Soybean Fatty Acid Research Articles

The influence of planting density on the oil and fatty acids content of soybean in hydroponic and soil conditions of the Ararat Valley

Background: Plant seed oils are major renewable resources from nature that can be used in multiple applications. Oils and fats are a vital component of human diets. About 1/3 of the daily energy requirements of a healthy diet come from fats. Each branch of the food industry has its own demands for the fatty acid composition of seed oils. High amounts of polyunsaturated fatty acids are a requirement of healthy diets. However, food manufacturers need highly saturated fatty acids because they need oil that is resistant to high temperatures and oxidation. Soybean oil is second only behind palm oil as the most commercially abundant plant oil globally. Soybean has been recognized for its agricultural importance and their various positive effects on human health. Objective: to identify the quantitative changes of soybean fatty acids depending on the planting density and growing conditions in the Ararat Valley of Armenia. Methods: Soybean was grown in hydroponic and soil conditions of the Ararat Valley. The seeds were sown in the middle of April. Experiments were carried out in hydroponic equipment and soil with different planting densities (30, 50, 70 and 90 plants/m2). The oil content was measured by using the pulsed nuclear magnetic resonance method. The composition of fatty acids in seed oil was analyzed by chromatographic method on "Chromatec Crystal 5000" GC. Results: Oil content ranged from 19.6% to 24.4% in all variants. The maximum content was registered in soil with 30 plant/m2 planting density. Linoleic acid content ranged from 46.35 to 51.28% of the total fatty acid content, regardless of planting density and growing media. The content of linolenic acid ranged from 4.13 to 5.11% in all variants. The maximum content of ω-3 and the minimal content of ω-9 was recorded in hydroponics at 30 plants/m2 planting density. The highest percentage of stearic acid was observed in the soil variant with a planting density of 30 plants/m2. Interestingly, the variants did not differ in the total content of the 5 main fatty acids (98.84-98.95%). Growing medium had no significant effect on the total saturated fatty acids content; in all variants it ranged from 14.29% to 14.83%. Total unsaturated fatty acids content ranged from 85.2% to 85.7%. The hydroponic variant with a planting density of 30 plants/m2 had the lowest ratio of ω-6 and ω-3 fatty acids (10:1). Conclusion: Planting density and growing media had a significant influence on the content of monounsaturated and polyunsaturated fatty acids. Under hydroponic conditions the lowest content of monounsaturated and the highest content of polyunsaturated fatty acids were observed at the lowest planting density (30 plants/m2), which can be used as a functional food for daily consumption. Keywords: Glycine max (L.) Merr., soilless culture, ω-3, ω-6, ω-9, saturated acids.

NIRS-Based Prediction for Protein, Oil, and Fatty Acids in Soybean (Glycine max (L.) Merrill) Seeds

NIRS-Based Prediction for Protein, Oil, and Fatty Acids in Soybean (Glycine max (L.) Merrill) Seeds

Identification of candidate genes and genomic prediction of soybean fatty acid components in two soybean populations.

