CP4 EPSPS Gene Research Articles

Effects of transgenic modification on the bacterial communities in different niches of maize under glyphosate toxicity

Transgenic glyphosate-resistant maize has emerged as a way to expand the use of glyphosate for weed control. Studying the microbiome in the tissues and rhizosphere soil of transgenic plants is vital for understanding the glyphosate-resistant mechanism and optimizing the transgenic design of crops. In our study, the expression of a mutant cp4epsps gene in transgenic maize, which confers tolerance to glyphosate, was performed using the maize variety Xianyu 335 as the genetically modified acceptor line. This transgenic modification did not affect the initial bacterial community in the leaf, stem, or root of maize, but promoted a differential bacterial community in the rhizosphere soil. Under glyphosate application, the abundance of beneficial bacteria involved in N fixation and P solubilization in plant tissues and the rhizosphere soil of glyphosate-resistance maize were higher than those in the glyphosate-sensitive maize. In contrast, the abundance of pathogens had the opposite trend, suggesting that the enhanced health of transgenic maize prevented microbiome deterioration under glyphosate. The re-inoculation of bacterial strains isolated from glyphosate-resistance maize into the leaf and rhizosphere soil of glyphosate-sensitive maize resulted in an enhanced photosynthetic capacity in response to glyphosate, demonstrating the vital role of specific bacteria for glyphosate resistance. Our study provides important evidence of how transgenic maize tolerance to herbicides affects the bacterial communities across the maize niches under glyphosate toxicity.

Assessment of the potential risks in SD rats gavaged with genetically modified yeast containing the cp4-epsps gene.

Despite the absence of definitive evidence indicating that the cp4-epsps gene and its resultant recombinant proteins have significant harmful effects on either human or animal health, the safety assessment of genetically modified (GM) crops expressing the CP4-EPSPS proteins has been controversial. This study endeavor was aimed at evaluating the potential risks posed by the CP4-EPSPS protein in transgenic crops, thereby contributing to the advancement of risk assessment methodologies in the context of genetically engineered crops. To ascertain the appropriate daily dosages for oral gavage administration, the expression levels of the CP4-EPSPS protein in a recombinant yeast were quantified. Subsequently, physiological and biochemical analysis, metabolomics, and metagenomic analysis were conducted based on a 90-day Sprague-Dawley (SD) rats feeding experiment, respectively, thereby enhancing the depth and precision of our risk assessment framework. The results from the physiological and biochemical analysis, organ pathological, blood metabolism, gut microbiota, and correlation analysis of metabolites and gut microbiota revealed several biomarkers for further risk assessment. These biomarkers include clinical biochemical indexes such as total bilirubin (TBIL), direct bilirubin (DBIL), creatine kinase (CK), and lactate dehydrogenase (LDH); metabolites like Methionine, 2-Oxovaleric acid, and LysoPC (16:0); and gut microbiota including Blautia wexlerae, Holdemanella biformis, Dorea sp. CAG 317, Coriobacteriaceae and Erysipelotrichaceae. In conclusion, the risk can be significantly reduced by directly consuming inactivated recombinant CP4-EPSPS. Therefore, in everyday life, the risk associated with consuming GM foods containing recombinant CP4-EPSPS is substantially reduced after heat treatment.

Establishment of an Efficient Sugarcane Transformation System via Herbicide-Resistant CP4-EPSPS Gene Selection

Sugarcane (Saccharum spp.), a major cash crop that is an important source of sugar and bioethanol, is strongly influenced by the impacts of biotic and abiotic stresses. The intricate polyploid and aneuploid genome of sugarcane has shown various limits for conventional breeding strategies. Nonetheless, biotechnological engineering currently offers the best chance of introducing commercially significant agronomic features. In this study, an efficient Agrobacterium-mediated transformation system that uses the herbicide-resistant CP4-EPSPS gene as a selection marker was developed. Notably, all of the plants that were identified by PCR as transformants showed significant herbicide resistance. Additionally, this transformation protocol also highlighted: (i) the high yield of transgenic lines from calli (each gram of calli generated six transgenic lines); (ii) improved selection; and (iii) a higher transformation efficiency. This protocol provides a reliable tool for a routine procedure for the generation of resilient sugarcane plants.

