Transgenic Seeds Research Articles

Targeted engineering of camelina and pennycress seeds for ultrahigh accumulation of acetyl-TAG

Acetyl-TAG (3-acetyl-1,2-diacylglycerol), unique triacylglycerols (TAG) possessing an acetate group at the sn-3 position, exhibit valuable properties, such as reduced viscosity and freezing points. Previous attempts to engineer acetyl-TAG production in oilseed crops did not achieve the high levels found in naturally producing Euonymus seeds. Here, we demonstrate the successful generation of camelina and pennycress transgenic lines accumulating nearly pure acetyl-TAG at 93 mol% and 98 mol%, respectively. These ultrahigh acetyl-TAG synthesizing lines were created using gene-edited FATTY ACID ELONGASE1 (FAE1) mutant lines as an improved genetic background to increase levels of acetyl-CoA available for acetyl-TAG synthesis mediated by the expression of EfDAcT, a high-activity diacylglycerol acetyltransferase isolated from Euonymus fortunei. Combining EfDAcT expression with suppression of the competing TAG-synthesizing enzyme DGAT1 further enhanced acetyl-TAG accumulation. These ultrahigh levels of acetyl-TAG exceed those in earlier engineered oilseeds and are equivalent or greater than those in Euonymus seeds. Imaging of lipid localization in transgenic seeds revealed that the low amounts of residual TAG were mostly confined to the embryonic axis. Similar spatial distributions of specific TAG and acetyl-TAG molecular species, as well as their probable diacylglycerol (DAG) precursors, provide additional evidence that acetyl-TAG and TAG are both synthesized from the same tissue-specific DAG pools. Remarkably, this ultrahigh production of acetyl-TAG in transgenic seeds exhibited minimal negative effects on seed properties, highlighting the potential for production of designer oils required for economical biofuel industries.

Transgenic rice with microbial high-temperature-resistant β-glucosidase gene significantly improved 2-acetyl-1-pyrroline content and edible quality.

The content of 2-acetyl-1-pyrroline (2-AP) directly affects the aroma and taste of rice. Δ1-Pyrroline and methylglyoxal are the precursors of 2-AP synthesis, and β-glucosidase plays an important role in the synthesis of methylglyoxal. In this study, β-glucosidase gene cloned from Pyrococcus furiosus was molecularly modified to obtain the high-temperature-resistant β-glucosidase gene 371-β-glucosidase (T371A), which was transformed into kitaake varieties (Oryza sativa L. subsp. japonica) by Agrobacterium-mediated transformation method, and transgenic rice with heterologous expression of T371A was obtained. Experiments were conducted in transgenic rice to investigate whether this gene had an effect on the synthesis of 2-AP. Under the optimum reaction temperature of 50°C and cooking temperature of 100°C, the enzyme activity of β-glucosidase in transgenic rice seeds was prominently increased by 260-280% and 419-426% over that of the control, respectively. The content of 2-AP in transgenic rice seeds significantly increased by 75-105% under normal temperature and high-temperature cooking conditions compared with the control. It was also found that transgenic rice increased the content of methylglyoxal and decreased the expression of betaine aldehyde dehydrogenase (BADH2). The high-temperature-tolerant β-glucosidase gene obtained in this study provides an innovative technical strategy for molecular breeding of high-edible aroma crops and has wide application potential. © 2024 Society of Chemical Industry.

Agrobacterium-mediated Cuscuta campestris transformation as a tool for understanding plant-plant interactions.

Cuscuta campestris, a stem parasitic plant, has served as a valuable model plant for the exploration of plant-plant interactions and molecular trafficking. However, a major barrier to C. campestris research is that a method to generate stable transgenic plants has not yet been developed. Here, we describe the development of a Cuscuta transformation protocol using various reporter genes (GFP, GUS, or RUBY) and morphogenic genes (CcWUS2 and CcGRF/GIF), leading to a robust protocol for Agrobacterium-mediated C. campestris transformation. The stably transformed and regenerated RUBY C. campestris plants produced haustoria, the signature organ of parasitic plants, and these were functional in forming host attachments. The locations of T-DNA integration in the parasite genome were confirmed through TAIL-PCR. Transformed C. campestris also produced flowers and viable transgenic seeds exhibiting betalain pigment, providing proof of germline transmission of the RUBY transgene. Furthermore, RUBY is not only a useful selectable marker for the Agrobacterium-mediated transformation, but may also provide insight into the movement of molecules from C. campestris to the host during parasitism. Thus, the protocol for transformation of C. campestris reported here overcomes a major obstacle to Cuscuta research and opens new possibilities for studying parasitic plants and their interactions with hosts.