Soybean, a source of plant-derived lipids, contains an array of fatty acids essential for health. A comprehensive understanding of the fatty acid profiles in soybean is crucial for enhancing soybean cultivars and augmenting their qualitative attributes. Here, 180 F10 generation recombinant inbred lines (RILs), derived from the cross-breeding of the cultivated soybean variety 'Jidou 12' and the wild soybean 'Y9,' were used as primary experimental subjects. Using inclusive composite interval mapping (ICIM), this study undertook a quantitative trait locus (QTL) analysis on five distinct fatty acid components in the RIL population from 2019 to 2021. Concurrently, a genome-wide association study (GWAS) was conducted on 290 samples from a genetically diverse natural population to scrutinize the five fatty acid components during the same timeframe, thereby aiming to identify loci closely associated with fatty acid profiles. In addition, haplotype analysis and the Kyoto Encyclopedia of Genes and Genomes pathway analysis were performed to predict candidate genes. The QTL analysis elucidated 23 stable QTLs intricately associated with the five fatty acid components, exhibiting phenotypic contribution rates ranging from 2.78% to 25.37%. In addition, GWAS of the natural population unveiled 102 significant loci associated with these fatty acid components. The haplotype analysis of the colocalized loci revealed that Glyma.06G221400 on chromosome 6 exhibited a significant correlation with stearic acid content, with Hap1 showing a markedly elevated stearic acid level compared with Hap2 and Hap3. Similarly, Glyma.12G075100 on chromosome 12 was significantly associated with the contents of oleic, linoleic, and linolenic acids, suggesting its involvement in fatty acid biosynthesis. In the natural population, candidate genes associated with the contents of palmitic and linolenic acids were predominantly from the fatty acid metabolic pathway, indicating their potential role as pivotal genes in the critical steps of fatty acid metabolism. Furthermore, genomic selection (GS) for fatty acid components was conducted using ridge regression best linear unbiased prediction based on both random single nucleotide polymorphisms (SNPs) and SNPs significantly associated with fatty acid components identified by GWAS. GS accuracy was contingent upon the SNP set used. Notably, GS efficiency was enhanced when using SNPs derived from QTL mapping analysis and GWAS compared with random SNPs, and reached a plateau when the number of SNP markers exceeded 3,000. This study thus indicates that Glyma.06G221400 and Glyma.12G075100 are genes integral to the synthesis and regulatory mechanisms of fatty acids. It provides insights into the complex biosynthesis and regulation of fatty acids, with significant implications for the directed improvement of soybean oil quality and the selection of superior soybean varieties. The SNP markers delineated in this study can be instrumental in establishing an efficacious pipeline for marker-assisted selection and GS aimed at improving soybean fatty acid components.

The catalytic deoxygenation reaction temperature and N2 gas flow rate influence the conversion of soybean fatty acids into Green Diesel

BackgroundGreen diesel is a promising alternative as a petroleum replacement given the worldwide demand for petroleum fuel. Environmental issues have drawn public attention and concerns towards advancing renewable energy development. A catalytic deoxygenation (deCOx) was carried out to produce green diesel from soybean oil (SO) using a low-cost NiO-doped calcined dolomite (NiOCD) catalyst. MethodThe structure, chemical composition and morphology of NiOCD were comprehensively characterized by XRF, BET, TPD-CO2, SEM and TEM. In this study, the effect of two operating parameters, reaction temperature and flow rate of nitrogen, was discovered using a one-factor-at-a-time (OFAT) optimisation study. In addition, the life cycle cost analysis (LCCA) of stepwise catalyst preparation and green diesel production has been performed. Significant findingsAn optimal reaction temperature of 420 °C was found to provide the highest yield of green diesel (47.13 wt.%) with an 83.51% hydrocarbon composition. The ideal nitrogen flow rate, however, was found to be 50 cm3/min, which produced 41.80 wt.% of green diesel with an 88.63% hydrocarbon composition. The deoxygenation reaction was significantly impacted by both reaction temperature and nitrogen flow rate. According to LCCA, NiOCD catalyst has potential to lower the overall cost of producing green diesel compared to commercial zeolite catalysts.

Effects of High Hydrostatic Pressure on the Distribution of Oligosaccharides, Pinitol, Soysapapogenol A, and Fatty Acids in Soybean.

The effects of high hydrostatic pressure (HHP) treatment (100-600 MPa for 10-60 min) and thermal treatment (boiling for 10-60 min) on oligosaccharides, pinitol, and soyasapogenol A as taste ingredients in soybean (Glycine max (L.) Merr.) (cv. Yukihomare) were evaluated. Additionally, soybean-derived fatty acids such as α-linolenic acid, linoleic acid, oleic acid, palmitic acid, and stearic acid in pressurized soybeans were quantitatively analyzed. Sucrose, stachyose, and raffinose concentrations were decreased in all tested pressure and time combinations; however, pinitol concentrations were increased by specific pressure and time combinations at 100-400 MPa for 10-60 min. While the soyasapogenol A content in boiled soybeans decreased with increasing boiling time, that of pressurized soybeans was altered by specific pressure and time combinations. At the lower pressure and shorter time combinations, the essential fatty acids such as α-linolenic acid and linoleic acid showed higher contents. Stearic acid and oleic acid contents of pressurized soybeans increased at mild pressure levels (300-500 MPa). In contrast, the combination of higher pressure and longer time results in lower essential fatty acid contents. Non-thermal-pressurized soybeans have the potential to be a high-value food source with better taste due to the enrichment of low molecular weight components such as pinitol, free amino acids, and the reduction of isoflavones and Group A soyasapogenol.