Effect of glyphosate management, formulations and rates on the agronomic performance of maize with the cp4epsps gene

Effect of glyphosate management, formulations and rates on the agronomic performance of maize with the cp4epsps gene

From genes to fields: Environmental compatibility of herbicide tolerant transgenic cotton

IntroductionThere are several enticing reasons to use genetic engineering to develop transgenic cotton plants with economically relevant new traits. The use of transgenic cotton to protect crops from biotic stressors like weeds may prove to be a cost-effective solution. ObjectivesThe development of herbicide-resistant cotton variants is the primary motivation for gene transfer in this crop. In the current study, we conducted a battery of experiments on the herbicide-tolerant cotton variety MNH-786 to investigate its expression patterns and environmental adaptation. Material and methodsThe 35S promoter controls the expression of the codon-optimized Cp4-EPSPS gene that is present in this variety. Several different molecular approaches, such as polymerase chain reaction (PCR), Florescent In-Situ Hybridization analysis, and enzyme-linked immunosorbent assay (ELISA), are used to confirm the presence of transgenic cotton plants. The glyphosate spray assay was performed using molecular analysis, employing a dosage of 2000 ml per acre. The suitability of transgenic and non-transgenic cotton plants in four unique environments—the Multan, Vehari, Bahawalpur, and Faisalabad Districts of Punjab province, Pakistan was assessed using a comparative comparison. A gene x environment interaction (GEI) biplot was used to analyze the correlation between genes and their environments. ResultsTransgenic cotton plants screened using PCR to target 1.467 kb regions showed effective integration into cotton genomes. Results showed that Cp4-EPSPS proteins fluctuated and associated with environmental variables across all three generations (T0, T1, and T2). In the herbicide spray assay, all transgenic cotton plants survived, but non-transgenic cotton and all weeds died. According to the "which-won-where" polygon, transgenic cotton plant Events 3, 4, 5, 6, and 7 performed better in three settings, except Faisalabad. According to correlation and heat-map yield research, transgenic cotton variety MNH-786 Events 4, 5 and 6 have a higher consistency and influence in Vehari district's natural habitats. Multan and Bahawalpur also show encouraging results. In Faisalabad, no transgenic events perform well. ConclusionWe conclude that Events 4 and 5 show a substantial abundance of genetic potential that is robust over a wide range of environments thanks to our thorough study of the outcomes obtained under these conditions. SignificanceOur study emphasises the importance of transgenic cotton, its performance, and its exposure to appropriate agroecological zones. This knowledge can be advantageous for farmers in attaining optimal conditions to maximize productivity.

Single primer isothermal amplification for real-time and visual on-site detection of genetically modified crops by recognizing CP4-EPSPS gene

In response to the potential controversies on increasing circulation of genetically modified organisms (GMOs) in the global markets, the rapid and sensitive detection methods are essential for monitoring of GMOs to maintain consumers' rights. Herein, we first developed a rapid technique for the real-time and visual on-site detection of the genetically modified crops (specifically targeting CP4-EPSPS gene) by using single primer isothermal amplification (SPIA) combined with SYBR Green II. 19 GM crop events and 12 non-GM crop events were used to assess the specificity of the developed SPIA assay, which presented excellent specificity. The limit of detection of the developed SPIA method was as low as 4 copies/μL, which was better than that of the real-time PCR method (20 copies/μL). The SPIA method was further used to assess 30 of actual samples, and the experimental results revealed that 3 positive samples were found, which were in good agreement with those of the real-time PCR methods. Overall, the major advantages of the developed SPIA over real-time PCR are the lower LOD, simplicity, and inexpensiveness. The developed SPIA method provides a potential alternative for the real-time and visual on-site detection of GMOs, which also accelerates the development of highly sensitive nucleic acid testing techniques.