Investigation of the toxicity and safety concerns of transgenic maize seeds expressing immunogenic F and HN protein genes against Newcastle disease virus.

The presented study investigated the potential toxicity and safety concerns associated with transgenic maize seeds expressing immunogenic F and HN protein genes against Newcastle disease virus (NDV). The experiment involved feeding Sprague-Dawley rats with transgenic maize seeds formulated into standard diets at levels of 30% (w/w) for a duration of 90days. The rats were divided into three groups, with 10 rats per group. We assessed various parameters including overall appearance, feed consumption, body weight, organ weight, hematological parameters, serum chemistry, and histopathology. The results of these assessments were compared between the control group and the treatment groups. The study findings revealed that there were no significant differences between the control and treatment groups in terms of overall appearance, feed consumption, body weight, organ weight, hematological parameters, serum chemistry, microscopic histopathology, and gross appearance of tissues. These observations suggest that the consumption of transgenic maize seeds did not lead to any treatment-related adverse effects or toxicological issues. Furthermore, the transgenic maize seeds were found to be nutritionally equivalent to their non-transgenic counterpart. Overall, no physiological, pathological, or molecular toxicity was observed in the Rats fed with transgenic feed.However, it is important to note that this study focused specifically on the parameters measured and the outcomes observed in Sprague-Dawley rats, and further research and studies are necessary to fully evaluate the safety and potential applications of transgenic edible vaccines in humans or other animals.

Overexpression of protection of telomeres 1 (POT1), a single-stranded DNA-binding proteins in alfalfa (Medicago sativa), enhances seed vigor

Extensive bodies of research are dedicated to the study of seed aging with a particular focus on the roles of reactive oxygen species (ROS), and the ensuing oxidative damage during storage, as a primary cause of seed vigor decreasing. ROS diffuse to the nucleus and damage the telomeres, resulting in a loss of genetic integrity. Protection of telomeres 1 (POT1) is a telomeric protein that binds to the telomere region, and plays an essential role in maintaining genomic stability in plants. In this study, there were totally four MsPOT1 genes obtained from alfalfa genome. Expression analysis of four MsPOT1 genes in germinated seed presented the different expressions. Four MsPOT1 genes displayed high expression levels at the early stage of seed germination, Among the four POT1 genes, it was found that MS. gene040108 was significantly up-regulated in the early germination stage of CK seeds, but down-regulated in aged seeds. RT-qPCR assays and RNA-seq data revealed that MsPOT1-X gene was significantly induced by seed aging treatment. Transgenic seeds overexpressing MsPOT1-X gene in Arabidopsis thaliana and Medicago trunctula exhibited enhanced seed vigor, telomere length, telomerase activity associated with reduced H2O2 content. These results would provide a new way to understand aging stress-responsive MsPOT1 genes for genetic improvement of seed vigor. Although a key gene regulating seed vigor was identified in this study, the specific mechanism of MsPOT1-X gene regulating seed vigor needs to be further explored.

MYB4 in Lilium pumilum affects plant saline-alkaline tolerance

ABSTRACT Lilium pumilum DC (L. pumilum DC) plays an important role in the rational utilization of salinized soil. To explore the molecular mechanism of salt-tolerant L. pumilum, the LpMYB4 was cloned. LpMYB4 close relationship with Bambusa emeiensis and Zea mays MYB4 throughout the phylogenetic tree construction. LpMYB4 protein was found to be localized in the nucleus. Prokaryotic and eukaryotic bacterial solution resistance experiments proved that the exogenous introduction of LpMYB4 made the overexpression strains obtain better survival ability under saline-alkaline stress. Compared with wild-type plants, tobacco plants overexpressing LpMYB4 had better growth and lower leaf wilting and lodging, the content of chlorophyll was higher, the content of hydrogen peroxide and superoxide anion was lower, the activity of peroxidase and superoxide dismutase was higher and the relative conductivity was lower under saline-alkaline stress. The analysis of seed germination and seedling resistance of transgenic plants under salt stress showed that LpMYB4 transgenic seeds were more tolerant to salt stress during germination and growth. Yeast two-hybrid and two-luciferase complementation experiments showed that LpMYB4 interacted with yeast two-hybrid and LpGPX6. The analysis of the role of LpMYB4 in improving plant saline-alkali resistance is helpful to the transformation of plant germplasm resources and has great significance for agriculture and sustainable development.

