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The expanding cultivation of grains to meet agro-industry demands, the implementation of more efficient, sustainable, and integrative management practices capable of mitigating the selection of resistant pests and weeds becomes necessary. Due to the intensive application of chemical herbicides, in addition to environmental impacts, led to the development of resistance mechanisms in weed populations, such as sourgrass ( Digitaria insularis ). These mechanisms complicate management efforts, escalate production costs, and diminish productivity. In this context, RNA interference (RNAi) technology has emerged as a promising molecular tool for the targeted control of weeds, owing to its specificity in the post-transcriptional silencing of vital genes. This study investigated the application of RNAi technology for the suppression of 5-enolpyruvylshikimate-3-phosphate synthase ( EPSPS ) gene expression in D. insularis , a weed species of significant agronomic importance in Latin America. A double-stranded RNA (dsRNA) sequence, specifically designed for the EPSPS gene, was synthesized via bacterial fermentation with a strain of E. coli HT115, extracted using TRIzol™ and purified with phenol: chloroform: isoamyl alcohol, and applied topically in D. insularis leaves. The subsequent phenotypic and molecular effects were evaluated. The spray application of the dsRNA resulted in a 44% reduction in the plant's shoot dry mass and a 75% reduction in the number of tillers, thereby indicating consistent physiological impacts due to gene silencing. Quantitative reverse transcription PCR (qRT-PCR) analysis confirmed a significant suppression of EPSPS transcript levels following treatment, suggesting partial gene silencing. These findings collectively demonstrate the efficacy of RNAi in modulating gene expression in D. insularis , thereby underscoring its potential as a sustainable biotechnological strategy for the development of novel weed control methodologies.

Drought is one of the most stressful environmental factor causing yield and economic losses in many soybean-producing regions. In the last decades, transcription factors (TFs) are being used to develop genetically modified plants more tolerant to abiotic stresses. Dehydration responsive element binding (DREB) and ABA-responsive element-binding (AREB) TFs were introduced in soybean showing improved drought tolerance, under controlled conditions. However, these results may not be representative of the way in which plants behave over the entire season in the real field situation. Thus, the objectives of this study were to analyze agronomical traits and physiological parameters of (1Ab58), (1Bb2193), and (1Ea2939) GM lines under irrigated (IRR) and non-irrigated (NIRR) conditions in a field experiment, over two crop seasons and quantify transgene and drought-responsive genes expression. Results from season 2013/2014 revealed that line 1Ea2939 showed higher intrinsic water use and leaf area index. Lines 1Ab58 and 1Bb2193 showed a similar behavior to wild-type plants in relation to chlorophyll content. Oil and protein contents were not affected in transgenic lines in NIRR conditions. Lodging, due to plentiful rain, impaired yield from the 1Ea2939 line in IRR conditions. qPCR results confirmed the expression of the inserted TFs and drought-responsive endogenous genes. No differences were identified in the field experiment performed in crop season 2014/2015, probably due to the optimum rainfall volume during the cycle. These field screenings showed promising results for drought tolerance. However, additional studies are needed in further crop seasons and other sites to better characterize how these plants may outperform the WT under field water deficit.

Soybean is one of the most common grain crops worldwide, representing an important protein and oil source. Although genetic variability in the chemical composition of grains is seen in soybean, the mean levels of proteins have remained stagnant or, in some cases, have decreased over time, arousing concern in the agricultural industry. Furthermore, environmental conditions influence the chemical composition of grains. Thus, the present study evaluated the effect of water deficit (WD) induced at the vegetative period (vegetative stress (VS)) and reproductive period (reproductive stress (RS)) on the protein and oil contents of grains in different soybean genotypes. Yield and its components were evaluated to evaluate the interrelation of these traits. The experiment was completed over three crop seasons under field conditions in Londrina, Paraná (PR), Brazil. WD was induced using rainout shelters and then stress treatments with irrigated and non-irrigated conditions were compared. WD negatively affected yield and its components. All evaluated genotypes showed similar responses for oil and protein contents under different water conditions. Higher protein content and lower oil content were observed in grains under RS. Such a relationship was not equally established under VS. Additionally, negative relationships between protein and oil content and between protein content and yield were confirmed.

The yield and protein content of soybean seeds are challenging traits for breeding since they have a negative correlation. To solve this issue, it is essential to understand the protein transport in soybean seeds. Here we performed an analysis of 3 datasets containing transcriptional data from soybean seeds and pods, without any treatment, aiming to select genes related to the transport of organic nitrogen compounds. We identified 69 transporters, and among them, seven were chosen to be validated by qRT-PCR. Seeds and pods of soybean cultivars BRS 232 and BRS 284, respectively, presenting higher and lower protein content, were assayed under normal growth condition. Results showed in BRS 232 soybean cultivar, a positive correlation between seed protein content and gene expression for five out of the seven genes analyzed. These nitrogen compounds transporters can play an essential role in the storage of proteins, thus increases the protein content of soybean seeds and contributes to the decrease the negative correlation between yield and protein content of soybean seeds.

