted equations for oleic acid were selected on the basis of minimizing the standard error of cross-validation and increasing the coefficient of determination. Validation demonstrated that the equations for determining total oleic acid and over 50 % oleic acid content had high predictive ability (r2 = 0.91 and r2 = 0.99, respectively). To validate the newly developed equation, F2 seeds from a different genetic background were tested. Again, high oleic acid from single soybean seeds was accurately predicted from various genetic backgrounds. Therefore, applying the calibration equations to NIR will be useful to rapidly and efficiently select high oleic acid soybean genotypes in breeding programs.

High-throughput/low-cost/low-tech methods for phytic acid determination that are sufficiently accurate and reproducible would be of value in plant genetics, crop breeding and in the food and feed industries. Variants of two candidate methods, those described by Vaintraub and Lapteva (Anal Biochem 175:227–24, 1988; “VL” methods) and Huang and Lantzsch (J Sci Food Agric 34:1423–1426, 1983; “HL” methods), were evaluated. The primary concern with these methods is that, due to interference of matrix constituents including inorganic P, they can overestimate phytic acid and are ineffective at low levels of phytic acid. Twelve seed flours, representing lines of soybean, maize, barley and dry bean, containing a wide range of phytic acid levels, were analyzed by a minimum of eight cooperating laboratories using three variants of the VL method and two variants of the HL method. No method had consistently acceptable (˂2.0”) “Horwitz ratios”, a measure of reproducibility, although some treatments approached that. For example, one variant of the VL method when used to assay a soybean flour with a “standard” level of phytic acid had a Horwitz ratio of 2.15. Some variants of the VL method were adequate for analyses of cereal grains regardless of phytic acid level but none accurately measured phytic acid when at low levels in soybean flours. One variant of the HL method in which the 0.2 N HCl extraction media is modified to contain 10% Na2SO4, did accurately measure phytic acid levels in both cereal and legume flours regardless of endogenous phytic acid levels or matrix constituents.

Soybeans are an important source of protein-rich meal for livestock feed formulations. Recent changes in the cost of commodity-based sources of metabolizable energy (ME) inputs has put pressure on soybean meal to deliver both protein and ME in feed formulations. The non-oil fraction of soybean contains approximately 12% soluble carbohydrates, principally sucrose, raffinose, and stachyose. Of these carbohydrates, only sucrose is positive for ME. Both raffinose and stachyose, belonging to the raffinose family of oligosaccharides (RFOs), are considered antinutritional because of the negative consequences of their fermentation in the gut of monogastric animals when RFOs are consumed in the diet. Therefore, there is an interest in improving soybean seed composition so that it contains higher ME and fewer antinutritional components by increasing the sucrose content while lowering the RFOs. Several soybean lines have been discovered that contain altered levels of RFOs, and recent molecular genetic investigations have shown the phenotype to be caused by mutations in a raffinose synthase 2 (RS2) gene encoding the enzyme that is the committed step for RFO biosynthesis. The objective of this research was to determine the variation in carbohydrate profile for different soybean lines grown in a single location containing one of three different alleles of the RS2 gene. The results indicate that, although there is variation in the carbohydrate profiles for each line, different lines with the same RS2 genotype tend to produce a characteristic carbohydrate profile. Although the carbohydrate profile for each RS2 genotype class was consistent in different genetic backgrounds under two conditions grown at one location, more research will be necessary to determine the environmental stability of the carbohydrate profiles in multiple locations over different years.

