Pyramiding of transgenic immune receptors from primary and tertiary wheat gene pools improves powdery mildew resistance in the field.

Introgression of resistance genes from wild or related species is a common strategy to improve disease resistance of wheat cultivars. Pm17 is a gene that confers powdery mildew resistance in wheat. It encodes an NLR type of immune receptor and was introgressed from rye to wheat as part of the 1RS chromosome arm translocation several decades ago. So far it has not been possible to separate Pm17 from its co-introgressed rye genes due to suppressed recombination. Here we tested in the field transgenic Bobwhite wheat overexpressing Pm17 without any other rye genes. Four transgenic events showed high levels of PM17 protein accumulation, strong powdery mildew resistance, and no pleiotropic effects during three field seasons. We used a combined approach of transgene insertion and cross-breeding to generate lines co-expressing Pm17 and Pm3, or Pm17 and Pm8. Blumeria graminis f. sp. tritici infection tests confirmed additive, race-specific resistance of the two pyramided transgenes in lines Pm17+Pm3b and Pm17+Pm8. Furthermore, pyramided lines showed strong powdery mildew resistance during three field seasons. We conclude that the combination of overexpressed NLR genes from the extended gene pool broadens and diversifies wheat disease resistance.

Breeding for durable resistance against biotrophic fungal pathogens using transgenes from wheat.

Breeding for resistant crops is a sustainable way to control disease and relies on the introduction of novel resistance genes. Here, we tested three strategies on how to use transgenes from wheat to achieve durable resistance against fungal pathogens in the field. First, we tested the highly effective, overexpressed single transgene Pm3e in the background of spring wheat cultivar Bobwhite in a long-term field trial over many years. Together with previous results, this revealed that transgenic wheat line Pm3e#2 conferred complete powdery mildew resistance during a total of nine field seasons without a negative impact on yield. Furthermore, overexpressed Pm3e provided resistance to powdery mildew isolates from our worldwide collection when crossed into the elite wheat cultivar Fiorina. Second, we pyramided the four overexpressed transgenes Pm3a, Pm3b, Pm3d, and Pm3f in the background of cultivar Bobwhite and showed that the pyramided line Pm3a,b,d,f was completely resistant to powdery mildew in five field seasons. Third, we performed field trials with three barley lines expressing adult plant resistance gene Lr34 from wheat during three field seasons. Line GLP8 expressed Lr34 under control of the pathogen-inducible Hv-Ger4c promoter and provided partial barley powdery mildew and leaf rust resistance in the field with small, negative effects on yield components which might need compensatory breeding. Overall, our study demonstrates and discusses three successful strategies for achieving fungal disease resistance of wheat and barley in the field using transgenes from wheat. These strategies might confer long-term resistance if applied in a sustainable way. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-024-01451-2.

Bioregulators: unlocking their potential role in regulation of the plant oxidative defense system.

KEY MESSAGE: Plant bioregulators play an important role in managing oxidative stress tolerance in plants. Utilizing their ability in stress sensitive crops through genetic engineering will be a meaningful approach to manage food production under the threat of climate change. Exploitation of the plant defense system against oxidative stress to engineer tolerant plants in the climate change scenario is a sustainable and meaningful strategy. Plant bioregulators (PBRs), which are important biotic factors, are known to play a vital role not only in the development of plants, but also in inducing tolerance in plants against various environmental extremes. These bioregulators include auxins, gibberellins, cytokinins, abscisic acid, brassinosteroids, polyamines, strigolactones, and ascorbic acid and provide protection against the oxidative stress-associated reactive oxygen species through modulation or activation of a plant's antioxidant system. Therefore, exploitation of their functioning and accumulation is of considerable significance for the development of plants more tolerant of harsh environmental conditions in order to tackle the issue of food security under the threat of climate change. Therefore, this review summarizes a new line of evidence that how PBRs act as inducers of oxidative stress resistance in plants and how they could be modulated in transgenic crops via introgression of genes. Reactive oxygen species production during oxidative stress events and their neutralization through an efficient antioxidants system is comprehensively detailed. Further, the use of exogenously applied PBRs in the induction of oxidative stress resistance is discussed. Recent advances in engineering transgenic plants with modified PBR gene expression to exploit the plant defense system against oxidative stress are discussed from an agricultural perspective.