An integrated strategy involving high-throughput sequencing to characterize an unknown GM wheat event in Canada.

Glyphosate-resistant wheat plants were discovered in southern Alberta in 2017, representing an unauthorized GM release in Canada. The Canadian Food Inspection Agency undertook a series of experiments to characterize and identify this unknown GM wheat, as well as to develop and validate construct-specific and event-specific qPCR assays. Results of PCR-based assays and Sanger sequencing indicated the presence of CaMV 35S promoter (p35S), Rice Actin 1 intron (RactInt1), CP4-EPSPS gene and nopaline synthase terminator (tNOS) elements in the unknown GM wheat. Genome walking and bead capture strategies, combined with high-throughput sequencing, were used to identify the 5' and 3' wheat junctions and the subsequent mapping of the insert to chromosome 3B of the wheat genome. A probable transformation vector, pMON25497, was recognized, and further testing identified the unknown GM wheat as MON71200 event, one of two events obtained with the pMON25497 vector. The two construct-specific assays targeted the junctions of the RactInt1 and the CP4-EPSPS elements and the CP4-EPSPS and tNOS elements, while the event-specific assay was located at the 3' junction into the wheat genome. Both construct-specific and event-specific assays had limits of detection of 0.10% of MON71200 in a seed pool. As expected, the two construct-specific assays cross-reacted with other wheat and corn events containing the same elements in the same order. No cross-reactivity was observed for the event-specific assay. The integrated strategy employed in this study can serve as a model for other cases when facing similar challenges involving unknown GM events.

Paternal chromosome elimination of inducer triggers induction of double haploids in Brassica napus

A synthetic octoploid rapeseed, Y3380, induces maternal doubled haploids when used as a pollen donor to pollinate plant. However, the mechanism underlying doubled haploid formation remains elusive. We speculated that double haploid induction occurs as the inducer line’s chromosomes pass to the maternal egg cell, and the zygote is formed through fertilization. In the process of zygotic mitosis, the paternal chromosome is specifically eliminated. Part of the paternal gene might have infiltrated the maternal genome through homologous exchange during the elimination process. Then, the zygote haploid genome doubles (early haploid doubling, EH phenomenon), and the doubled zygote continues to develop into a complete embryo, finally forming doubled haploid offspring. To test our hypothesis, in the current study, the octoploid Y3380 line was back bred with the 4122-cp4-EPSPS exogenous gene used as a marker into hexaploid Y3380-cp4-EPSPS as paternal material to pollinate three different maternal materials. The fertilization process of crossing between the inducer line and the maternal parent was observed 48 h after pollination, and the fertilization rate reached 97.92% and 98.72%. After 12 d of pollination, the presence of cp4-EPSPS in the embryo was detected by in situ PCR, and at 13–23 d after pollination, the probability of F1 embryos containing cp4-EPSPS gene was up to 97.27%, but then declined gradually to 0% at 23–33 d. At the same time, the expression of cp4-EPSPS was observed by immunofluorescence in the 3rd to 29th day embryo. As the embryos developed, cp4-EPSPS marker genes were constantly lost, accompanied by embryonic death. After 30 d, the presence of cp4-EPSPS was not detected in surviving embryos. Meanwhile, SNP detection of induced offspring confirmed the existence of double haploids, further indicating that the induction process was caused by the loss of specificity of the paternal chromosome. The tetraploid-induced offspring showed infiltration of the induced line gene loci, with heterozygosity and homozygosity. Results indicated that the induced line chromosomes were eliminated during embryonic development, and the maternal haploid chromosomes were synchronously doubled in the embryo. These findings support our hypothesis and lay a theoretical foundation for further localization or cloning of functional genes involved in double haploid induction in rapeseed.

