GM Crop Database

Database Product Description

T120-7 (ACS-BVØØ1-3)
Host Organism
Beta vulgaris (Sugar Beet)
Trait
Phosphinothricin (PPT) herbicide tolerance, specifically glufosinate ammonium.
Trait Introduction
Agrobacterium tumefaciens-mediated plant transformation.
Proposed Use

Production for human consumption.

Product Developer
Bayer CropScience (Aventis CropScience(AgrEvo))

Summary of Regulatory Approvals

Country Food Feed Environment Notes
Canada 2000 2001 2001
Japan 1999 1999
United States 1998 1998 1998

Introduction Expand

Sugar beet line T120-7 was developed using recombinant DNA techniques to allow for the use of glufosinate ammonium, the active ingredient in phosphinothricin herbicides (Basta®, Ignite®, Rely®, Liberty®, Harvest®, and Finale®) as a weed control option. This line was genetically engineered to express a new enzyme, phosphinothricin-N-acetyltransferase (PAT), derived from the common aerobic soil actinomycete, Streptomyces viridochromogenes.

Glufosinate is a short name for the ammonium salt, glufosinate-ammonium. It is a broad-spectrum contact herbicide and is used to control a wide range of weeds after the crop emerges or for total vegetation control on land not used for cultivation. Glufosinate is a natural compound isolated from two species of Streptomyces fungi. It inhibits the activity of an enzyme, glutamine synthetase, which is necessary for the production of glutamine and for ammonia detoxification. The application of glufosinate leads to reduced glutamine and increased ammonia levels in the plant tissues. This causes photosynthesis to stop and the plant dies within a few days. Glufosinate also inhibits the same enzyme in animals. It is highly biodegradable, has no residual activity, and very low toxicity for humans and wild fauna.

The PAT enzyme detoxifies phosphinothricin via acetylation into an inactive compound. An antibiotic resistance marker gene (neo) encoding the enzyme neomycin phosphotransferase II (NPTII), which inactivates aminoglycoside antibiotics such as kanamycin and neomycin, was also introduced into the genome of line T120-7. This gene was derived from a bacterial transposon (Tn5 transposable element from Escherichia coli) and was included as a selectable marker to identify transformed plants during tissue culture regeneration and multiplication. The expression of the neo gene in these plants has no agronomic significance and the safety of the NPTII enzyme as a food additive was evaluated by the United States Food and Drug Administration in 1994 (US FDA, 1994).

Summary of Introduced Genetic Elements Expand

Code Name Type Promoter, other Terminator Copies Form
pat phosphinothricin N-acetyltransferase HT CaMV 35S CaMV 35S poly(A) signal 1 Synthetic version of native gene
nptII neomycin phosphotransferase II SM nopaline synthase (nos) from A. tumefaciens Native

Characteristics of Beta vulgaris (Sugar Beet) Expand

Center of Origin Reproduction Toxins Allergenicity

Beta is considered an Old World genus basically confined to the Mediterranean Basin and Middle East.

Sugar beet is an outcrossing, largely wind pollinated, plant. During the reproductive phase, large amounts of pollen are produced which can travel long distances. Sugar beet hybridizes freely with all members of the section Beta (within genus Beta).

Saponins (triterpenoid glycosides) are the only known toxicants found in sugar beet and are actively eliminated in sugar processing.

There are rare reports of allergic reactions in sugar beet field workers, associated with exposure to the seeds of the plant.

Donor Organism Characteristics Expand

Latin Name Gene Pathogenicity
Streptomyces viridochromogenes pat

S. viridochromogenes is ubiquitous in the soil. It exhibits very slight antimicrobial activity, is inhibited by streptomycin, and there have been no reports of adverse affects on humans, animals, or plants.

Modification Method Expand

Event T120-7 was produced by Agrobacterium-mediated transformation of calli from the parent line R01 with plasmid vector pOCA18/Ac. The transfer-DNA (T-DNA) portion of the tumour inducing (Ti) bacterial plasmid was engineered to contain a modified form the PAT encoding gene as well as the nptII gene. In order to enhance plant expression of the protein, the nucleotide sequence of the pat gene was modified using site-directed mutagenesis to contain plant-preferred codons. These modifications did not result in changes to the predicted amino acid sequence of the PAT enzyme. Expression of NPTII was used as a selectable marker to screen for transformed plants during tissue culture regeneration and multiplication.