Linking land use patterns and pest outbreaks in Bt maize.

Western corn rootworm, Diabrotica virgifera virgifera LeConte (Coleoptera: Chrysomelidae), is a major pest of maize in the United States and is an invasive pest in Europe. Maize is the only agricultural crop on which western corn rootworm larvae can survive and this insect requires two consecutive years of maize cultivation to complete its life cycle. Transgenic maize producing insecticidal proteins derived from the bacterium Bacillus thuringiensis (Bt) is often used to manage rootworm populations. The first Bt trait, Cry3Bb1, was introduced in 2003, but larval resistance to this toxin appeared in northeastern Iowa in 2009. Rootworm management occurs on a field-by-field basis, but adult rootworm may disperse among fields. It is known that growing consecutive years of Cry3Bb1 maize within a field can lead to resistance, but the relationship of the surrounding landscape to the development of resistance is unknown. Using geospatial tools and publicly available land-use data, we examined circular areas (buffers) surrounding fields that had previously experienced high levels of rootworm injury to Cry3Bb1 maize and rootworm resistance to Cry3Bb1 maize (problem fields). We calculated the proportion of area inside each buffer planted to maize continuously for 1-9 yr, and compared these values to those for randomly selected control points throughout the state. We also calculated the proportion of the state planted to maize for at least three consecutive years for 2003 through 2018, and its relationship with the annual value of maize. We found that areas surrounding problem fields had significantly more continuous maize compared to controls, with the most continuous maize within 1.6 km of problem fields. We also found that the cultivation of continuous maize in Iowa increased significantly between 2003 and 2018, and this was correlated with average annual price of maize. We hypothesize a scenario in which continuous cultivation of Cry3Bb1 maize in local landscapes, driven in part by the increased value of maize, facilitated selection for Cry3Bb1 resistance. These results suggest that land use in areas surrounding problem fields affect the rate of resistance evolution and approaches for resistance management can be enhanced by taking a landscape-level perspective.

Current understanding of pattern-triggered immunity and hormone-mediated defense in rice (Oryza sativa) in response to Magnaporthe oryzae infection.

Plant pathogens represent a huge threat to world food security, affecting both crop production and quality. Although significant progress has been made in improving plant immunity by expressing key, defense-related genes and proteins from different species in transgenic crops, a challenge remains for molecular breeders and biotechnologists to successfully engineer elite, transgenic crop varieties with improved resistance against critical plant pathogens. Upon pathogen attack, including infection of rice (Oryza sativa) by Magnaporthe oryzae, host plants initiate a complex defense response at molecular, biochemical and physiological levels. Plants perceive the presence of pathogens by detecting microbe-associated molecular patterns via pattern recognition receptors, and initiate a first line of innate immunity, the so-called pattern-triggered immunity (PTI). This results in a series of downstream defense responses, including the production of hormones, which collectively function to fend off pathogen attacks. A variety of studies have demonstrated that many genes are involved in the defense response of rice to M. oryzae. In this review, the current understanding of mechanisms that improve rice defense response to M. oryzae will be discussed, with special focus on PTI and the phytohormones ethylene, jasmonic acid, salicylic acid, and abscisic acid; as well as on the mediation of defense signaling mechanisms by PTI and these hormones. Potential target genes that may serve as promising candidates for improving rice immunity against M. oryzae will also be discussed.

Insights obtained using different modules of the cotton uceA1.7 promoter.