Soybean is the primary source of vegetable protein and is used for various purposes, mainly to feed animals. This crop can have diverse seed coat colors, varying from yellow, black, brown, and green to bicolor. Black seed coat cultivars have already been assigned as favorable for both seed and grain production. Thus, this work aimed to identify genes associated with soybean seed quality by comparing the transcriptomes of soybean seeds with contrasting seed coat colors. The results from RNA-seq analyses were validated with real-time PCR using the cultivar BRS 715A (black seed coat) and the cultivars BRS 413 RR and DM 6563 IPRO (yellow seed coat). We found 318 genes differentially expressed in all cultivars (freshly harvested seeds and seeds stored in cold chamber). From the in silico analysis of the transcriptomes, the following genes were selected and validated with RT-qPCR: ACS1, ACSF3, CYP90A1, CYP710A1, HCT, CBL, and SAHH. These genes are genes induced in the black seed coat cultivar and are part of pathways responsible for ethylene, lipid, brassinosteroid, lignin, and sulfur amino acid biosynthesis. The BRSMG 715A gene has almost 4times more lignin than the yellow seed coat cultivars. These attributes are related to the BRSMG 715A cultivar’s higher seed quality, which translates to more longevity and resistance to moisture and mechanical damage. Future silencing studies may evaluate the knockout of these genes to better understand the biology of soybean seeds with black seed coat.

The present study aimed to identify the relationship between lignin content in soybean pods, pod dehiscence rate, gene expression from phenylpropanoid pathway, cell wall lignification, and deterioration of seed due to weather. To evaluate the weathering deterioration, an experiment simulating 150 mm of rainfall was performed in greenhouse conditions. Gene expression was assessed by RT-qPCR. Results showed that soybean cultivars BRS Jiripoca and BRS 388 RR considered tolerant to weathering deterioration, presented higher levels of lignin content in pods. The data showed that dehiscence rate was neither a trait involved in weathering deterioration tolerance nor in lignin content in soybean pods. Lignin content and weathering deterioration were highly negatively correlated, probably due to changes in cell wall permeability to water accordingly to lignin level. Higher expression levels of genes involved in lignin biosynthesis and cell wall lignification were identified in both tolerant cultivars, however, with opposite expression profiles and in different developmental phases, suggesting a possible temporal strategy to cope with environmental adverse conditions.

Soybean (Glycine max) is one of the major crops worldwide and flooding stress affects the production and expansion of cultivated areas. Oxygen is essential for mitochondrial aerobic respiration to supply the energy demand of plant cells. Because oxygen diffusion in water is 10,000 times lower than in air, partial (hypoxic) or total (anoxic) oxygen deficiency is important component of flooding. Even when oxygen is externally available, oxygen deficiency frequently occurs in bulky, dense or metabolically active tissues such as phloem, meristems, seeds, and fruits. In this study, we analyzed conserved and divergent root transcriptional responses between flood-tolerant Embrapa 45 and flood-sensitive BR 4 soybean cultivars under hypoxic stress conditions with RNA-seq. To understand how soybean genes evolve and respond to hypoxia, stable and differentially expressed genes were characterized structurally and compositionally comparing its mechanistic relationship. Between cultivars, Embrapa 45 showed less up- and more down-regulated genes, and stronger induction of phosphoglucomutase (Glyma05g34790), unknown protein related to N-terminal protein myristoylation (Glyma06g03430), protein suppressor of phyA-105 (Glyma06g37080), and fibrillin (Glyma10g32620). RNA-seq and qRT-PCR analysis of non-symbiotic hemoglobin (Glyma11g12980) indicated divergence in gene structure between cultivars. Transcriptional changes for genes in amino acids and derivative metabolic process suggest involvement of amino acids metabolism in tRNA modifications, translation accuracy/efficiency, and endoplasmic reticulum stress in both cultivars under hypoxia. Gene groups differed in promoter TATA box, ABREs (ABA-responsive elements), and CRT/DREs (C-repeat/dehydration-responsive elements) frequency. Gene groups also differed in structure, composition, and codon usage, indicating biological significances. Additional data suggests that cis-acting ABRE elements can mediate gene expression independent of ABA in soybean roots under hypoxia.