Associated with the pat gene were promoter sequences, as well as transcription termination and polyadenylation signal sequences, derived from the 35S transcript of cauliflower mosaic virus (CaMV). Expression of the NPTII encoding gene was regulated using promoter sequences from the A. tumefaciens nopaline synthase encoding gene (nos).

Characteristics of the Modification Expand

The Introduced DNA

Southern and PCR analyses of genomic DNA from event T120-7 indicated the presence of a single insertion of the T-DNA region, and no evidence of insertion of sequences outside the T-DNA borders.

Genetic Stability of the Introduced Trait

Stable insertion of the T-DNA was demonstrated by comparing the original transformant with four progenies produced either by self-pollination or crosses with nontransgenic sugar beet lines. From Southern blot analyses and trait segregation data, it was concluded that the trait was maintained over multiple generations and inherited in a Mendelian fashion.

Expressed Material

The expression levels of PAT enzyme were quantitated using enzyme linked immunosorbent assay (ELISA) and found to average 137 ng/g (fresh wt tissue; range 74-208 ng/g) in the roots, 966 ng/g (fresh wt tissue; range 732-1176 ng/g) in the tops, and undetectable in the pulp, molasses or refined sugar. The concentrations of NPTII averaged 20 ng/g (fresh wt tissue) in the roots, 44 ng/g (fresh wt tissue) in tops, and undetectable in molasses or refined sugar.

Environmental Safety Considerations Expand

Field Testing

The transgenic sugar beet line T120-7 was field tested in the United States (1994, 1996, 1997) and in Canada, Western and Eastern Europe, and in the former Soviet Union. T120-7 was evaluated extensively and no differences were found in the agronomic characteristics, plant emergence and seedling vigour for line T120-7 compared to non-transformed counterpart beets and standard commercial sugar beet varieties growing in nearby fields.

Outcrossing

Sugar beet is an outcrossing, largely wind pollinated plant. It is normally a biennial, developing a large succulent root in the first year and a seed stalk the second. Certain conditions such as low temperatures after planting and longer day length may induce bolting and produce a seed stalk during the first growing season. Beet is also highly sensitive to frost and a poor competitor with other plants.

During the reproductive phase, large amounts of pollen are produced which can travel long distances. The genus Beta, including the wild relatives, is divided into four sections: Beta, Corolinnae, Procumbentes, and Nanae. Sugar beet hybridizes freely with all members of the section Beta and the resulting progeny are fertile but hybridization is unlikely with other members of the Chenopodiaceae family. Hybrids between sugar beet and members of the other three sections do not naturally occur without human intervention. Assuming proximity, synchrony of flowering and suitable conditions, B. vulgaris may freely hybridize with other varieties.

During production of T120-7, for purposes of than seed production, plants are harvested before the natural onset of the reproductive phase in the same manner that unmodified cultivars are grown. Since it is uncommon for sugar beets to bolt, except in fields or plots grown specifically for seed production, there is little opportunity for uncontrolled pollen flow due to adequate isolation distances enforced by seed certification agencies.

Specific concerns have been raised about potential outcrossing with subsp. macrocarpa, in the Imperial Valley, California. Isozyme studies indicated the introgression of genes from commercial sugar beets has occurred, although other reports show that gene flow between these two plant populations is not likely due to non-synchronous flowering periods, sterile F1 hybrids, and poor growth in F2 hybrids.

It was determined that the transgenic sugar beet line T120-7 was no more likely to become a weed than herbicide tolerant cultivars currently in use or that can be developed by traditional breeding techniques. T120-7 was unlikely to increase the weediness potential of any other cultivated plant or native wild species with which it may interbreed.

Weediness

Sugar beet plants are not a serious weed, although sugar beets have escaped cultivation and their progeny have persisted in the environment for many years. Occasionally, sugar beet volunteers may arise from the presence of wild beet, the bolting of fodder beet plants, the development of groundkeepers, which arise initially from vegetative growth of beet crowns or tops left after harvest or the germination of seed (which may be dormant in the soil for up to 10 years). Volunteer plants may be controlled by mechanical means or the use of registered herbicides that can be used on sugar beet volunteers.