Multi-omics analysis reveals novel loci and a candidate regulatory gene of unsaturated fatty acids in soybean (Glycine max (L.) Merr)

BackgroundSoybean is a major oil crop; the nutritional components of soybean oil are mainly controlled by unsaturated fatty acids (FA). Unsaturated FAs mainly include oleic acid (OA, 18:1), linoleic acid (LLA, 18:2), and linolenic acid (LNA, 18:3). The genetic architecture of unsaturated FAs in soybean seeds has not been fully elucidated, although many independent studies have been conducted. A 3 V multi-locus random single nucleotide polymorphism (SNP)-effect mixed linear model (3VmrMLM) was established to identify quantitative trait loci (QTLs) and QTL-by-environment interactions (QEIs) for complex traits.ResultsIn this study, 194 soybean accessions with 36,981 SNPs were calculated using the 3VmrMLM model. As a result, 94 quantitative trait nucleotides (QTNs) and 19 QEIs were detected using single-environment (QTN) and multi-environment (QEI) methods. Three significant QEIs, namely rs4633292, rs39216169, and rs14264702, overlapped with a significant single-environment QTN.ConclusionsFor QTNs and QEIs, further haplotype analysis of candidate genes revealed that the Glyma.03G040400 and Glyma.17G236700 genes were beneficial haplotypes that may be associated with unsaturated FAs. This result provides ideas for the identification of soybean lipid-related genes and provides insights for breeding high oil soybean varieties in the future.

Mapping and identification of QTLs for seed fatty acids in soybean (Glycine max L.)

Soybean is one of the world's most important sources of vegetable oil. The oil content and fatty acid ratio have attracted significant attention due to their impact on the shelf-life of soybean oil products and the consumers' health. In this study, the high-density genetic map derived from Guizao 1 and Brazil 13 was utilised to analyse the quantitative trait loci of palmitic acid (PA), stearic acid (SA), oleic acid (OA), linoleic acid (LA), linolenic acid (LNA) and oil content. A total of 54 stable QTLs were detected in genetic map linkage analysis, which shared six bin intervals. Among them, the bin interval on chromosome 13 (bin106-bin118 and bin123-bin125) was detected as stable QTLs in multiple environments, linked to OA, LA, and LNA. Eight differentially expressed genes (DEGs) within these QTLs interval were determined as candidate genes according to the joint investigation of parental resequencing, bioinformatics and RNA sequencing data. All these results will be conducive to breeding soybean with the ideal fatty acid ratio for food, and provide the genetic information basis for mining genes related to fatty acid and oil content traits in soybean.

Enhancing the recovery of human-edible nutrients in milk and nitrogen efficiency throughout the lactation cycle by feeding fatty acid supplements