MAIN CONCLUSION: The structure of the cotton uceA1.7 promoter and its modules was analyzed; the potential of their key sequences has been confirmed in different tissues, proving to be a good candidate for the development of new biotechnological tools. Transcriptional promoters are among the primary genetic engineering elements used to control genes of interest (GOIs) associated with agronomic traits. Cotton uceA1.7 was previously characterized as a constitutive promoter with activity higher than that of the constitutive promoter from the Cauliflower mosaic virus (CaMV) 35S gene in various plant tissues. In this study, we generated Arabidopsis thaliana homozygous events stably overexpressing the gfp reporter gene driven by different modules of the uceA1.7 promoter. The expression level of the reporter gene in different plant tissues and the transcriptional stability of these modules was determined compared to its full-length promoter and the 35S promoter. The full-length uceA1.7 promoter exhibited higher activity in different plant tissues compared to the 35S promoter. Two modules of the promoter produced a low and unstable transcription level compared to the other promoters. The other two modules rich in cis-regulatory elements showed similar activity levels to full-length uceA1.7 and 35S promoters but were less stable. This result suggests the location of a minimal portion of the promoter that is required to initiate transcription properly (the core promoter). Additionally, the full-length uceA1.7 promoter containing the 5'-untranslated region (UTR) is essential for higher transcriptional stability in various plant tissues. These findings confirm the potential use of the full-length uceA1.7 promoter for the development of new biotechnological tools (NBTs) to achieve higher expression levels of GOIs in, for example, the root or flower bud for the efficient control of phytonematodes and pest-insects, respectively, in important crops.

The quest for osmosensors in plants.

Osmotic stress has severe effects on crop productivity. Since climate change is predicted to exacerbate this problem, the development of new crops that are tolerant to osmotic stresses, especially drought and salinity stress, is required. However, only limited success has been achieved to date, primarily because of the lack of a clear understanding of the mechanisms that facilitate osmosensing. Here, we discuss the potential mechanisms of osmosensing in plants. We highlight the roles of proteins such as receptor-like kinases, which sense stress-induced cell wall damage, mechanosensitive calcium channels, which initiate a calcium-induced stress response, and phospholipase C, a membrane-bound enzyme that is integral to osmotic stress perception. We also discuss the roles of aquaporins and membrane-bound histidine kinases, which could potentially detect changes in extracellular osmolarity in plants, as they do in prokaryotes and lower eukaryotes. These putative osmosensors have the potential to serve as master regulators of the osmotic stress response in plants and could prove to be useful targets for the selection of osmotic stress-tolerant crops.

Integrating the dynamics of yield traits in rice in response to environmental changes.

Reductions in crop yields as a consequence of global climate change threaten worldwide food security. It is therefore imperative to develop high-yielding crop plants that show sustainable production under stress conditions. In order to achieve this aim through breeding or genetic engineering, it is crucial to have a complete and comprehensive understanding of the molecular basis of plant architecture and the regulation of its sub-components that contribute to yield under stress. Rice is one of the most widely consumed crops and is adversely affected by abiotic stresses such as drought and salinity. Using it as a model system, in this review we present a summary of our current knowledge of the physiological and molecular mechanisms that determine yield traits in rice under optimal growth conditions and under conditions of environmental stress. Based on physiological functioning, we also consider the best possible combination of genes that may improve grain yield under optimal as well as environmentally stressed conditions. The principles that we present here for rice will also be useful for similar studies in other grain crops.

Structure-Function Insights of Jaburetox and Soyuretox: Novel Intrinsically Disordered Polypeptides Derived from Plant Ureases.