Introduction, James Orf Classical Breeding and Genetics of Soybean, Andrew M. Scaboo, Pengyin Chen, David A. Sleper, and Kerry M. Clark Identification of Genes Underlying Simple Traits in Soybean, David Lightfoot Molecular Genetic Linkage Maps of Soybean, Sachiko Isobe and Satoshi Tabata Molecular Mapping of Quantitative Trait Loci, Dechun Wang and David Grant Molecular Breeding, David R. Walker, Maria J. Monteros, and Jennifer L. Yates Map-Based Cloning of Genes and QTLs in Soybean, Madan K. Bhattacharyya Candidate Gene Analysis of Mutant Soybean Germplasm, R.E. Dewey and P. Zhang Functional Genomics-Transcriptomics in Soybean, Sangeeta Dhaubhadel, Frederic Marsolais, Jennifer Tedman-Jones, and Mark Gijzen The Draft Soybean Genome Sequence, Jeremy Schmutz, Steven B. Cannon, Therese Mitros, Will Nelson, Shengqiang Shu, David Goodstein, and Dan Rokhsar Soybean Comparative Genomics, Jianxin Ma, Steven Cannon, Scott A. Jackson, and Randy C. Shoemaker Role of Bioinformatics as a Tool, Julie M. Livingstone, Kei Chin C. Cheng, and Martina V. Stromvik Soybean Proteomics, Savithiry S. Natarajan, Thomas J. Caperna, Wesley M. Garrett, and Devanand L. Luthria Metabolomics Approach in Soybean, Takuji Nakamura, Keiki Okazaki, Noureddine Benkeblia, Jun Wasaki, Toshihiro Watanabe, Hideyuki Matsuura, Hirofumi Uchimiya, Setsuko Komatsu, and Takuro Shinano Soybean Future Prospects, Ed Ready

Soybean (Glycine max (L.) Merr.) oil from conventional cultivars typically contains ~30 g kg −1 stearic acid of the total seed oil. Increased stearic acid concentration in the seed oil of soybeans is desirable for certain food and industrial uses. To date a small number of mutants have been developed with increased stearic acid levels three to six times that of normal. At least two such lines were found to possess separate mutations in the delta-9- stearoyl-acyl carrier protein desaturase C gene (SACPD-C) that dramatically increased seed stearic acid concentration. We now report additional independent mutations in this gene that increase seed stearic acid levels of the soybean germplasm line RG7 to ~116 g kg −1 . An F 5 recombinant inbred line (RIL) population was developed to determine the relationship between the RG7 SACPD-C mutation and stearic acid concentration. Transgressive segregation in the progeny of the cross between mutant lines RG7 and RG2 (low palmitic acid) further increases seed stearic acid to almost 180 g kg −1 , suggesting the presence of yet another gene having a signifi cant effect on stearic acid accumulation. We also discerned an independent, presumably allelic, mutation within the SACPD-C gene in line RG8, which has about 106 g kg −1 stearic acid. Molecular markers diagnostic for the RG7 and RG8 SACPD-C mutations were developed, enabling rapid selection for these mutations in the development of cultivars with increased seed stearic acid content in the seed oil.

Elevating oleic acid in seed oil improves oxidative stability and is desirable for expanding edible and industrial uses of soybean [Glycine max (L.) Merr.]. Soybean lines with up to 800 g kg –1 oleic acid concentration were developed by combining a recessive mutant allele at the FAD2-1A locus (Glyma10 g42470) with either of two different recessive mutant alleles at the FAD2-1B locus (Glyma20 g24530). However, oleic acid concentration for some higher oleic acid genotypes can be affected by growing conditions. The objective of this study was to evaluate the stability of oleic acid concentration among 10 soybean lines with elevated oleic acid and eight checks with typical oleic acid levels over six environments. High oleic genotypes with genes combined from crossing M23 with the FAD2-1AΔ mutant and PI567189A with the FAD2-1B mutant I143T accumulated less oleic acid, and means for oleic acid concentration ranged from 551 to 729 g kg –1 across environments compared to 202 to 263 g kg –1 in the check cultivars. Signifi cant variation among environments indicated that genes from different sources can affect both concentration and stability of oleic acid of these soybean genotypes grown in different environments.

The alteration of fatty acid profiles in soybean to improve soybean oil quality has been a long-time goal of soybean researchers. Soybean oil with elevated oleic acid is desirable because this monounsaturated fatty acid improves the nutrition and oxidative stability of soybean oil compared to other oils. In the lipid biosynthetic pathway, the enzyme fatty acid desaturase 2 (FAD2) is responsible for the conversion of oleic acid precursors to linoleic acid precursors in developing soybean seeds. Two genes encoding FAD2-1A and FAD2-1B were identified to be expressed specifically in seeds during embryogenesis and have been considered to hold an important role in controlling the seed oleic acid content. A total of 22 soybean plant introduction (PI) lines identified to have an elevated oleic acid content were characterized for sequence mutations in the FAD 2-1A and FAD2-1B genes. PI 603452 was found to contain a deletion of a nucleotide in the second exon of FAD2-1A. These important SNPs were used in developing molecular marker genotyping assays. The assays appear to be a reliable and accurate tool to identify the FAD 2-1A and FAD2-1B genotype of wild-type and mutant plants. PI 603452 was subsequently crossed with PI 283327, a soybean line that has a mutation in FAD2-1B. Interestingly, soybean lines carrying both homozygous insertion/deletion mutation (indel) FAD2-1A alleles and mutant FAD2-1B alleles have an average of 82-86% oleic acid content, compared to 20% in conventional soybean, and low levels of linoleic and linolenic acids. The newly identified indel mutation in the FAD2-1A gene offers a simple method for the development of high oleic acid commercial soybean varieties.