Dairy cows convert low-quality feedstuffs unsuitable for human consumption into milk, a high-quality protein source for humans. Even under grazing conditions of well-managed pastures, dietary energy limits the efficiency of animal production, contributing to the excretion of nutrients to the environment. Thus, our study aimed to assess the effect of supplying sources and levels of supplemental fat to grazing dairy cows during early lactation on nutrient excretion and recovery of human-edible (HE) nutrients in milk throughout the lactation cycle. Two experiments were conducted with grazing Jersey × Holstein dairy cows. In Experiment 1, five dairy cows were used in a 5 × 5 Latin Square design. Treatments were a control diet (concentrate without supplemental fat) or diets supplemented with calcium salts of either soybean fatty acids (CSSO) or palm fatty acids (CSPO) fed at two levels of supplementation (400 g/day or 700 g/day) for 24 days. In general, the level of supplemental fat had a minor effect on production parameters and methane emission and intensity. Feeding cows with CSPO increased milk yield by 8.1%, milk energy output by 20%, and milk fat yield by 35.2% compared with CSSO. A tendency was observed for CSPO to decrease methane per unit of milk energy compared with CSSO. In experiment 2, 24 dairy cows were used in a randomized block design. Treatments were a control diet or diets supplemented with calcium salts of either CSSO or CSPO fed at 400 g/day for 13 weeks. The carryover effect of the diets was evaluated until 42 weeks of lactation. During the supplementation period, feeding CSPO improved the recovery of HE energy, protein, and indispensable amino acids compared with control and CSSO. Interestingly, we demonstrated that feeding CSPO improved the recovery of HE energy, protein, and indispensable amino acids compared with control over the lactation cycle. Overall, our results suggested that the dose of supplemental fat used in our study had a minor influence on production and rumen fermentation parameters. We demonstrated that feeding dairy cows with a fatty acid source based on palm oil under tropical grazing conditions reduced nutrient excretion and improved the recovery of HE nutrients throughout the lactation cycle.

Genome-wide identification and expression analysis of diacylglycerol acyltransferase genes in soybean (Glycine max).

Soybean (Glycine max) is a major protein and vegetable oil source. In plants, diacylglycerol acyltransferase (DGAT) can exert strong flux control, which is rate-limiting for triacylglycerol biosynthesis in seed oil formation. Here, we identified soybean DGAT genes via a bioinformatics method, thereby laying a solid foundation for further research on their function. Based on our bioinformatics analyses, including gene structure, protein domain characteristics, and phylogenetic analysis, 26 DGAT putative gene family members unevenly distributed on 12 of the 20 soybean chromosomes were identified and divided into the following four groups: DGAT1, DGAT2, WS/DGAT, and cytoplasmic DGAT. The Ka/Ks ratio of most of these genes indicated a significant positive selection pressure. DGAT genes exhibited characteristic expression patterns in soybean tissues. The differences in the structure and expression of soybean DGAT genes revealed the diversity of their functions and the complexity of soybean fatty acid metabolism. Our findings provide important information for research on the fatty acid metabolism pathway in soybean. Furthermore, our results will help identify candidate genes for potential fatty acid-profile modifications to improve soybean seed oil content. This is the first time that in silico studies have been used to report the genomic and proteomic characteristics of DGAT in soybean and the effect of its specific expression on organs, age, and stages.

Recovery of Copper from Low Grade Ores by HNO3 Leaching: Process Optimization and Solvent Extraction Studies using Soybean Fatty Acid

This paper presents the recovery of copper from a Nigerian low-grade copper ore containing 4.08 wt. % Cu via nitric acid leaching, followed by solvent extraction using soybeans oil fatty acid extractant. The leaching tests on the milled ore were done according to a 3-factor Central Composite Design with time, acid concentration and temperature as the selected process variables. Regression models were developed from the experimental data for the ore dissolution and the quantity of copper extracted, and the effects of the individual process variables and their interactions were determined from the leaching studies using analysis of variance (ANOVA). Results from the leaching tests indicate that the ore dissolution depends mainly on the leaching time. About 92 percent of the copper contents were recovered at optimum leaching time, temperature and nitric acid concentration of 29.5 min, 81.25 oC and 2.18 M, respectively. The solvent extraction studies show that copper can be extracted from aqueous media using soybeans fatty acid, with the performance of the diluents in terms of the quantity of Cu extracted being MIBK > kerosene > xylene.