Intrinsically disordered proteins (IDPs) and intrinsically disordered regions (IDRs) do not have a stable 3D structure but still have important biological activities. Jaburetox is a recombinant peptide derived from the jack bean (Canavalia ensiformis) urease and presents entomotoxic and antimicrobial actions. The structure of Jaburetox was elucidated using nuclear magnetic resonance which reveals it is an IDP with small amounts of secondary structure. Different approaches have demonstrated that Jaburetox acquires certain folding upon interaction with lipid membranes, a characteristic commonly found in other IDPs and usually important for their biological functions. Soyuretox, a recombinant peptide derived from the soybean (Glycine max) ubiquitous urease and homologous to Jaburetox, was also characterized for its biological activities and structural properties. Soyuretox is also an IDP, presenting more secondary structure in comparison with Jaburetox and similar entomotoxic and fungitoxic effects. Moreover, Soyuretox was found to be nontoxic to zebra fish, while Jaburetox was innocuous to mice and rats. This profile of toxicity affecting detrimental species without damaging mammals or the environment qualified them to be used in biotechnological applications. Both peptides were employed to develop transgenic crops and these plants were active against insects and nematodes, unveiling their immense potentiality for field applications.

Osmoprotection in plants under abiotic stresses: new insights into a classical phenomenon.

MAIN CONCLUSION: Plant osmoprotectants protect against abiotic stresses. Introgression of osmoprotectant genes into crop plants via genetic engineering is an important strategy in developing more productive plants. Plants employ adaptive mechanisms to survive various abiotic stresses. One mechanism, the osmoprotection system, utilizes various groups of low molecular weight compounds, collectively known as osmoprotectants, to mitigate the negative effect of abiotic stresses. Osmoprotectants may include amino acids, polyamines, quaternary ammonium compounds and sugars. These nontoxic compounds stabilize cellular structures and enzymes, act as metabolic signals, and scavenge reactive oxygen species produced under stressful conditions. The advent of recent drastic fluctuations in the global climate necessitates the development of plants better adapted to abiotic stresses. The introgression of genes related to osmoprotectant biosynthesis from one plant to another by genetic engineering is a unique strategy bypassing laborious conventional and classical breeding programs. Herein, we review recent literature related to osmoprotectants and transgenic plants engineered with specific osmoprotectant properties.

Alkaline phosphatase 2 is a functional receptor of Cry1Ac but not Cry2Ab in Helicoverpa zea.

Although membrane-bound alkaline phosphatases (ALPs) have been proposed as a receptor for Cry1Ac in a few lepidopteran species, their potential functions as a Cry2Ab receptor are yet to be verified. To determine if ALP2 also serves as a receptor for Cry1Ac and even for Cry2Ab in Helicoverpa zea, we measured the potency of activated Cry1Ac and Cry2Ab against midgut and fat body cell lines of H. zea and the ovarian cell line of Spodoptera frugiperda (Sf9) expressing H. zea ALP2 (HzALP2) or transfected with HzALP2 double-stranded RNA (dsRNA). Relative to the control cells, the three cell lines expressing HzALP2 were more susceptible to Cry1Ac but there was no difference for Cry2Ab. By contrast, the two H. zea cell lines transfected with HzALP2 dsRNA were resistant to Cry1Ac while kept susceptible to Cry2Ab. Furthermore, RNA interference knockdown of HzALP2 in H. zea larvae enhanced larval survival on Cry1Ac-containing diets. These findings indicate that HzALP2 functions as a receptor of Cry1Ac but not Cry2Ab.

Examining the behavior of crop-derived antibiotic resistance genes in anaerobic sludge batch reactors under thermophilic conditions.

The consumption of transgenic crops and their by-products has become increasingly common in the United States. Yet, uncertainty remains regarding the fate and behavior of DNA within food matrices once it exits the digestive track and enters into wastewater treatment plants (WWTPs). Because many transgenic crops have historically contained antibiotic resistance genes as selection markers, understanding the behavior and uptake of these transgenes by environmental microbes is of critical importance. To investigate the behavior of free transgenic crop DNA, thermophilic anaerobic batch reactors were amended with varying concentrations of transgenic crop genes (i.e., LUG, nptII, and bla) and the persistence of those genes was monitored over 60 days using quantitative PCR. Significant levels of nptII and bla were detected in extracellular DNA (eDNA). Furthermore, LUG maize marker genes were also detected in the control reactors, suggesting that other crop-derived transgenes contained within digested transgenic foods may also enter WWTPs. Possible bacterial transformation events were detected within the highest dose treatments at Days 30 and 60 of incubation. These findings suggest that within the average conventional digester residence times in the United States (30 days), there is a potential for bacterial transformation events to occur with crop-derived transgenes found in eDNA.