A 90-day feeding study of genetically modified maize LP007-1 in wistar han RCC rats.

LP007-1 is a variety of insect-resistant and herbicide-tolerant maize containing the modified cry1Ab, cry2Ab, vip3Aa and cp4-epsps genes. The food safety assessment of the maize LP007-1 was conducted in Wistar Han RCC rats by a 90-days feeding study. Maize grains from both LP007-1 or its corresponding non-genetically modified control maize AX808 were incorporated into rodent diets at 25% and 50% concentrations by mass and administered to rats (n=10/sex/group) for 90 days. A commercialized rodent diet was fed to an additional group as the basal-diet group. The diets of all groups were nutritionally balanced. No biologically relevant differences were observed in rats fed with maize LP007-1 compared to rats fed with AX808 and the basal-diet with respect to body weight/gain, food consumption/utilization, clinical signs, mortality, ophthalmology, clinical pathology (hematology, prothrombin time, activation of partial thrombin time, serum chemistry, urinalysis), organ weights, and gross and microscopic pathology. Considering the circumstances of this study, the results provided evidence that LP007-1 maize did not exhibit toxicity in the 90-day feeding study.

Glyphosate in agronomic performance and seed quality of soybean with cp4-EPSPs and cry1Ac genes

Glyphosate in agronomic performance and seed quality of soybean with cp4-EPSPs and cry1Ac genes

Survivorship and food consumption of immatures and adults of Apis mellifera and Scaptotrigona bipunctata exposed to genetically modified eucalyptus pollen

Eucalyptus comprises the largest planted area of cultivated production forest in Brazil. Genetic modification (GM) of eucalyptus can provide additional characteristics for increasing productivity and protecting wood yield, as well as potentially altering fiber for a diversity of industrial uses. However, prior to releasing a new GM plant, risk assessments studies with non-target organisms must be undertaken. Bees are prominent biological models since they play an important role in varied ecosystems, including for Eucalyptus pollination. The main goal of this study was to evaluate whether a novel event (Eucalyptus 751K032), which carries the cp4-epsps gene that encodes the protein CP4-EPSPS and nptII gene that encodes the protein NPTII, might adversely affect honey bees (Apis mellifera) and stingless bees (Scaptotrigona bipunctata). The experiments were performed in southern Brazil, as follows: (i) larvae and adults were separately investigated, (ii) three or four different pollen diets were offered to bees, depending on larval or adult status, and (iii) two biological attributes, i.e., survivorship of larvae and adults and food intake by adults were evaluated. The diets were prepared with pollen from GM Eucalyptus 751K032; pollen from conventional Eucalyptus clone FGN-K, multifloral pollen or pure larval food. The insecticide dimethoate was used to evaluate the sensitivity of bees to toxic substances. Datasets were analyzed with Chi-square test, survival curves and repeated measures ANOVA. Results indicated no evidence of adverse effects of Eucalyptus pollen 751K032 on either honey bees or stingless bees assessed here. Therefore, the main findings suggest that the novel event may be considered harmless to these organisms since neither survivorship nor food consumption by bees were affected by it.