Cloning and functional characterization of the legumin A gene (EuLEGA) from Eucommia ulmoides Oliver

Legumin A is a seed storage protein that provides nutrients for seed germination. The purpose of this study was to describe the structure and expression pattern of the EuLEGA gene in Eucommia ulmoides Oliver (E. ulmoides) and to infer its functional role. The 1287 bp coding sequence of the EuLEGA CDS of the EuLEGA gene, encoding a protein containing 428 amino acid residues, was cloned. The structure predicted that the protein belonged to the RmlC (deoxythymidine diphosphates, dTDP)-4-dehydrorhamnose 3,5-epimerase)-like cupin conserved domain family, which contains both RmlC, a key enzyme for the synthesis of rhamnose and legumin A. The overexpression (OE) vector of the EuLEGA gene was constructed and genetically transformed into tobacco and E. ulmoides; the RNA interference (RNAi) vector of the EuLEGA gene was constructed and genetically transformed into E. ulmoides; and the contents of legumin A and rhamnose were detected. The results showed that the EuLEGA gene could significantly increase the content of legumin A in transgenic tobacco leaves and transgenic E. ulmoides regenerative buds, and the OE of this gene in E. ulmoides could promote an increase in rhamnose content. RNAi caused a significant decrease in the legumin A content in the regenerated buds of E. ulmoides. These was a significant increase in legumin A in the transgenic tobacco seeds, and these results indicate that the expression of the EuLEGA gene is closely related to the accumulation of legumin A. Subcellular localization studies revealed that EuLEGA is localized to the cytoplasm with the vacuolar membrane. Analysis of the EuLEGA gene expression data revealed that the expression level of the EuLEGA gene in the samaras was significantly greater than that in the leaves and stems. In addition, the study also demonstrated that GA3 can upregulate the expression levels of the EuLEGA gene, while ABA and MeJA can downregulate its expression levels.

Movement of ACC oxidase 3 mRNA from seeds to flesh promotes fruit ripening in apple

Xenia, the phenomenon in which the pollen genotype directly affects the phenotypic characteristics of maternal tissues (i.e., fruit ripening), has applications in crop production and breeding. However, the underlying molecular mechanism has yet to be elucidated. Here, we investigated whether mobile mRNAs from the pollen affect the ripening and quality-related characteristics of the fruit using cross-pollination between distinct Malus domestica (apple) cultivars. We demonstrated that hundreds of mobile mRNAs originating from the seeds are delivered to the fruit. We found that the movement of one of these mRNAs, ACC oxidase 3 (MdACO3), is coordinated with fruit ripening. Salicylic acid treatment, which can cause plasmodesmal closure, blocks MdACO3 movement, indicating that MdACO3 transcripts may move through the plasmodesmata. To assess the role of mobile MdACO3 transcripts in apple fruit, we created MdACO3-GFP-expressing apple seeds using MdACO3-GFP-overexpressing pollen for pollination and showed that MdACO3 transcripts in the transgenic seeds move to the flesh, where they promote fruit ripening. Furthermore, we demonstrated that MdACO3 can be transported from the seeds to fruit in the fleshy-fruited species tomato and strawberry. These results underscore the potential of mobile mRNAs from seeds to influence fruit characteristics, providing an explanation for the xenia phenomenon. Notably, our findings highlight the feasibility of leveraging diverse pollen genomic resources, without resorting to genome editing, to improve fruit quality.

Transgenic Soybean for Production of Thermostable α-Amylase.