Effects of Different Gene Editing Modes of CRISPR/Cas9 on Soybean Fatty Acid Anabolic Metabolism Based on GmFAD2 Family

Δ12-fatty acid dehydrogenase (FAD2) is the essential enzyme responsible for catalyzing the formation of linoleic acid from oleic acid. CRISPR/Cas9 gene editing technology has been an essential tool for molecular breeding in soybeans. To evaluate the most suitable type of gene editing in soybean fatty acid synthesis metabolism, this study selected five crucial enzyme genes of the soybean FAD2 gene family-GmFAD2-1A, GmFAD2-1B, GmFAD2-2A, GmFAD2-2B, and GmFAD2-2C-and created a CRISPR/Cas9-mediated single gene editing vector system. The results of Sanger sequencing showed that 72 transformed plants positive for T1 generation were obtained using Agrobacterium-mediated transformation, of which 43 were correctly edited plants, with the highest editing efficiency of 88% for GmFAD2-2A. The phenotypic analysis revealed that the oleic acid content of the progeny of GmFAD2-1A gene-edited plants had a higher increase of 91.49% when compared to the control JN18, and the rest of the gene-edited plants in order were GmFAD2-2A, GmFAD2-1B, GmFAD2-2C, and GmFAD2-2B. The analysis of gene editing type has indicated that base deletions greater than 2bp were the predominant editing type in all editing events. This study provides ideas for the optimization of CRISPR/Cas9 gene editing technology and the development of new tools for precise base editing in the future.

Bioinformatics and Functional Analysis of High Oleic Acid-Related Gene GmSAM22 in Soybean [Glycine max (L.) Merr.

High yield, high quality, stable yield, adaptability to growth period, and modern mechanization are the basic requirements for crops in the 21st century. Soybean oleic acid is a natural unsaturated fatty acid with strong antioxidant properties and stability. Known as a safe fatty acid, it has the ability to successfully prevent cardiovascular and cerebrovascular disorders. Improving the fatty acid composition of soybean seeds, can not only speed up the breeding process of high-quality high-oil and high-oleic soybeans, but also have important significance in human health, and provide the possibility for the development of soybean oil as a new energy source. Hence, the aim of this study was to analyze the high oleic acid elated gene GmSAM22 in soybean. In this research the soybean oleic acid-related gene GmSAM22 was screened out by Genome-wide association analysis, a 662 bp fragment was acquired by specific PCR amplification, and the pMD18T cloning vector was linked by the use of a seamless cloning technique. Bioinformatics analysis of the signal peptide prediction, subcellular localization, protein hydrophobicity, transmembrane region analysis, a phosphorylation site, protein secondary and tertiary structure and protein interaction analysis of the protein encoded by the SAM22 gene was carried out. The plasmid of the gene editing vector is pBK041. The overexpression vector was transformed from pCAMBIA3301 as the base vector, and overexpression vector were designed. Positive plants were obtained by genetic transformation by the pollen tube channel method. Fluorescence quantitative PCR was performed on the T2 generation plants to detect the relative expression levels in different tissues. Southern Blot was used to detect the presence of hybridization signal. Screening genes BAR, 35S, and NOS in plants were identified by conventional PCR. 10 seeds with high and low oleic acid content were chosen for quantitative PCR identification, and finally, the concentration and morphology of soybean fatty acids were identified by near-far infrared spectroscopy. On 10 seeds with an upper and lower oleic acid content, a quantitative fluorescence analysis was done. In Southern blot hybridization, the SAM22 gene was integrated into the recipient soybean plant in hands of a sole copy. Fluorescence quantitative PCR appeared that the average relative expression of the SAM22 gene in roots, stems, leaves, and seeds was 1.70, 1.67, 3.83, and 4.41, respectively. Positive expression seeds had a 4.77% increase in oleic acid content. The level of oleic acid in the altered seeds was reduced by 4.13% when compared to CK, and it was discovered that the GmSAM22 gene could be a regulatory and secondary gene that promotes the conversion of stearic acid to oleic acid in soybean. There has not been a discussion of gene cloning or functional verification. The cloning and genetic transformation of the soybean SAM22 gene can effectively increase the content of oleic acid, which lays a foundation for the study of soybean with high oleic acid.