The soybean genome duplicated ∼14 and 45 million years ago and has many paralogous genes, including those in urease activation (emplacement of Ni and CO 2 in the active site). Activation requires the UreD and UreF proteins, each encoded by two paralogues. UreG, a third essential activation protein, is encoded by the single-copy Eu3, and eu3 mutants lack activity of both urease isozymes. eu2 has the same urease-negative phenotype, consistent with Eu2 being a single-copy gene, possibly encoding a Ni carrier. Unexpectedly, two eu2 alleles co-segregated with missense mutations in the chromosome 2 UreF paralogue (Ch02UreF), suggesting lack of expression/function of Ch14UreF. However, Ch02UreF and Ch14UreF transcripts accumulate at the same level. Further, it had been shown that expression of the Ch14UreF ORF complemented a fungal ureF mutant. A third, nonsense (Q2 * ) allelic mutant, eu2-c, exhibited 5- to 10-fold more residual urease activity than missense eu2-a or eu2-b, though eu2-c should lack all Ch02UreF protein. It is hypothesized that low-level activation by Ch14UreF is 'spoiled' by the altered missense Ch02UreF proteins ('epistatic dominant-negative'). In agreement with active 'spoiling' by eu2-b-encoded Ch02UreF (G31D), eu2-b/eu2-c heterozygotes had less than half the urease activity of eu2-c/eu2-c siblings. Ch02UreF (G31D) could spoil activation by Chr14UreF because of higher affinity for the activation complex, or because Ch02UreF (G31 D) is more abundant than Ch14UreF. Here, the latter is favoured, consistent with a reported in-frame AUG in the 5' leader of Chr14UreF transcript. Translational inhibition could represent a form of 'functional divergence' of duplicated genes.

Soybean (Glycine max (L.) Merr.) oil typically contains about 7% of linolenic acid, an oxida- tively unstable fatty acid that is undesirable for many food applications. For cooking oil, reduc- tion of the linolenic acid content by partial hydro- genation produces trans fatty acids, which are now known to increase the risk of coronary heart disease. Genetic reduction of linolenic acid was achieved decades ago, and recently the molecu- lar genetic basis for both 3% and 1% linolenic acid soybean germplasm was revealed to be combinations of mutations in three independent members of the soybean omega-3 fatty acid desaturase (FAD3) genes that corresponded to the fan1, fan2, and fan3 loci. When the mutant GmFAD3A and GmFAD3C genes are found in combination in the breeding line CX1512-44, the linolenic acid content is reduced to approxi- mately 3% of the of the seed oil. When the mutant GmFAD3A and GmFAD3B genes are in combination in the breeding line RG-10, a similar fatty acid profi le is produced with approximately 3% linolenic acid. The objective of this work was to determine the ability of different combinations of mutant FAD3 alleles from CX1512-44 and an RG-10-derived soybean line to produce less than 3% linolenic acid in the seed oil. The results indicated that novel combinations of mutations in the three FAD3 genes are capable of produc- ing soybean seeds containing only 1% linolenic acid in the seed oil.