Detection of Genetically Modified Glyphosate-Resistant Soybean Sold in Sarawak

A genetically modified (GM) crop is an organism whose genetic makeup has been altered to express the desired physiological traits. Soybean (Glycine max) is a common GM crop. Its genome has been genetically engineered to confer resistance to herbicides, pests and extreme environmental conditions. Mislabelling of food products as GM-free has triggered insecurity among consumers. In addition, the health effects due to consumption of GM foods remains controversial. Therefore, this study aimed to identify the presence of GM soybean in animal feeds and several food products such as raw soybean, tempeh, and tofu collected from Sarawak traditional markets, grocery stores, and supermarkets. The presence of the regulatory elements CaMV 35S Promoter (P35S) and NOS Terminator (TNOS) were initially screened using conventional Polymerase Chain Reaction (PCR). Then, all samples were subjected to the PCR-based construct-specific method by targeting the cp4 epsps gene, which confers glyphosate-resistance. Positive samples were validated through DNA sequencing. The result demonstrated that 56 out of 65 samples including 17 soybean, 12 animal feeds, 7 tofu and 20 tempeh samples were positive for cp4 epsps. Furthermore, 2 out of 20 raw soybean samples were labelled as GM-free. However, validation using DNA sequencing indicates 100% identity to cp4 epsps gene in comparison with the Genbank database. This study demonstrated the significance of GM detection in soybean and the importance of accurate food labelling.

Stacking herbicide detoxification and resistant genes improves glyphosate tolerance and reduces phytotoxicity in tobacco (Nicotiana tabacum L.) and rice (Oryza sativa L.)

Glyphosate residues retained in the growing meristematic tissues or in grains of glyphosate-resistant crops affect the plants physiological functions and crop yield. Removing glyphosate residues in the plants is desirable with no penalty on crop yield and quality. We report a new combination of scientific strategy to detoxify glyphosate that reduces the residual levels and improve crop resistance. The glyphosate detoxifying enzymes Aldo-keto reductase (AKR1) and mutated glycine oxidase (mGO) with different modes of action were co-expressed with modified EPSPS, which is insensitive to glyphosate in tobacco (Nicotiana tabacum L.) and rice (Oryza sativa L.). The transgenic tobacco plants expressing individual PsAKR1, mGO, CP4-EPSPS, combinations of PsAKR1:CP4EPSPS, PsAKR1:mGO, and multigene with PsAKR1: mGO: CP4EPSPS genes were developed. The bio-efficacy studies of in-vitro leaf regeneration on different concentrations of glyphosate, seedling bioassay, and spray on transgenic tobacco plants demonstrate that glyphosate detoxification with enhanced resistance. Comparative analysis of the transgenic tobacco plants reveals that double and multigene expressing transgenics had reduced accumulation of shikimic acid, glyphosate, and its primary residue AMPA, and increased levels of sarcosine were observed in all PsAKR1 expressing transgenics. The multigene expressing rice transgenics showed improved glyphosate resistance with yield maintenance. In summary, results suggest that stacking genes with two different detoxification mechanisms and insensitive EPSPS is a potential approach for developing glyphosate-resistant plants with less residual content.

Compositional and Animal Feeding Assessments of a Novel Herbicide-Tolerant Maize Variety

ZDAX5 is a variety of herbicide-tolerant maize that contains the modified P450-N-Z1 gene isolated from Cynodon dactylon (L.) Pers. and the cp4 epsps gene isolated from the Agrobacterium tumefaciens strain CP4 and exhibits high tolerances to flazasulfuron and glyphosate under field conditions. Once ZDAX5 corn is available on the market, the evolution of herbicide-resistant weeds will be delayed by applying glyphosate and flazasulfuron to corn fields. Prior to commercialization, it is critical to assess the safety of ZDAX5 maize. Compositional analysis and feed consumption studies in rodents are an important consideration in the safety assessment of genetically modified crops. The nutritional components of ZDAX5 were analyzed and compared with those of its non-transgenic counterpart. The data showed that all the analyzed components in the herbicide-tolerant maize plants were substantially equivalent to those of its non-transgenic counterpart. Furthermore, most of the measured values from ZDAX5 were within the range of values reported for other commercial maize varieties. The sub-chronic feeding trial was carried out with grains from GM, and non-GM maize were independently added into rodent diets at concentrations of 12.5%, 25% and 50%. As a control, another set of rats was fed with a marketed diet. At the end of the 90-day feeding study, no negative effects associated with the consumption of GM maize were found. These results indicate that the herbicide-tolerant maize ZDAX5 is as nutritious and safe as non-transgenic maize.