CRISPR/Cas9-Mediated Mutagenesis of GmFAD2-1A and/or GmFAD2-1B to Create High-Oleic-Acid Soybean

Soybean (Glycine max (L.) Merr.) oil is an important source of vegetable oil for supporting the human diet. However, the high level of polyunsaturated fatty acids in natural soybean oil renders the oil unstable and thus susceptible to the development of unpalatable flavors and trans fatty acids. Therefore, reducing the content of polyunsaturated fatty acids and increasing the content of monounsaturated fatty acids is a longstanding and important target for soybean breeding. However, soybean varieties with a high oleic acid content are rare in soybean germplasm resources, which introduces substantial difficulties in the cultivation of high-oleic-acid soybeans. In this study, CRISPR/Cas9-mediated gene-editing technology was used to create targeted knockout of the soybean fatty acid desaturase encoding genes GmFAD2-1A and GmFAD2-1B that contribute to the formation of polyunsaturated fatty acids. We obtained fad2-1a, fad2-1b, and fad2-1a/fad2-1b homozygous mutants using two sgRNAs. We found that the oleic acid content increased from 11% to 40-50% in the fad2-1a and fad2-1b mutants and to 85% in the fad2-1a/fad2-1b mutants. We also generated transgene-free double mutants that conferred higher oleic acid, and the fad2-1a/fad2-1b mutant had no adverse phenotyping compared with the wild type. Our study provided new materials for the selection and breeding of high-oleic-acid soybean varieties.

Fatty acids methyl esters from soybean oil for biobased surfactants industry: Obtention C16 / C18 concentrate for use as feedstock

Soybean fatty acid methyl esters (FAMEs) are used as biodiesel, a co-fuel according to current consumption figures. In addition, FAMEs are becoming increasingly important as biodegradable substitutes for mineral oils, such as oil for chainsaws, gearboxes, hydraulics, and lubricants for crude oil production. Therefore, the possibility of producing a wide spectrum of value-added products in oleochemical industry will guarantee the competitiveness of biodiesel plants in the market. In this way, soybean FAMEs of local production were modified to obtain the desired characteristics to produce biobased surfactants as methyl ester sulfonated (MES). In order to incorporate soybean FAMEs into the conventional MES manufacturing process, their lipid profile must be similar to that of the traditionally used raw material, that is, palm oil. This oil is composed of acids with a length between C12 and C18 and 44–50 % of unsaturated acids, a higher concentration of unsaturated acids is not convenient because generated impurities could affect the final color of the product. Thus, the objective of this work is to achieve a ratio between soybean C16/C18 FAMEs of 60/40, the maximum possible ratio from the initial C16/C18 of 11.0/87.6. In this study, molecular distillation and urea complexation were used as individual and combined techniques to obtain C16/C18 concentrate. In the molecular distillation a ratio of 16.2 / 83.1 was obtained with a yield of 23.2 % (evaporation temperature of 70 °C) while by urea complexation a ratio of 35.4/ 60.8 was obtained with a yield of 32.0 % (urea: FAMEs mass ratio of 1: 1). The most suitable combination of both techniques was a urea complexation stage followed by a molecular distillation stage. The obtained product had a ratio of 56.7 / 43.3 and a yield of 41.8 %. The results of this study could be applied to other vegetable sources to obtain an appropriate profile of FAMEs. Also, the developed process in this work could be used in the industry of green surfactants to produce the segmentation of biodiesel industry. • FAMEs from soybean oil could be used in the biobased surfactants industry. • Urea Complexation and Molecular Distillation combined could join to biodiesel industry. • This method of purification allows a greater productive diversification of biodiesel. • The combination of two techniques allows modifying the FAMEs profile of soybean oil.

QTL Mapping of Palmitic Acid Content Using Specific-Locus Amplified Fragment Sequencing (SLAF-Seq) Genotyping in Soybeans (Glycine max L.).