Sugar beet plants have escaped from past commercial cultivation in the San Francisco Bay area and persist to this day. However, transgene movement via pollen to these plants is highly unlikely as sugar beets are no longer in commercial production in the Bay area. Other populations of sexually compatible plants are located in the Imperial Valley of California, but no wild populations exist outside of California.

The movement of the glufosinate tolerance trait from T120-7 to any other sexually compatible plant should not have a significant impact as the glufosinate tolerance would not confer any competitive advantage to these plants, especially if glufosinate is not applied to these plants. This would only occur in managed ecosystems where glufosinate is applied for broad-spectrum weed control, or in plant varieties developed to exhibit glufosinate tolerance and in which glufosinate is used to control weeds. As with glufosinate tolerant sugar beet volunteers, these individuals, should they arise, would be controlled using other available chemical means. Hybrids, if they developed, could potentially result in the loss of glufosinate as a tool to control these species. However, this can be avoided by the use of sound crop management practices including not using the same herbicide every year.

Secondary and Non-Target Adverse Effects

It was concluded that the genes inserted into the transgenic sugar beet line T120-7 would not result in any deleterious effects or significant impacts on nontarget organisms, including threatened and endangered species or beneficial organisms. Field observations of event T120-7 revealed no negative effects on nontarget organisms. The lack of known toxicity for the introduced PAT enzyme suggests no potential for deleterious effects on beneficial organisms such as bees and earthworms. The high specificity of the enzyme for its substrates makes it unlikely that the introduced enzyme would metabolize endogenous substrates to produce compounds toxic to beneficial organisms.

Impact on Biodiversity

Genetically engineered event T120-7 sugar beet is no more likely to become a weed than lines developed by traditional breeding techniques. It is unlikely to increase the weediness potential of any other cultivated plant or native wild species with which it may interbreed. It will not harm threatened and endangered species and non-target organisms. It was concluded that there was no potential impact of event T120-7 on biodiversity.

Food and/or Feed Safety Considerations Expand

Dietary Exposure

Approximately 26% of the world sugar consumed and 47% of the sugar consumed in the United States is produced from sugar beets. Sugar beet is generally converted directly to refined white sugar (which is composed almost entirely of sucrose) through extensive purification processes. The PAT and NPTII proteins were expressed at low levels in the sugar beet roots and tops, and neither protein was detected in refined sugar or molasses derived from sugar beet T120-7, indicating no likelihood for exposure to these proteins from these products.

Sugar beet pulp is a by-product and is normally dried and pelleted for use as a livestock feed. In recent years, it has been purified and used as an additive, at less than 1%, in some specific foods such as food fibre for breakfast cereals etc. The PAT protein was not detected in the pulp and NPTII was detected at very low levels.

Nutritional Data

The transgenic T120-7 line was compared with nontransgenic sugar beets on the basis of numerous compositional components, including crude fat, crude protein, fibre, ash, carbohydrate, calories, calcium, magnesium, phosphorus, potassium, and sodium. With the exception of fibre content, no statistically significant differences were observed. The differences in fibre content were ascribed to environmental factors associated with different growing locations and conditions.

The process fractions, refined sugar, molasses and dried pulp, were assessed for the proximate variables: fatty acids, amino acids, minerals and sugar profiles. Analysis of the dried pulp fraction revealed differences for ash content and some amino acids. It was determined that these differences in nutrient content would not impact on animal nutrition, as dried pulp comprises only a portion, less than 25%, of the total diet for cattle feed use. The molasses fraction revealed differences in fat, protein, calcium, C10:0 and some amino acids. These differences were small and not expected to impact the nutritional content of animal feed. Analysis of the refined sugar fraction showed a difference in tryptophan levels. Importantly, there were no significant differences between transgenic and nontransgenic sugar beets in carbohydrate content, which is the major nutrient in refined sugar (98%).