Engineered resistance and risk assessment associated with insecticidal and weeds resistant transgenic cotton using wister rat model

Stacking multiple genes into cotton crop to cop up multiple biotic stresses such as insects and weeds is a promising tool to save crop from losses. Transgenic cotton variety, VH-289, with double Bt and cp4EPSPS genes under the control of 35S promoter was used for the expression analyses and biosafety studies. The transgenic cotton plants were screened through PCR amplification of fragments, 1.7 kb for Cry1Ac, 582 bp for Cry2A and 250 bp for cp4EPSPS; which confirmed the presence of all genes transformed in transgenic cotton. The Cry1Ac + Cry2A and cp4EPSPS proteins were quantified through ELISA in transgenic cotton plants. The Glyphosate assay performed by spraying 1900 mL per acre of glyphosate Roundup further confirmed complete survival of transgenic cotton plants as compared to the non-transgenic cotton plants and all weeds. Similarly, insect infestation data determined that almost 99% insect mortality was observed in controlled field grown transgenic cotton plants as compared to the non-transgenic control plants. Evaluation of effect of temperature and soil nutrients availability on transgene expression in cotton plants was done at two different cotton growing regions, Multan and Lahore, Pakistan and results suggested that despite of higher temperature in Multan field, an increased level of Cry and cp4EPSPS proteins was recorded due to higher soil organic matter availability compared to Lahore field. Before commercialization of any transgenic variety its biosafety study is mandatory so, a 90 days biosafety study of the transgenic cotton plants with 40% transgenic cottonseeds in standard diet showed no harmful effect on wister rat model when studied for liver function, renal function and serum electrolyte.

Rapid detection of genetically modified products based on CRISPR-Cas12a combined with recombinase polymerase amplification

With the large-scale planting of genetically modified (GM) crops, consumers were more aware of biosafety. Onsite rapid diagnostic methods were advantageous to the regulation of GM products. In this study, a rapid, sensitive and portable detection method based on recombinase polymerase amplification were proposed based on RPA reaction and Cas12a cleavage reaction for GM ingredients, named RPA-Cas12a-GM. The results would be displayed by fluorescence signal (FS) and visual bands of lateral flow strip (LFS). RPA-Cas12a-GM method could be completed within 45 min, and the detection limit was as low as 45 copies/μL of the standard plasmid containing CP4-EPSPS gene and Cry1Ab/Ac gene. Furthermore, the detection coincidence rate of RPA-Cas12a-GM method was 100%. In conclusion, the proposed RPA-Cas12a-GM method based FS and LFS were sensitive, specific, rapid and visible for diagnosis of CP4-EPSPS gene and Cry1Ab/Ac gene without complex equipment, which provides technical support for the regulation of GM products in the field.

The Naturally Evolved EPSPS From Goosegrass Confers High Glyphosate Resistance to Rice.

Glyphosate-resistant crops developed by the CP4-EPSPS gene from Agrobacterium have been planted on a massive scale globally, which benefits from the high efficiency and broad spectrum of glyphosate in weed control. Some glyphosate-resistant (GR) genes from microbes have been reported, which might raise biosafety concerns. Most of them were obtained through a hygromycin-HPT transformation system. Here we reported the plant source with 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) gene from goosegrass endowed rice with high resistance to glyphosate. The integrations and inheritability of the transgenes in the rice genome were investigated within two generations. The EiEPSPS transgenic plants displayed similar growth and development to wild type under no glyphosate selection pressure but better reproductive performance under lower glyphosate selection pressure. Furthermore, we reconstructed a binary vector pCEiEPSPS and established the whole stage glyphosate selection using the vector. The Glyphosate-pCEiEPSPS selection system showed a significantly higher transformation efficiency compared with the hygromycin-HPT transformation system. Our results provided a promising alternative gene resource to the development of GR plants and also extended the plant transformation toolbox.