Soybeans are essential crops that supply protein and oil. The composition and contents of soybean fatty acids are relevant to human health and have a significant relationship with soybean oil processing and applications. Identifying quantitative trait locus (QTL) genes related to palmitic acid could facilitate the development of a range of nutritive soybean cultivars using molecular marker-assisted selection. In this study, we used a cultivar with higher palmitic acid content, ‘Dongnong42’, and a lower palmitic acid content cultivar, ‘Hobbit’, to establish F2:6 recombinant inbred lines. A high-density genetic map containing 9980 SLAF markers was constructed and distributed across 20 soybean chromosomes. The genetic map contained a total genetic distance of 2602.58 cM and an average genetic distance of 0.39 cM between adjacent markers. Two QTLs related to palmitic acid content were mapped using inclusive composite interval mapping, explaining 4.2–10.1% of the phenotypic variance in three different years and environments, including the QTL included in seed palmitic 7-3, which was validated by developing SSR markers. Based on the SNP/Indel and significant differential expression analyses of Dongnong42 and Hobbit, two genes, Glyma.15g119700 and Glyma.15g119800, were selected as candidate genes. The high-density genetic map, QTLs, and molecular markers will be helpful for the map-based cloning of palmitic acid content genes. These could be used to accelerate breeding for high nutritive value cultivars via molecular marker-assisted breeding.

PSVI-15 Effects of Different Dietary Sources of Lipids for Lactating Ewes

Abstract The aim of this study was to evaluate the effects of diets with fatty acid profiles on lactation performance and milk composition of ewes. Dorper vs. Santa Inês ewes (n=40; 54.9 ± 8.3 kg BW and 14 days in milk) were penned individually and used in a randomized complete block design. Ewes were fed a basal diet (19.87 ± 0.62% CP) containing 70% concentrate and 30% coastcross hay, and no added oil (CON). The remaining diets were added with 4.0% of soybean oil (SO), calcium salts of palm fatty acids (CSPF), or calcium salts of soybean fatty acids (CSSF) (DM basis). Ewes were fed the diets from the week 2 to 10 of lactation. Milk production was determined every 7 days during the experiment. Ewes were separated from their lambs and mechanically milked for emptying of the mammary gland. After 3 hours, using the same procedure, milk production was recorded and samples collected for milk composition analysis. The addition of SO decreased (P< 0.001) DM intake, with no difference between the other treatments. Milk production, as well as the production of fat, protein, lactose and total solids (g/3 hours) were not affected by the treatments. Regarding milk composition, the percentage of milk fat and total solids increased (P< 0.0001) with CSSF. The percentage of protein was less (P< 0.001) in the milk from ewes that received the diet containing CSPF, with no difference between the other diets. Finally, the percentage of lactose was less (P< 0.001) for ewes fed the diet containing calcium salts of palm fatty acids. In conclusion, soybean oil had the greatest negative effect on dry matter intake. All the oil sources modulated the milk composition, with better results in this regard for the calcium salts of palm fatty acids, considering that it increased the fat and total solids content of the milk.

Construction and Functional Analysis of CRISPR/Cas9 Vector of FAD2 Gene Family in Soybean

Soybean oleic acid content is one of the important indexes to evaluate the quality of soybean oil. In the synthesis pathway of soybean fatty acids, the FAD2 gene family is the key gene that regulates the production of linoleic acid from soybean oleic acid. In this study, CRISPR/Cas9 gene editing technology was used to regulate FAD2 gene expression. Firstly, the CRISPR/Cas9 single knockout vectors GmFAD2-1B and GmFAD2-2C and double knockout vectors GmFAD2-2A-3 were constructed. Then, the three vectors were transferred into the recipient soybean variety Jinong 38 by Agrobacterium-mediated cotyledon node transformation, and the mutant plants were obtained. Functional analysis and comparison of the mutant plants of the T2 and T3 generations were carried out. The results showed that there was no significant difference in agronomic traits between the CRISPR/Cas9 single and double knockout vectors and the untransformed CRISPR/Cas9 receptor varieties. The oleic acid content of the plants that knocked out the CRISPR/Cas9 double gene vector was significantly higher than that of the single gene vector.