Water deficit may occur at any stage of crop development, affecting productivity and causing economic losses. In response to drought, raffinose family oligosaccharides (RFOs) are accumulated in plant tissues stabilizing and protecting cell membranes and keeping the vital functions. The enzyme galactinol synthase (GolS, EC 2.4.1.123) catalyzes the first step in the biosynthesis of RFOs. In our study, soybean events overexpressing 35S:AtGolS2 were molecularly, physiological, and agronomical characterized, under drought simulated in greenhouse and in field conditions during the crop season 2014/2015. The conventional soybean cultivar BRS 184 was transformed and five positive events were obtained. Four events transmitted the transgene to further generations and in the events 2Ia1 and 2Ia4, two to four copies of AtGols2 gene were observed. Results in greenhouse showed that the overexpression of AtGolS2 in genetically modified (GM) plants led to increased galactinol transcripts, probably resulting in changes in carbohydrate metabolism. Accumulation of these transcripts that may have acted as osmoprotectors, lead to higher drought tolerance and survival rate of 2Ia4 plants. In addition, in field conditions, higher yield was observed for 2Ia4 plants under irrigated (IRR) and non-irrigated (NIRR) treatments. This result can be due to the increased synthesis of RFOs even under well-watered conditions. This field screening showed promising results for drought tolerance, suggesting that 2Ia4 plants may be useful in a breeding program for the development of drought-tolerant plants. However, additional studies are needed in further crop seasons and other sites to better characterize how these plants may outperform the WT plants under water deficit.

Rainfall is among the climatic factors that most affect production, as in the Brazilian Cerrado. Non-destructive and automated phenotyping methods are fast and efficient for genotype selection. The objective of this work was to evaluate, under field conditions, the morphophysiological changes, yield, and grain quality of soybean (Glycine max L. Merrill) under water stress in the Brazilian Cerrado. The plots comprised six soybean cultivars and the subplots of four water regimes, corresponding to 31, 44, 64 and 100% of crop evapotranspiration replacement. The experiments were conducted from May to September 2018 and 2019. An irrigation system with a bar of sprinklers with different flow rates was used. Gas exchange, vegetation indices (measured using a hyperspectral sensor embedded in a drone), yield and grain quality were evaluated. Water stress had different effects on gas exchange, vegetation indices, grain yield and chemical composition among the cultivars. Embrapa cultivar BRS 7280 Roundup ready (RR) and Nidera cultivar NA 5909 RG (glyphosate resistant) are yield stable and have a greater tolerance to drought. BRS 7280RR showed a higher tolerance to drought and higher water use efficiency (WUE) than all other tested cultivars. Vegetation indices, such as the NDVI (Normalized Difference Vegetation Index), correlated with the morphophysiological traits, such as plant height, were the most responsive variables to water stress. The NDVI can be used to predict soybean yield as a tool in a selection program under drought.

BackgroundThe drought condition is responsible for considerable losses in soybean production, which in turn may result in billionaire losses. After drought perception, plants activate a cascade of protecting genes against water deficit (WD), many of which are responsive to abscisic acid, the most important phytohormone to plants’ adaptation. This work aimed to recover abscisic acid (ABA)-responsive differentially expressed genes (DEG) from an RNA-Seq, carried out from leaves and roots of drought-sensitive (BR16) and tolerant (Embrapa 48) soybean cultivars under mild (MiWD), moderate (MoWD), and severe (SWD) water-deficit treatments. Enriched ABA-responsive pathways important to drought tolerance in soybean were also identified.ResultsIn drought-sensitive and tolerant soybean cultivars, approximately 75% of genes were identified as ABA-responsive by containing more than two ABRE (ABA-responsive elements) in the promoter region. Most of these genes were positively regulated. Roots were the tissue with more ABA-responsive genes and pathways triggered in response to WD in both cultivars, although, on the tolerant cultivar, these pathways were higher expressed. The most important enriched ABA pathways observed in the roots of both cultivars were involved in sugar and sulfur amino acid biosynthesis, osmoregulation, and crosstalk among ABA and ethylene, jasmonate, auxin, and cytokinin. Other pathways enriched were involved in phytoalexin production, ROS homeostasis, and membrane stability by glycerolipid and glycerophospholipid production. ABA-responsive genes were also ordered based on their expression profile in tissue and cultivar, and nine confidence groups could be observed. More than 80% of these clustered genes showed the same regulation profile under MiWd, MoWD, and SWD treatments. Activation of ABA biosynthesis under water deficit was validated by RT-qPCR by increasing the expression level of NCED3, an important enzyme in this pathway, and GOLS, a known ABA-responsive gene.ConclusionsA robust catalog of ABA-responsive genes was made available in this work. Considering ABA’s role in drought-response mechanisms, the genes in the groups pointed out in this study would be reliable candidates to be used in strategies to develop soybean lines more tolerant to drought. This paper, presented for the first time, ABA-responsive genes and ABA-enriched pathways in contrasting soybean cultivars for drought tolerance.