Alpha-amylases are crucial hydrolase enzymes which have been widely used in food, feed, fermentation, and pharmaceutical industries. Methods for low-cost production of α-amylases are highly desirable. Soybean seed, functioning as a bioreactor, offers an excellent platform for the mass production of recombinant proteins for its ability to synthesize substantial quantities of proteins. In this study, we generated and characterized transgenic soybeans expressing the α-amylase AmyS from Bacillus stearothermophilus. The α-amylase expression cassettes were constructed for seed specific expression by utilizing the promoters of three different soybean storage peptides and transformed into soybean via Agrobacterium-mediated transformation. The event with the highest amylase activity reached 601 U/mg of seed flour (one unit is defined as the amount of enzyme that generates 1 micromole reducing ends per min from starch at 65 °C in pH 5.5 sodium acetate buffer). The optimum pH, optimum temperature, and the enzymatic kinetics of the soybean expressed enzyme are similar to that of the E. coli expressed enzyme. However, the soybean expressed α-amylase is glycosylated, exhibiting enhanced thermostability and storage stability. Soybean AmyS retains over 80% activity after 100 min at 75 °C, and the transgenic seeds exhibit no significant activity loss after one year of storage at room temperature. The accumulated AmyS in the transgenic seeds represents approximately 15% of the total seed protein, or about 4% of the dry seed weight. The specific activity of the transgenic soybean seed flour is comparable to many commercial α-amylase enzyme products in current markets, suggesting that the soybean flour may be directly used for various applications without the need for extraction and purification.

Cytochrome GmGLY1 is Involved in the Biosynthesis of Glycitein in Soybean.

Isoflavones, the major secondary metabolites of interest due to their benefits to both human and plant health, are exclusively produced by legumes. In this study, we profiled the isoflavone content in dry seeds from 211 soybean [Glycine max (L.) Merr.] accessions grown across five environments. Broad and discernible phenotypic variations were observed among accessions, regions, and years of growth. Twenty-six single-nucleotide polymorphisms (SNPs) associated with the sum of glycitein (GLE), glycitin (GL), 6″-O-acetylglycitin (AGL), and 6″-O-malonylglycitin (MGL) contents were detected in multiple environments via a genome-wide association study (GWAS). These SNPs were located on chromosome 11 (8,148,438 bp to 8,296,956 bp, renamed qGly11-01). Glyma.11g108300 (GmGLY1), a gene that encodes a P450 family protein, was identified via sequence variation analysis, functional annotation, weighted gene coexpression network analysis (WGCNA), and expression profile analysis of candidate gene, and hairy roots transformation in soybean. Overexpression of GmGLY1 increased the glycitein content (GLC) in soybean hairy roots and transgenic seeds, while CRISPR/Cas9-generated mutants exhibited decreased GLC and increased daidzein content (DAC). Haplotype analysis revealed that GmGLY1 allelic variations significantly affect the GLC accumulation. These findings enhance our understanding of genes influencing GLC in soybean and may guide breeding for lines with high and stable GLC.

Verification of the peanut vitamin C synthesis-related gene AhPMM and its role in stress resistance

Vitamin C plays an important role in plant antioxidation, photosynthesis, growth and development, and metabolism. In this study, a gene AhPMM, which is involved in vitamin C synthesis and responds significantly to low temperature, NaCl, polyethylene glycol (PEG) and abscisic acid (ABA) treatments, was cloned from peanut. An AhPMM overexpression vector was constructed, and transferred to a peanut variety Junanxiaohong using the pollen tube injection method. PCR test on the T3 generation transgenic peanut plants showed a transgenics positive rate of 42.3%. HPLC was used to determine the content of reducing vitamin C (AsA) and total vitamin C in the leaves of transgenic plants. The results showed that the content of AsA in some lines increased significantly, up to 1.90 times higher than that of the control, and the total vitamin content increased by up to 1.63 times compared to that of the control. NaCl and ABA tolerance tests were carried out on transgenic seeds. The results showed that the salt tolerance of transgenic seeds was significantly enhanced and the sensitivity to ABA was weakened compared to that of the non-transgenic control. Moreover, the salt tolerance of the transgenic plants was also significantly enhanced compared to that of the non-transgenic control. The above results showed that AhPMM gene not only increased the vitamin C content of peanut, but also increased the salt tolerance of transgenic peanut seeds and plants. This study may provide a genetic source for the molecular breeding of peanut for enhanced salt tolerance.

Peanut LEAFY COTYLEDON1-type genes participate in regulating the embryo development and the accumulation of storage lipids.