The response of transgenic Brassica species to salt stress: a review.

Salt stress is considered one of the main abiotic factors to limit crop growth and productivity by affecting morpho-physiological and biochemical processes. Genetically, a number of salt tolerant Brassica varieties have been developed and introduced, but breeding of such varieties is time consuming. Therefore, current focus is on transgenic technology, which plays an important role in the development of salt tolerant varieties. Various salt tolerant genes have been characterized and incorporated into Brassica. Therefore, such genetic transformation of Brassica species is a significant step for improvement of crops, as well as conferring salt stress resistance qualities to Brassica species. Complete genome sequencing has made the task of genetically transforming Brassica species easier, by identifying desired candidate genes. The present review discusses relevant information about the principles which should be employed to develop transgenic Brassica species, and also will recommend tools for improved tolerance to salinity.

Pyrethroid and carbamate resistance in Australian Helicoverpa armigera (Lepidoptera: Noctuidae) from 2008 to 2015: what has changed since the introduction of Bt cotton?

Pyrethroid and carbamate resistance was evaluated in Helicoverpa armigera from 2008 to 2015. Insects were collected as eggs primarily from cultivated hosts in the major cropping areas of New South Wales and Queensland, Australia. Larvae reared from eggs were tested for resistance to fenvalerate, bifenthrin or methomyl in the F0 generation using a topical application of a discriminating dose of insecticide. In 2008-2009, resistance to fenvalerate was 71% and no resistance to bifenthrin was recorded. In the following two seasons, resistance to pyrethroids was relatively stable with fenvalerate resistance ranging from 63% to 67% and bifenthrin resistance ranging from 5.6% and 6.4% in 2009-2010 and 2010-2011, respectively. However, in 2011-2012, pyrethroid resistance had increased to 91% and 36% for fenvalerate and bifenthrin, respectively. Resistance remained above 90% for fenvalerate and above 35% for bifenthrin in the following three seasons from 2012 to 2015. In 2008-2009, methomyl resistance was 33% and declined to 22% and 15% in 2009-2010 and 2010-2011, respectively. Methomyl resistance remained at moderate levels from 2011-12 to 2014-15, ranging from 21% to 40%. Factors that influenced selection pressure of pyrethroid and carbamate insecticides and impacted resistance frequency in H. armigera may have been associated with changes in the composition of the cropping landscape. The rapid expansion of the pulse industry and the commensurate increased use of insecticide may have played a role in reselection of high-level pyrethroid resistance, and highlights the need for an urgent and strategic response to insecticide resistance management in the Australian grains industry.

The genome of cultivated sweet potato contains Agrobacterium T-DNAs with expressed genes: An example of a naturally transgenic food crop.