The catabolism of cytokinins is a vital component of hormonal regulation, contributing to the control of active forms of cytokinins and their cellular distribution. The enzyme catalyzing the irreversible cleavage of N6-side chains from cytokinins is a flavoprotein classified as cytokinin dehydrogenase (CKX, EC 1.5.99.12). CKXs also show low cytokinin oxidase activity, but molecular oxygen is a comparatively poor electron acceptor. The CKX gene family of Arabidopsis thaliana comprises seven members. Four code for proteins secreted to the apoplast, the remainder are not secreted. Two are targeted to the vacuoles and one is restricted to the cytosol. This study presents the purification and characterization of each of these non-secreted CKX enzymes and substrate specificities are discussed with respect to their compartmentation. Vacuolar enzymes AtCKX1 and AtCKX3 were produced in Pichia pastoris and cytosolic enzyme AtCKX7 was expressed in Escherichia coli. The recombinant proteins were purified by column chromatography. All enzymes preferred synthetic electron acceptors over oxygen, namely potassium ferricyanide and 2,3-dimetoxy-5-methyl-1,4-benzoquinone (Q0). In slightly acidic conditions (pH 5.0), N6-(2-isopentenyl)adenine 9-glucoside (iP9G) was the best substrate for AtCKX1 and AtCKX7, whereas AtCKX3 preferentially degraded N6-(2-isopentenyl)adenine 9-riboside-5′-monophosphate (iPMP). Moreover, vacuolar AtCKX enzymes in certain conditions degraded N6-(2-isopentenyl)adenine di- and triphosphates two to five times more effectively than its monophosphate.

Raffinose family oligosaccharides (RFOs), which include raffinose and stachyose, are thought to be an important source of energy during seed germination. In contrast to their potential for promoting germination, RFOs represent anti-nutritional units for monogastric animals when consumed as a component of feed. The exact role for RFOs during soybean seed development and germination has not been experimentally determined; but it has been hypothesized that RFOs are required for successful germination. Previously, inhibition of RFO breakdown during imbibition and germination was shown to significantly delay germination in pea seeds. The objective of this study was to compare the germination potential for soybean seeds with either wild-type (WT) or low RFO levels and to examine the role of RFO breakdown in germination of soybean seeds. There was no significant difference in germination between normal and low RFO soybean seeds when imbibed/germinated in water. Similar to the situation in pea, soybean seeds of wild-type carbohydrate composition experienced a delay in germination when treated with a chemical inhibitor of alpha-galactosidase activity (1-deoxygalactonojirimycin or DGJ) during imbibition. However, low RFO soybean seed germination was not significantly delayed or reduced when treated with DGJ. In contrast to the situation in pea, the inhibitor-induced germination delay in wild-type soybean seeds was not partially overcome by the addition of galactose or sucrose. We conclude that RFOs are not an essential source of energy during soybean seed germination.

Cytokinin dehydrogenase (CKX; EC 1.5.99.12) degrades cytokinin hormones in plants. There are several differently targeted isoforms of CKX in plant cells. While most CKX enzymes appear to be localized in the apoplast or vacuoles, there is generally only one CKX per plant genome that lacks a translocation signal and presumably functions in the cytosol. The only extensively characterized maize CKX is the apoplastic ZmCKX1; a maize gene encoding a non-secreted CKX has not previously been cloned or characterized. Thus, the aim of this work was to characterize the maize non-secreted CKX gene (ZmCKX10), elucidate the subcellular localization of ZmCKX10, and compare its biochemical properties with those of ZmCKX1. Expression profiling of ZmCKX1 and ZmCKX10 was performed in maize tissues to determine their transcript abundance and organ-specific expression. For determination of the subcellular localization, the CKX genes were fused with green fluorescent protein (GFP) and overexpressed in tomato hairy roots. Using confocal microscopy, the ZmCKX1-GFP signal was confirmed to be present in the apoplast, whereas ZmCKX10-GFP was detected in the cytosol. No interactions of ZmCKX1 with the plasma membrane were observed. While roots overexpressing ZmCKX1-GFP formed significantly more mass in comparison with the control, non-secreted CKX overexpression resulted in a small reduction in root mass accumulation. Biochemical characterization of ZmCKX10 was performed using recombinant protein produced in Pichia pastoris. In contrast to the preference for 2,6-dichlorophenolindophenol (DCPIP) as an electron acceptor and trans-zeatin, N(6)-(Delta(2)-isopentenyl)adenine (iP) and N(6)-(Delta(2)-isopentenyl)adenosine (iPR) as substrates for ZmCKX1, the non-secreted ZmCKX10 had a range of suitable electron acceptors, and the enzyme had a higher preference for cis-zeatin and cytokinin N-glucosides as substrates.