Two peanut LEC1-type genes exhibit partial functional redundancy. AhNFYB10 could complement almost all the defective phenotypes of lec1-2 in terms of embryonic morphology, while AhNF-YB1 could partially affect these phenotypes. LEAFY COTYLEDON1 (LEC1) is a member of the nuclear factor Y (NF-Y) family of transcription factors and has been identified as a key regulator of embryonic development. In the present study, two LEC1-type genes from Arachis hypogeae were identified and designated as AhNF-YB1 and AhNF-YB10; these genes belong to subgenome A and subgenome B, respectively. The functions of AhNF-YB1 and AhNF-YB10 were investigated by complementation analysis of their defective phenotypes of the Arabidopsis lec1-2 mutant and by ectopic expression in wild-type Arabidopsis. The results indicated that both AhNF-YB1 and AhNF-YB10 participate in regulating embryogenesis, embryo development, and reserve deposition in cotyledons and that they have partial functional redundancy. In contrast, AhNF-YB10 complemented almost all the defective phenotypes of lec1-2 in terms of embryonic morphology and hypocotyl length, while AhNF-YB1 had only a partial effect. In addition, 30-40% of the seeds of the AhNF-YB1 transformants exhibited a decreasing germination ratio and longevity. Therefore, appropriate spatiotemporal expression of these genes is necessary for embryo morphogenesis at the early development stage and is responsible for seed maturation at the mid-late development stage. On the other hand, overexpression of AhNF-YB1 or AhNF-YB10 at the middle to late stages of Arabidopsis seed development improved the weight, oil content, and fatty acid composition of the transgenic seeds. Moreover, the expression levels of several genes associated with fatty acid synthesis and embryogenesis were significantly greater in developing AhNF-YB10-overexpressing seeds than in control seeds. This study provides a theoretical basis for breeding oilseed crops with high yields and high oil content.

ABA Inhibits Rice Seed Aging by Reducing H2O2 Accumulation in the Radicle of Seeds

The seed, a critical organ in higher plants, serves as a primary determinant of agricultural productivity, with its quality directly influencing crop yield. Improper storage conditions can diminish seed vigor, adversely affecting seed germination and seedling establishment. Therefore, understanding the seed-aging process and exploring strategies to enhance seed-aging resistance are paramount. In this study, we observed that seed aging during storage leads to a decline in seed vigor and can coincide with the accumulation of hydrogen peroxide (H2O2) in the radicle, resulting in compromised or uneven germination and asynchronous seedling emergence. We identified the abscisic acid (ABA) catabolism gene, abscisic acid 8′-hydroxylase 2 (OsABA8ox2), as significantly induced by aging treatment. Interestingly, transgenic seeds overexpressing OsABA8ox2 exhibited reduced seed vigor, while gene knockout enhanced seed vigor, suggesting its role as a negative regulator. Similarly, seeds pretreated with ABA or diphenyleneiodonium chloride (DPI, an H2O2 inhibitor) showed increased resistance to aging, with more robust early seedling establishment. Both OsABA8ox2 mutant seeds and seeds pretreated with ABA or DPI displayed lower H2O2 content during aging treatment. Overall, our findings indicate that ABA mitigates rice seed aging by reducing H2O2 accumulation in the radicle. This study offers valuable germplasm resources and presents a novel approach to enhancing seed resistance against aging.

Biofortified Rice Provides Rich Sakuranetin in Endosperm

Sakuranetin plays a key role as a phytoalexin in plant resistance to biotic and abiotic stresses, and possesses diverse health-promoting benefits. However, mature rice seeds do not contain detectable levels of sakuranetin. In the present study, a transgenic rice plant was developed in which the promoter of an endosperm-specific glutelin gene OsGluD-1 drives the expression of a specific enzyme naringenin 7-O-methyltransferase (NOMT) for sakuranetin biosynthesis. The presence of naringenin, which serves as the biosynthetic precursor of sakuranetin made this modification feasible in theory. Liquid chromatography tandem mass spectrometry (LC–MS/MS) validated that the seeds of transgenic rice accumulated remarkable sakuranetin at the mature stage, and higher at the filling stage. In addition, the panicle blast resistance of transgenic rice was significantly higher than that of the wild type. Specially, the matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) imaging was performed to detect the content and spatial distribution of sakuranetin and other nutritional metabolites in transgenic rice seeds. Notably, this genetic modification also did not change the nutritional and quality indicators such as soluble sugars, total amino acids, total flavonoids, amylose, total protein, and free amino acid content in rice. Meanwhile, the phenotypes of the transgenic plant during the whole growth and developmental periods and agricultural traits such as grain width, grain length, and 1000-grain weight exhibited no significant differences from the wild type. Collectively, the study provides a conceptual advance on cultivating sakuranetin-rich biofortified rice by metabolic engineering. This new breeding idea may not only enhance the disease resistance of cereal crop seeds but also improve the nutritional value of grains for human health benefits.