Agrobacterium rhizogenes and Agrobacterium tumefaciens are plant pathogenic bacteria capable of transferring DNA fragments [transfer DNA (T-DNA)] bearing functional genes into the host plant genome. This naturally occurring mechanism has been adapted by plant biotechnologists to develop genetically modified crops that today are grown on more than 10% of the world's arable land, although their use can result in considerable controversy. While assembling small interfering RNAs, or siRNAs, of sweet potato plants for metagenomic analysis, sequences homologous to T-DNA sequences from Agrobacterium spp. were discovered. Simple and quantitative PCR, Southern blotting, genome walking, and bacterial artificial chromosome library screening and sequencing unambiguously demonstrated that two different T-DNA regions (IbT-DNA1 and IbT-DNA2) are present in the cultivated sweet potato (Ipomoea batatas [L.] Lam.) genome and that these foreign genes are expressed at detectable levels in different tissues of the sweet potato plant. IbT-DNA1 was found to contain four open reading frames (ORFs) homologous to the tryptophan-2-monooxygenase (iaaM), indole-3-acetamide hydrolase (iaaH), C-protein (C-prot), and agrocinopine synthase (Acs) genes of Agrobacterium spp. IbT-DNA1 was detected in all 291 cultigens examined, but not in close wild relatives. IbT-DNA2 contained at least five ORFs with significant homology to the ORF14, ORF17n, rooting locus (Rol)B/RolC, ORF13, and ORF18/ORF17n genes of A. rhizogenes. IbT-DNA2 was detected in 45 of 217 genotypes that included both cultivated and wild species. Our finding, that sweet potato is naturally transgenic while being a widely and traditionally consumed food crop, could affect the current consumer distrust of the safety of transgenic food crops.

GIS assessment of the risk of gene flow from Brassica napus to its wild relatives in China.

Risk of gene flow from canola (Brassica napus) to species of wild relatives was used as an example to evaluate the risk of gene flow of transgenic crops. B. juncea and B. rapa were the most common weedy Brassica species in China, which were both sexually compatible with canola. Data on canola cultivation in China were collected and analyzed using geographic information system (GIS), and the distribution of its wild relatives was predicted by MaxEnt species distribution model. Based on biological and phenological evidence, our results showed that gene flow risk exists in most parts of the country, especially in places with higher richness of wild Brassica species. However, risk in dominant canola cultivation regions is relatively low owing to the reduced distribution density of wild species in these regions. Three regions of higher risk of gene flow had been identified. Risk of gene flow is relatively high in certain areas. China has been assumed to be the original center of B. juncea and B. rapa, and gene flow may lead to negative effects on the conservation of biodiversity of local species. Strategies had been proposed to reduce the possibility of gene flow either by monitoring introgression from crops to wild relatives in the areas of high adoption of the crop or by taking measures to limit the releasing of new crops or varieties in the areas with abundant wild relatives.

Mechanisms and strategies of plant defense against Botrytis cinerea.

Biotic factors affect plant immune responses and plant resistance to pathogen infections. Despite the considerable progress made over the past two decades in manipulating genes, proteins and their levels from diverse sources, no complete genetic tolerance to environmental stresses has been developed so far in any crops. Plant defense response to pathogens, including Botrytis cinerea, is a complex biological process involving various changes at the biochemical, molecular (i.e. transcriptional) and physiological levels. Once a pathogen is detected, effective plant resistance activates signaling networks through the generation of small signaling molecules and the balance of hormonal signaling pathways to initiate defense mechanisms to the particular pathogen. Recently, studies using Arabidopsis thaliana and crop plants have shown that many genes are involved in plant responses to B. cinerea infection. In this article, we will review our current understanding of mechanisms regulating plant responses to B. cinerea with a particular interest on hormonal regulatory networks involving phytohormones salicylic acid (SA), jasmonic acid (JA), ethylene (ET) and abscisic acid (ABA). We will also highlight some potential gene targets that are promising for improving crop resistance to B. cinerea through genetic engineering and breeding programs. Finally, the role of biological control as a complementary and alternative disease management will be overviewed.

Field-evolved resistance by western corn rootworm to multiple Bacillus thuringiensis toxins in transgenic maize.