The effects of high temperature treatment on soybean [Glycine max (L.) Merr.] seed composition, vigor, and proteome were investigated using mature dry seeds harvested from plants grown in environment-controlled chambers. High day/night temperatures (37/30°C) from stages R5 through R8 altered ratios of individual fatty acids to total fatty acid compared to the control (27/18°C). Concentration of sugars decreased, but total protein and phytic acid concentration were unchanged. High temperature resulted in a greater propor- tion of abnormal seeds, but normal-appearing seed exhibited reduced germination and vigor. Proteomic analysis detected 20 protein identities whose accumulations were changed by the high temperature. Fourteen spots were identified as seven subunits of seed storage proteins. The remaining six proteins were identified as those responding to abiotic stresses or having a function in respiration: (i) sucrose binding protein, (ii) Class III acidic endochitinase, (iii) heat shock protein (HSP22), (iv) late embryo abundant protein, (v) Bowman-Birk proteinase inhibitor, and (vi) formate dehydrogenase. High temperature during seed development changed soybean seed composition and decreased seed vigor, but also changed seed protein expression profiles.

Soybean [Glycine max (L.) Merr.] is important for the high protein meal used for livestock feed formulations. Carbohydrates contribute positively or negatively to the potential metabolizable energy in soybean meal. The positive carbohydrate present in soybean meal consists primarily of sucrose, whereas the negative carbohydrate components are the raffinose family of oligosaccharides (RFOs), raffinose and stachyose. Increasing sucrose and decreasing raffinose and stachyose are critical targets to improve soybean. In three recently characterized lines, variant alleles of the soybean raffinose synthase 2 (RS2) gene were associated with increased sucrose and decreased RFOs. The objective of this research was to compare the environmental stability of seed carbohydrates in soybean lines containing wild-type or variant alleles of RS2 utilizing a field location study and a date of planting study. The results define the carbohydrate variation in distinct regional and temporal environments using soybean lines with different alleles of the RS2 gene.

Characterization of genetic mutations at the molecular level allows for the development of perfect genetic markers and rapid assays to detect those markers, which enables breeders to directly select for desired alleles and accelerate the breeding process. The objectives of this study were to identify genetic lesions found in the GmFAD3A gene in the low linolenic acid soybean [Glycine max (L.) Merr] line PI 361088B, which contains a fan allele, and to develop rapid molecular marker assays that distinguish between the wild-type and PI 361088B alleles. The entire genomic sequence of the GmFAD3A gene was determined for PI 361088B. Two thymine nucleotides are inserted into a run of four thymines from nucleotide position 307 to 310 of the GmFAD3A coding sequence, resulting in a frameshift and the introduction of a premature stop codon at nucleotide 328. Two molecular marker assays were developed that rely on polymerase chain reaction (PCR) amplifi cation of the region of interest followed by restriction endonuclease digestion and agarose gel electrophoresis or melting curve analysis. A third assay is an allele-specifi c PCR-based assay that does not require any endonuclease step and requires only melting curve analysis. In conclusion, the low linolenic acid soybean line PI 361088B contains a coding mutation in GmFAD3A; this allele can easily be distinguished from the corresponding wild-type allele using any of three rapid molecular marker assays that were developed. USDA-ARS, Plant Genetics Research Unit, 110 Waters Hall, Univ. of Missouri, Columbia, MO 65211. Mention of a trademark, vendor, or proprietary product does not constitute a guarantee or warranty of the product by the USDA and does not imply its approval to the exclusion of other products or vendors that may also be suitable. Received 11 Dec. 2006. *Corresponding author (bilyeuk@missouri.edu).