Overexpression of the OsFes1A increased the phytosterols content and enhanced drought and salt stress tolerance in Arabidopsis.

Overexpression of the rice gene, OsFes1A, increased phytosterol content and drought and salt stress tolerance in Arabidopsis. Phytosterols are key components of the phospholipid bilayer membrane and regulate various processes of plant growth and response to biotic and abiotic stresses. In this study, it was demonstrated that the overexpression of OsFes1A (Hsp70 nucleotide exchange factor Fes1) increased phytosterols content and enhanced tolerance to salt and drought stress in Arabidopsis. In transgenic plants, the average content of campesterol was 17.6% higher than that of WT, and the average content of β-sitosterol reached 923.75μg/g, with an increase of 1.33-fold. In fes1a seeds, the contents of campesterol and β-sitosterol reduced by 20% and 10.93%, respectively. In OsFes1A transgenic seeds, the contents of campesterol and β-sitosterol increased by 1.38-fold and 1.25-fold respectively. Furthermore, the germination rate of transgenic Arabidopsis was significantly higher than WT under stress (salt, ABA, and drought treatment). Under salt stress, transgenic plants accumulated a lower MDA content, higher chlorophyll content, and POD activity relative to the wild type, while the mutants showed the opposite pattern Our study found multiple other functions of OsFes1A beyond the defined role of Fes1 in regulating Hsp70, contributing to the better understanding of the essential roles of Fes1 in plants. Meanwhile, it provides the theoretical basis for developing high phytosterol crop varieties.

OsCSD2 and OsCSD3 Enhance Seed Storability by Modulating Antioxidant Enzymes and Abscisic Acid in Rice.

Seed deterioration during storage poses a significant challenge to rice production, leading to a drastic decline in both edible quality and viability, thereby impacting overall crop yield. This study aimed to address this issue by further investigating candidate genes associated with two previously identified QTLs for seed storability through genome association analysis. Among the screened genes, two superoxide dismutase (SOD) genes, OsCSD2 (Copper/zinc Superoxide Dismutase 2) and OsCSD3, were selected for further study. The generation of overexpression and CRISPR/Cas9 mutant transgenic lines revealed that OsCSD2 and OsCSD3 play a positive regulatory role in enhancing rice seed storability. Subsequent exploration of the physiological mechanisms demonstrated that overexpression lines exhibited lower relative electrical conductivity, indicative of reduced cell membrane damage, while knockout lines displayed the opposite trend. Furthermore, the overexpression lines of OsCSD2 and OsCSD3 showed significant increases not only in SOD but also in CAT and POD activities, highlighting an augmented antioxidant system in the transgenic seeds. Additionally, hormone profiling indicated that ABA contributed to the improved seed storability observed in these lines. In summary, these findings provide valuable insights into the regulatory mechanisms of OsCSDs in rice storability, with potential applications for mitigating grain loss and enhancing global food security.

A universal design of restructured dimer antigens: Development of a superior vaccine against the paramyxovirus in transgenic rice

The development of vaccines, which induce effective immune responses while ensuring safety and affordability, remains a substantial challenge. In this study, we proposed a vaccine model of a restructured "head-to-tail" dimer to efficiently stimulate B cell response. We also demonstrate the feasibility of using this model to develop a paramyxovirus vaccine through a low-cost rice endosperm expression system. Crystal structure and small-angle X-ray scattering data showed that the restructured hemagglutinin-neuraminidase (HN) formed tetramers with fully exposed quadruple receptor binding domains and neutralizing epitopes. In comparison with the original HN antigen and three traditional commercial whole virus vaccines, the restructured HN facilitated critical epitope exposure and initiated a faster and more potent immune response. Two-dose immunization with 0.5 μg of the restructured antigen (equivalent to one-127th of a rice grain) and one-dose with 5 μg completely protected chickens against a lethal challenge of the virus. These results demonstrate that the restructured HN from transgenic rice seeds is safe, effective, low-dose useful, and inexpensive. We provide a plant platform and a simple restructured model for highly effective vaccine development.