Application of Rhizomucor miehei lipase-displaying Pichia pastoris whole cell for biodiesel production using agro-industrial residuals as substrate

This work aimed the application of a new biocatalyst for biodiesel production from residual agro-industrial fatty acids. A recombinant Pichia pastoris displaying lipase from Rhizomucor miehei (RML) on the cell surface, using the PIR-1 anchor system, were prepared using glycerol as the carbon source. The biocatalyst, named RML-PIR1 showed optimum temperature of 45 °C (74.0 U/L). The stability tests resulted in t1/2 of 3.49 and 2.15 h at 40 and 45 °C, respectively. RML-PIR1 was applied in esterification reactions using industrial co-products as substrates, palm fatty acid distillate (PFAD) and soybean fatty acid distillate (SFAD). The highest productivity was observed for SFAD after 48 h presenting 79.1% of conversion using only 10% of biocatalyst and free-solvent system. This is about ca. eight times higher than commercial free RML in the same conditions. The stabilizing agents study revealed that the treatment using glutaraldehyde (GA) and poly(ethylene glycol) (PEG) enabled increased stability and reuse of biocatalyst. It was observed by SEM analysis that the treatment modified the cell morphology. RML-PIR1-GA presented 87.9% of the initial activity after 6 reuses, whilst the activity of unmodified RML-PIR decreased by 40% after the first use. These results were superior to those obtained in the literature, making this new biocatalyst promising for biotechnological applications, such as the production of biofuels on a large scale.

Genome-wide identification and analysis of soybean acyl-ACP thioesterase gene family reveals the role of GmFAT to improve fatty acid composition in soybean seed.

Soybean acyl-ACP thioesterase gene family have been characterized; GmFATA1A mutants were discovered to confer high oleic acid, while GmFATB mutants presented low palmitic and high oleic acid seed content. Soybean oil stability and quality are primarily determined by the relative proportions of saturated versus unsaturated fatty acids. Commodity soybean typically contains 11% palmitic acid, as the primary saturated fatty acids. Reducing palmitic acid content is the principal approach to minimize the levels of saturated fatty acids in soybean. Though high palmitic acid enhances oxidative stability of soybean oil, it is negatively correlated with oil and oleic acid content and can cause coronary heart diseases for humans. For plants, acyl-acyl carrier protein (ACP) thioesterases (TEs) are a group of enzymes to hydrolyze acyl group and release free fatty acid from plastid. Among them, GmFATB1A has become the main target to genetically reduce the palmitic acid content in soybean. However, the role of members in soybean acyl-ACP thioesterase gene family is largely unknown. In this study, we characterized two classes of TEs, GmFATA, and GmFATB in soybean. We also denominated two GmFATA members and discovered six additional members that belong to GmFATB gene family through phylogenetic, syntenic, and in silico analysis. Using TILLING-by-Sequencing+, we identified an allelic series of mutations in five soybean acyl-ACP thioesterase genes, including GmFATA1A, GmFATB1A, GmFATB1B, GmFATB2A, and GmFATB2B. Additionally, we discovered mutations at GmFATA1A to confer high oleic acid (up to 34.5%) content, while mutations at GmFATB presented low palmitic acid (as low as 5.6%) and high oleic acid (up to 36.5%) phenotypes. The obtained soybean mutants with altered fatty acid content can be used in soybean breeding program for improving soybean oil composition traits.

Circular RNAs acting as ceRNAs mediated by miRNAs may be involved in the synthesis of soybean fatty acids.

Soybean oil is composed of fatty acids and glycerol. The content and composition of fatty acids partly determine the quality of soybean seeds. Circular RNAs (circRNAs) are endogenous non-coding RNAs that competitively bind to microRNAs (miRNAs) through miRNA recognition elements, thereby acting as sponges to regulate the expression of target genes. Although circRNAs have been identified previously in soybean, only their expression has been investigated without exploration of the competitive endogenous RNAs (ceRNAs) network of circRNAs-miRNAs-mRNAs. In this study, circRNAs in immature pods of a low linolenic acid soybean Mutant 72' (MT72) and the wild-type control 'Jinong 18' (JN18) were systematically identified and analyzed at 30 and 40days after flowering using high-throughput sequencing technology. We identified 6377 circRNAs, of which 114 were differentially expressed. Gene ontology and KEGG pathway analyses of targeted mRNAs in the ceRNAs network indicated that the differentially expressed circRNAs may be involved in fatty acid transport, suggesting that circRNAs may play a post-transcriptional regulatory role in soybean oil synthesis. This study provides a foundation for future exploration of the function of circRNAs in soybean and presents novel insights to guide further studies of plant circRNAs.