Ionomics and lipidomics for evaluating the transgenic (cp4-EPSPS gene) and non-transgenic soybean seed generations

This work evaluates three generations of transgenic and non-transgenic soybean plants through multiomics namely ionomics and lipidomics. Transgenic variety was inserted with cp4-EPSPS gene, which confers resistance to glyphosate. Comparisons between soybeans were carried out from seeds obtained during summer season on south hemisphere, grown in the same soil and without intentional stress factors. Essential and non-essential elements, B, Al, Mn, Fe, Co, Ni, Cu, Zn, Sr, Mo, and Ba, and fatty acids were evaluated by inductively coupled plasma mass spectrometry (ICP-MS) and nuclear magnetic resonance (NMR), respectively. After checking the accuracy of the method employed, chemometric analysis (using Principal Component Analysis, Hierarchical CusterAnalysis, and correlation heatmaps) indicated that some elements may share mechanisms of transport and are to some extent, correlating with fatty acids, allowing for differentiation of all generations and soybean species. Results reforce the importance of multiomics approach to evaluate genetic modified organism such as plants.

Final Health and Environmental Risk Assessment of Genetically Modified Soybean MON 89788

Soybean MON 89788 expresses the cp4 epsps gene from the plant pathogenic bacterium Agrobacterium tumefaciens (Rhizobium radiobacter) sp. strain CP4. The encoded enzyme 5enolpyruvylshikimate-3-phosphate synthase (CP4 EPSPS) protein confers tolerance to the active herbicidal substance glyphosate. Updated bioinformatics analyses of the inserted DNA and flanking sequences in soybean MON 89788 have not indicated a potential production of putative harmful proteins or polypeptides caused by the genetic modification. Genomic stability of the functional insert and consistent expression of the cp4 epsps gene, have been shown over several generations of soybean MON 89788. With the exception of the intended changes caused by the trans-genetically introduced trait, data from several field trials performed in USA and Argentina show that soybean MON 89788 is compositionally, morphologically and agronomically equivalent to its conventional counterpart and other commercial soybean varieties. A sub-chronic feeding study with rats, as well as a nutritional assessment trial with broilers has not revealed adverse effects of soybean MON 89788. These studies indicate that soybean MON 89788 is nutritionally equivalent to, and as safe as conventional soybean varieties. The CP4 EPSPS protein produced in soybean MON 89788 does not show sequence resemblance to known toxins or IgE-dependent allergens, nor has it been reported to cause IgE-mediated allergic reactions. Soybean is not cultivated in Norway, and there are no cross-compatible wild or weedy relatives of soybean in Europe. 
 
 Based on current knowledge, the VKM GMO Panel concludes that with the intended usage, there are no discernible safety concerns associated with soybean MON 89788 regarding human or animal health or to the environment in Norway.

Ongoing ecological and evolutionary consequences by the presence of transgenes in a wild cotton population

After 25 years of genetically modified cotton cultivation in Mexico, gene flow between transgenic individuals and their wild relatives represents an opportunity for analysing the impacts of the presence of novel genes in ecological and evolutionary processes in natural conditions. We show comprehensive empirical evidence on the physiological, metabolic, and ecological effects of transgene introgression in wild cotton, Gossypium hirsutum. We report that the expression of both the cry and cp4-epsps genes in wild cotton under natural conditions altered extrafloral nectar inducibility and thus, its association with different ant species: the dominance of the defensive species Camponotus planatus in Bt plants, the presence of cp4-epsps without defence role of Monomorium ebeninum ants, and of the invasive species Paratrechina longicornis in wild plants without transgenes. Moreover, we found an increase in herbivore damage to cp4-epsps plants. Our results reveal the influence of transgene expression on native ecological interactions. These findings can be useful in the design of risk assessment methodologies for genetically modified organisms and the in situ conservation of G. hirsutum metapopulations.