Overexpression of the alfalfa (Medicago sativa) gene, MsKMS1, negatively regulates seed germination in transgenic tobacco (Nicotiana tabacum).

Soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE)-associated proteins are a class of transmembrane proteins involved in intracellular trafficking pathways. However, the functions of many SNARE domain-containing proteins remain unclear. We have previously identified a SNARE-associated gene in alfalfa (Medicago sativa ) KILLING ME SLOWLY1 (MsKMS1 ), which is involved in various abiotic stresses. In this study, we investigated the function of MsKMS1 in the seed germination of transgenic tobacco (Nicotiana tabacum ). Phylogenetic analysis showed that MsKMS1 was homologous to the SNARE-associated or MAPR component-related proteins of other plants. Germination assays revealed that MsKMS1 negatively regulated seed germination under normal, D-mannitol and abscisic acid-induced stress conditions, yet MsKMS1 -overexpression could confer enhanced heat tolerance in transgenic tobacco. The suppressive effect on germination in MsKMS1 -overexpression lines was associated with higher abscisic acid and salicylic acid contents in seeds. This was accompanied by the upregulation of abscisic acid biosynthetic genes (ZEP and NCED ) and the downregulation of gibberellin biosynthetic genes (GA20ox2 and GA20ox3 ). Taken together, these results suggested that MsKMS1 negatively regulated seed germination by increasing abscisic acid and salicylic acid contents through the expression of genes related to abscisic acid and gibberellin biosynthesis. In addition, MsKMS1 could improve heat tolerance during the germination of transgenic tobacco seeds.

Overexpression of GhSWEET42, a SWEET-like gene from cotton, enhances the oil content and seed size

SWEET (‘sugars will eventually be exported transporters’) family genes reportedly play a critical role in sugar translocation and oil biosynthesis in various plant species. However, their functions in cotton are unknown. The present study demonstrated that while GhSWEET42 was widely expressed in different cotton tissues, it had the highest expression level in the developing ovules. Hence, it performs a vital role in seed development. We constructed GhSWEET42 transgenic Arabidopsis lines to verify the biological function of this gene and found that the oil content and weight of the seeds produced by the overexpression lines were 18–23% and 19–20% higher, respectively than those of the wild-type. Gas chromatography–mass spectrometry (GC–MS) analysis revealed that it was mainly a relative increase in unsaturated fatty acids (FAs) that contributed to the relative increase in oil content in the transgenic seeds. Moreover, the latter exhibited comparative upregulation of certain genes associated with FA and triacylglycerol biosynthesis as well as cell expansion. GhSWEET42 might work synergistically with the aforementioned genes. This finding indicates that GhSWEET42 may be essential in oil biosynthesis and seed development in cotton. The results of the present work may facilitate further explorations into the molecular mechanism of cottonseed oil biosynthesis as well as the cultivation of novel oil-rich cotton varieties.

Defining the physical properties of blends of acetyl‐triacylglycerols derived from transgenic oil seeds

AbstractAcetyl‐TAG are unusually structured triacylglycerols (TAG) that possess an acetate group at their sn‐3 position. Acetyl‐TAG have different physical properties compared to regular TAG, including lower viscosity and improved cold temperature properties, making acetyl‐TAG useful for different applications, including as a diesel replacement. These unusual TAG molecules can be synthesized in the seeds of transgenic plants through the expression of diacylglycerol acetyltransferase (DAcT) enzymes isolated from different Euonymus species. In this study, the kinematic viscosity as well as the crystallization and melting behavior of blends of acetyl‐TAG and regular TAG were examined to define goals for acetyl‐TAG synthesis in transgenic plants. Even small amounts of regular TAG when blended with acetyl‐TAG had a disproportionate effect on the viscosity of mixture. This effect of regular TAG in increasing kinematic viscosity was more pronounced at lower temperatures. Under slow cooling conditions, the two different TAGs and their blends possessed two main crystallization events with different degree of separation of the thermal transition, and the lower crystallization temperature decreased with increasing amounts of acetyl‐TAG in the blend. At higher cooling rates, one broad and tailed crystallization peak was observed. Heating thermograms indicate similar polymorphic behavior of the blends and a general peak shift to lower transition range with increasing acetyl‐TAG compared to the two pure lipids. This information about the viscosity and thermal properties of blends of TAG and acetyl‐TAG will provide useful targets for engineering higher levels of acetyl‐TAG in transgenic seeds.