The widespread planting of crops genetically engineered to produce insecticidal toxins derived from the bacterium Bacillus thuringiensis (Bt) places intense selective pressure on pest populations to evolve resistance. Western corn rootworm is a key pest of maize, and in continuous maize fields it is often managed through planting of Bt maize. During 2009 and 2010, fields were identified in Iowa in which western corn rootworm imposed severe injury to maize producing Bt toxin Cry3Bb1. Subsequent bioassays revealed Cry3Bb1 resistance in these populations. Here, we report that, during 2011, injury to Bt maize in the field expanded to include mCry3A maize in addition to Cry3Bb1 maize and that laboratory analysis of western corn rootworm from these fields found resistance to Cry3Bb1 and mCry3A and cross-resistance between these toxins. Resistance to Bt maize has persisted in Iowa, with both the number of Bt fields identified with severe root injury and the ability western corn rootworm populations to survive on Cry3Bb1 maize increasing between 2009 and 2011. Additionally, Bt maize targeting western corn rootworm does not produce a high dose of Bt toxin, and the magnitude of resistance associated with feeding injury was less than that seen in a high-dose Bt crop. These first cases of resistance by western corn rootworm highlight the vulnerability of Bt maize to further evolution of resistance from this pest and, more broadly, point to the potential of insects to develop resistance rapidly when Bt crops do not achieve a high dose of Bt toxin.

Genetically transformed tobacco plants expressing synthetic EPSPS gene confer tolerance against glyphosate herbicide.

Glyphosate quashes the synthesis of 5-enolpyruvylshikimate-3- phosphate synthase (EPSPS) enzyme which intercedes the functioning of shikimate pathway for the production of aromatic amino acids. Herbicide resistant crops are developed using glyphosate insensitive EPSPS gene isolated from Agrobacterium sp. strain CP4, which give farmers a sustainable weed control option. Intentions behind this study were to design and characterize the synthetic herbicide resistant CP4-EPSPS gene in a model plant system and check the effectiveness of transformed tobacco against application of glyphosate. Putative transgenic plants were obtained from independent transformation events, and stable plant transformation, transgene expression and integration were demonstrated respectively by PCR, qRT-PCR and Southern hybridization. Gene transcript level and gene copy number (1-4) varied among the tested transgenic tobacco lines. Herbicide assays showed that transgenic plants were resistant to glyphosate after 12 days of spraying with glyphosate, and EPSPS activity remained at sufficient level to withstand the spray at 1000 ppm of the chemical. T1 plants analyzed through immunoblot strips and PCR showed that the gene was being translated into protein and transmitted to the next generation successfully. This codon optimized synthetic CP4-EPSPS gene is functionally equivalent to the gene for glyphosate resistance available in the commercial crops and hence we recommend this gene for transformation into commercial crops.

Genomic variation and DNA repair associated with soybean transgenesis: a comparison to cultivars and mutagenized plants.

BACKGROUND: The safety of mutagenized and genetically transformed plants remains a subject of scrutiny. Data gathered and communicated on the phenotypic and molecular variation induced by gene transfer technologies will provide a scientific-based means to rationally address such concerns. In this study, genomic structural variation (e.g. large deletions and duplications) and single nucleotide polymorphism rates were assessed among a sample of soybean cultivars, fast neutron-derived mutants, and five genetically transformed plants developed through Agrobacterium based transformation methods. RESULTS: On average, the number of genes affected by structural variations in transgenic plants was one order of magnitude less than that of fast neutron mutants and two orders of magnitude less than the rates observed between cultivars. Structural variants in transgenic plants, while rare, occurred adjacent to the transgenes, and at unlinked loci on different chromosomes. DNA repair junctions at both transgenic and unlinked sites were consistent with sequence microhomology across breakpoints. The single nucleotide substitution rates were modest in both fast neutron and transformed plants, exhibiting fewer than 100 substitutions genome-wide, while inter-cultivar comparisons identified over one-million single nucleotide polymorphisms. CONCLUSIONS: Overall, these patterns provide a fresh perspective on the genomic variation associated with high-energy induced mutagenesis and genetically transformed plants. The genetic transformation process infrequently results in novel genetic variation and these rare events are analogous to genetic variants occurring spontaneously, already present in the existing germplasm, or induced through other types of mutagenesis. It remains unclear how broadly these results can be applied to other crops or transformation methods.