Toxicity and Allergenicity

It was determined that the PAT enzyme has a very low potential for toxic or allergenic effects based on its physiochemical characteristics (e.g., rapid breakdown under mammalian digestive conditions using simulated gastric and intestinal fluids, and lack of heat stability), its low concentrations in plant tissues and thus food or livestock feed products derived from them, and the lack of amino acid sequence homology with any known protein toxins or allergens.

Abstract Collapse

Sugar beet (Beta vulgaris L. ssp. vulgaris var. altissima) is a botanical variety of B. vulgaris ssp. vulgaris, as are other comestible and fodder beets, and is a member of the goosefoot (Chenopodiaceae) family. It is an important source of sugar (sucrose), accounting for approximately 40 percent of global production.

Sucrose is present in limited quantities in many plants, including various palms and the sugar maple, but sugar beet and sugarcane are the only commercially important sources. More than half of the world sugar supply is obtained from sugarcane, which is grown in tropical and subtropical climates. The rest is supplied by the sugar beet, which is grown in temperate countries. Sugar is manufactured from the roots of the sugar beet; the leaves and tops are removed after harvesting and used as livestock feed. The roots are cut into cossettes, or chips, from which the juice is extracted. The juice is processed to yield sugar and beet molasses, and the remaining pulp is used for domestic animal feed. Beet molasses is also fed to livestock; table molasses is not made from beets because of difficulties in purification. The sugar that is produced from the sugar beet is identical to the sugar that is derived from sugarcane. It is widely used as a sweetener for foods, in preserves, and in the manufacture of candies, baked goods, and soft and alcoholic beverages. Sugar is also used as the raw material for fermentation products such as ethyl alcohol, butyl alcohol, glycerin, citric acid, and levulinic acid. Sugar is an ingredient in some transparent soaps, and it can be converted to esters and ethers, some of which yield tough, insoluble, and infusible resins.

Effective weed management is critical to sugar beet production. Weeds cause significant losses in sugar beet yield and crop quality, and also cause harvesting problems. The low growth habit of the sugar beet plant renders it more susceptible to competition from weeds compared to other crops, such as corn or soybean. Many weed species grow taller than sugar beet and thus aggressively compete for light, water and nutrients. The economic threshold for weed control in sugar beet production is therefore quite low, given the high value of the crop, and the significant losses caused by the presence of weeds. Weeds control strategies include the use of herbicides alone, and herbicides in combination with hand weeding. Sugar beet herbicides can be applied prior to crop emergence (e.g., triallate, EPTC, ethofumesate), and post-emergence (e.g., ethofumesate, triflusulfuron, desmedipham, phenimedipham, sethoxydim).
The sugar beet line T120-7 was genetically engineered to express tolerance to glufosinate ammonium, the active ingredient in phosphinothricin herbicides (Basta®, Rely®, Finale®, and Liberty®). Glufosinate ammonium acts by inhibiting the plant enzyme glutamine synthetase, a key enzyme that detoxifies ammonia by incorporating it into glutamine. Inhibition of this enzyme leads to an accumulation of ammonia in the plant tissues, which kills the plant within hours of application. Phosphinothricin herbicides applied at rates recommended for effective weed control are toxic to conventional sugar beet varieties. The modified T120-7 line permits growers to use phosphinothricin-containing herbicides for weed control in the cultivation of sugar beet.

Glufosinate tolerance in T120-7 sugar beet is the result of the introduction of the pat gene into the beet genome via Agrobacterium-mediated transformation. The pat gene was isolated from the common soil fungus, Streptomyces viridochromogenes, and encodes the enzyme phosphinothricin-N-acetyltransferase (PAT). PAT catalyses the acetylation of phosphinothricin which detoxifies it into an inactive compound.

The transgenic sugar beet line T120-7 was field tested in the United States from 1994 to 1997, as well as in Canada, Western and Eastern Europe, and in the former Soviet Union. T120-7 was evaluated extensively and no differences were found in agronomic characteristics, plant emergence and seedling vigour when compared to standard commercial sugar beet varieties growing in nearby fields. It was demonstrated that the transformed sugar beet line did not negatively affect beneficial or non-target organisms, and it was not expected to impact on threatened or endangered species.