Soybean is a primary source of vegetable oil, accounting for 53 % of the total vegetable oil consumption in the USA in 2013. Soybean oil with high oleic acid and low linolenic acid content is desired, because it not only improves the oxidative stability of the oil, but also reduces the amount of undesirable trans fat production by the hydrogenation processing. The mutant FAD2-1A and FAD2-1B alleles contribute to an elevated oleic acid level, while mutations in FAD3A and FAD3C lead to a reduction in linolenic acid levels of soybean seed oil. Although single nucleotide polymorphism (SNP) SimpleProbe assays have been developed for these genes, breeder-friendly and high-throughput marker systems are needed for large-scale selections in soybean breeding programs. TaqMan or Kompetitive Allele Specific PCR assays were successfully developed to detect and discriminate mutant alleles of FAD2-1A (17D and PI 603452 sources) and FAD2-1B (PI 283327 source) for the high oleic acid trait, as well as those of FAD3A (CX1512-44 and C1640 sources) and FAD3C (CX1512-44 source) for low linolenic acid content soybean. The accuracy of the assays was validated using multiple populations as well as a panel of diverse soybean lines, and a perfect association was observed between the SNP alleles and their corresponding fatty acid phenotypes. These marker assays have been proven to be robust and reliable for detecting mutant and wild-type alleles and thus will assist forward and backcrossing breeding of soybean lines with desired oil profiles in an accurate and efficient manner.

Agricultural productivity and sustainability are continually challenged by emerging and indigenous pathogens. Currently, many pathogens can be combated only with biocides or environmentally dangerous fumigants. Here, we report a rapid and pathogen-specific strategy to reduce infection by organisms that target plant roots. Combinatorially selected defense peptides, previously shown to effect premature encystment of Phytophthora capsici zoospores, were fused to maize cytokinin oxidase/dehydrogenase as a display scaffold. When expressed in tomato roots, the peptide-scaffold constructs were secreted and accumulated to sufficient concentrations in the rhizosphere to induce zoospore encystment and thereby deter taxis to the root surface. Pathogen infection was significantly inhibited in roots expressing bioactive peptides fused to the maize cytokinin oxidase/dehydrogenase scaffold. This peptide-delivery technology is broadly applicable for rapid development of plant defense attributes against plant pathogens.

Phytic acid (myo-inositol-1,2,3,4,5,6-hexakisphosphate) is the primary storage component of phosphorus in plant seeds. The first step in phytic acid biosynthesis is the de novo synthesis of myo-inositol, which is catalyzed by the enzyme D-myo-inositol 3-phosphate synthase (MIPS EC 5.5.1.4). Previous work detected four MIPS genes in soybean (Glycine max). However, only a limited amount of data were available for the MIPS gene family and some of the data were conflicting. The work described here clears up these data and characterizes the MIPS gene family for the purposes of reverse genetic technologies. The complete genomic sequence of all four genes was determined and their expression profile was examined by quantitative real-time reverse transcription-polymerase chain reaction. Our results indicate that the four MIPS genes are highly conserved and temporally and spatially expressed. The MIPS gene family in the low phytic acid soybean line, CX1834, was also characterized since this line displays a phenotype similar to previously characterized MIPS mutants. These data demonstrate that mutations in MIPS genes are not the cause of the low phytic acid phenotype.

Marker assisted selection (MAS) procedures potentially can make breeding more efficient, as genotypes can be identified before pollination allowing breeders to cross or backcross only suitable materials. Our objective was to assess the accuracy of our phenotypic selections in soybean [Glycine max (L.) Merr.] for linolenic acid using molecular markers specific for mutations in two fatty acid desaturase genes, GmFAD3A and GmFAD3C. The markers were not available earlier in the selection process and were used retrospectively to determine that phenotypic selection for seed linolenic acid # 35 g kg 21 oil was successful in capturing genotypes homozygous for mutant alleles at both loci, but phenotypic selection was not perfect. Chemical analysis for seed linolenic acid concentration was not an accurate predictor of genotype. The advancement of heterozygotes reduced the selection efficiency relative to what would have been possible using molecular markers specific for mutations in the two fatty acid desaturase genes. Errors are thought to have derived from inaccurate sample tracking or identification, contamination, or errors in chemical analyses. Use of mutation-specific molecular markers to identify F2 lines homozygous for mutant alleles in GmFAD3A and GmFAD3C, combined with diligence in reducing sampling errors, would eliminate the need for chemical testing for linolenic acid content in subsequent generations where screening can emphasize other traits. A DIFFICULT PROBLEM for the plant breeder has been to discern to what extent a trait is heritable and the result of gene action, and to what extent the trait is the result of an environmental influence. The distinction between what is a genetic vs. an environmental effect for quantitative traits is generally difficult to discern. The ideal situation for selection of a quantitative trait is