Sugar beet is an outcrossing species, and is mainly wind pollinated. It is a biennial, developing a rosette of leaves and a large taproot in the first year, and bolting (i.e., forming an inflorescence bearing stem) in the second year. Bolting is induced only after the plant has undergone vernalization at the end of the first season. Certain environmental conditions, such as low temperatures and longer daylength, may induce bolting in the first year. Sugar beet is also very sensitive to frost, and is a poor competitor with other plants. Large amounts of pollen are produced during the reproductive phase, which is carried by wind over long distances.

The genus Beta is comprised of four sections: i) Beta, to which sugar beet belongs; ii) Corollinae; iii) Nanae; and iv) Procumbentes. All species within the Beta section (e.g., B. vulgaris ssp. maritima, B. macrocarpa) are cross-compatible and produce fertile progeny. Hybrids between sugar beet and members of the other three sections do not occur naturally, and cross-breeding with other members of the Chenopodiaceae is unlikely. As with conventional sugar beet cultivars, T120-7 plants are harvested prior to the initiation of the bolting phase. Since sugar beets are usually not allowed to bolt, except in fields or plots grown for seed production, there is little opportunity for uncontrolled pollen flow. Sugar beets grown for seed production are also isolated from other sugar beet fields to minimize pollen flow. This is usually acheived by the use of isolation distances enforced by seed certification agencies.

The development of glufosinate-tolerant hybrids is possible due to the ability of sugar beet to outcross with related plants. However, the glufosinate-tolerance trait is not expected to provide a competitive advantage to hybrid plants unless grown in managed environments routinely subjected to glufosinate applications. In the event that a glufosinate-tolerant hybrid survived, the herbicide-tolerant individual could be controlled using mechanical and/or other available chemical means. T120-7 is considered unlikely to increase the weediness potential of any other cultivated plant or native wild species with which it may interbreed. T120-7 also does not possess growth and reproductive characteristics that would render it weedier than conventional sugar beet.

Sugar beet intended for human consumption is generally converted directly to refined white sugar through extensive purification processes. The PAT protein in T120-7 sugar beet was expressed at low levels in the sugar beet roots and tops, and was not detected in refined sugar or molasses derived from T120-7. Consequently there will be no human exposure to PAT protein as a result of consumption of refined sugar derived from T120-7 sugar beet.

Sugar beet pulp is a by-product of the refining process and is normally dried and pelleted for use as a livestock feed. In recent years, it has been purified and used as an additive, at levels of less than 1%, in some specific foods, such as food fibre for breakfast cereals. The PAT protein was not detected in the pulp produced from T120-7 sugar beet.

T120-7 line was compared with non-transgenic sugar beets in analyses of numerous compositional components, including crude fat, crude protein, fibre, ash, carbohydrate, calories, calcium, magnesium, phosphorus, potassium, and sodium. With the exception of fibre content, no statistically significant differences were observed. The differences in fibre content were ascribed to environmental factors associated with different growing locations and conditions. The process fractions, refined sugar, molasses and dried pulp produced from T120-7 sugar beet were assessed by proximate analysis (fatty acids, amino acids, minerals and sugar profiles). The dried pulp fraction revealed differences in ash content and some amino acids. It was determined that these differences in nutrient content would not impact on animal nutrition, as dried pulp comprises less than 25% of the content of livestock feed. The molasses fraction revealed differences in fat, protein, calcium, and some amino acids. These differences were small and not expected to impact the nutritional content of animal feed. Analysis of the refined sugar fraction showed a difference in tryptophan levels. Importantly, there were no significant differences in carbohydrate content, which is the major nutrient in refined sugar (98%).

Potential toxicity and allergenicity of the transformed sugar beet line was investigated through examination of the amino acid sequence and physiochemical characteristics of the introduced PAT protein. It was determined that the PAT enzyme has a very low potential for toxic or allergenic effects. No significant homologies with the amino acid sequences of known protein toxins or allergens were detected, the PAT enzyme did not display the heat stability characteristic of allergenic compounds, and was rapidly degraded under conditions simulating mammalian digestion. The low concentrations of the PAT enzyme found in T120-7 tissues, and so in the food or livestock feed products derived from them, further supported the conclusion that the PAT enzyme possessed little or no potential for allergenicity or toxicity.

Links to Further Information Expand


This record was last modified on Friday, March 26, 2010