GM Crop Database

Database Product Description

X81359
Host Organism
Helianthus annuus (Sunflower)
Trade Name
CLEARFIELD™
Trait
Herbicide tolerant, imidazolinone.
Trait Introduction
Selection for a naturally occurring mutation
Proposed Use

Production of oil for use in human food only.

Product Developer
BASF Inc.

Summary of Regulatory Approvals

Country Food Feed Environment Notes
Canada 2003 2005 2005

Introduction Expand

X81359 (Clearfield™) sunflower hybrid was developed to allow the use of imidazolinone herbicides as a post-emergence weed control option in sunflower production. The tolerance to imidazolinone herbicides in X81359 is due to a naturally occurring mutation in the AHAS gene discovered in a wild population of Helianthus annus. This trait was introduced into X81359 using conventional plant breeding techniques.

AHAS catalyses the first step in the biosynthesis of the branched-chain amino acids isoleucine, leucine, and valine, and is active in the glycolytic pathway of plant metabolism. When conventional sunflower plants are treated with an imidazolinone, the herbicide binds to a specific site on the AHAS enzyme, thereby inhibiting its activity. The result of this enzyme inhibition is a decrease in the synthesis of these amino acids, and an accumulation of toxic levels of alpha-ketoglutarate, all of which results in the eventual death of the plant.

Summary of Introduced Genetic Elements Expand

Code Name Type Promoter, other Terminator Copies Form
als acetolactate synthase MUT Imidazolinone selection for a naturally occurring mutant from wild sunflower populations.

Characteristics of Helianthus annuus (Sunflower) Expand

Center of Origin Reproduction Toxins Allergenicity
North America Primarily self-pollinated. Some outcrossing via insect-pollination. Antinutritional compounds phytic acid and trypsin inhibitor. In seeds, several IgE binding proteins (10-50 kD) have been identified, including high-methionine 2S albumin.

Modification Method Expand

The imidazolinone-tolerance trait in X81359 was discovered in a wild (weedy) population of sunflower (Helianthus annus). The natural mutation conferring the imidazolinone tolerance was detected in a field where soybeans has been cultivated and treated with imazethapyr for several seasons. The mutant sunflower population expressed a mutation in AHAS gene, conferring tolerance to applications of imazethapyr. Conventional plant breeding techniques were used to introduce the herbicide-tolerance trait into sunflower germplasm, including extensive backcrossing into NuSun sunflower varieties.

Characteristics of the Modification Expand

The Introduced DNA

Since the imidazolinone-tolerance trait in X8159 hybrid sunflower is from a natural point mutation of wild sunflower, there was no introduction or incorporation of heterologous DNA into the plant genome. The tolerance to imidazolinone is due to a single point mutation in the AHAS gene, such that the herbicide cannot inhibit the enzyme’s activity. Information on the sequence of the modified AHAS gene revealed the single point mutation.

Genetic Stability of the Introduced Trait

The results of segregation analyses using the wild population from which the imidazolinone trait was isolated demonstrate the inheritance of a single semi-dominant allele. The hybrid X81359, which is several generations removed from the original mutation, is consistently tolerant to imidazolinone herbicides (CFIA, 2005b).

Expressed material

The modified AHAS gene, conferring tolerance to imidazolinone herbicides, is under control of the native AHAS promoter and is believed to be constitutively expressed.

In the unmodified plant, the levels of valine, leucine and isoleucine are regulated by feedback inhibition of AHAS. A mutation in the AHAS enzyme could affect the regulation of the biosynthesis of these amino acids. Data were submitted to show that the modified AHAS was feedback inhibited by valine and leucine, and thus, similar to the unmodified enzyme. Levels of valine, leucine and isoleucine in X81359 sunflower were comparable to those in the unmodified plant.

Environmental Safety Considerations Expand

Only the meal and oil of X81359 hybrid sunflower will be imported into Canada since this hybrid is intended solely for use as human food and livestock feed. X81350 sunflower is not intended for cultivation in Canada. No environmental effects are therefore expected from the importation of X81359 sunflower into Canada.

Food and/or Feed Safety Considerations Expand

Dietary exposure

The modification to the AHAS enzyme in X81359 will not result in any change in the consumption or use pattern of sunflower oil and products derived from the oil and the meal. The oil and meal from X81359 are therefore expected to be used in applications similar to other sunflower and vegetable oils by the food and feed industry. X81359 is also the result of several backcrosses into NuSun sunflower lines. The oil from these sunflower lines have an altered fatty acid profile, and are termed “mid-oleic.” Due to this altered fatty acid profile, oil derived from Clearfield™ sunflower line X81359 will require a modified common name to differentiate the oil from traditional sunflower oils in the marketplace.

Nutritional and Compositional Data

The nutritional composition of X81359 sunflower was assessed in whole sunflower grain and compared to comparator line, 8377NS, and other commercial sunflower cultivars. Samples for analysis were obtained from trials at six location years in the United States. The nutritional components included: crude protein, crude fat, crude fibre; branched chain amino acids (valine, isoleucine, leucine); essential amino acids (cysteine, methionine, threonine and lysine); B vitamins (thiamine, niacin, pantothenic acid, pyridoxine, riboflavin, and folic acid); minerals (phosphorus, zinc, magnesium and iron); vitamin E; and fatty acids. Statistically significant differences were observed in the levels of crude protein, threonine, tryptophan, calcium and magnesium. These differences were not biologically significant, and were within the normal range of variation reported for NuSun hybrids. Significant differences were observed in the levels of some fatty acids, such as stearic acid; however, these levels also fell within the ranges determined for other mid-oleic NuSun sunflower oils. The overall nutritional composition of X81359 was therefore determined to be equivalent to the comparator line 8377NS, and to commercial sunflower cultivars, including NuSun hybrids.

The anti-nutrients phytic acid and trypsin inhibitor were measured in X81359 and 8377NS. Phytic acid occurs naturally in sunflower and in cereals. It is indigestible by humans and non-ruminant livestock, and inhibits the absorption of iron and other minerals. Trypsin inhibitor interferes with the enzymes involved in protein digestion. Phytic acid levels in X81359 were not different from those of the comparator line 8377NS, and levels of trypsin inhibitor in both lines were below the limit of detection.

Toxicity and Allergenicity

The impact of single point mutations that confer tolerance of various plant AHAS to imidazolinone herbicides, on the toxic and allergenic potential of the protein has been reviewed previously by Health Canada and the Canadian Food Inspection Agency. No toxicological or allergenic concern was introduced at the level of exposure to AHAS in those products. In the case of oil produced from X81359 sunflower, extensive processing removes protein residues from the oil, and consequently, exposure to the mutated AHAS protein is expected to be negligible.

The AHAS enzyme does not possess the physicochemical properties of toxins and allergens, such as resistance to heat and trypsin degradation. The unmodified form of AHAS is heat sensitive and susceptible to trypsin degradation. Data from studies on the AHAS from X81359 demonstrated the inactivation of the enzyme after 1 minute of heating at 100°C, and its complete degradation in trypsin within 60 minutes of exposure. The unmodified form of AHAS also shows no amino acid similarity to known toxins and allergens, and the modified AHAS differs by only one amino acid. Evidence was also provided to show that the protein components of X81359 were not altered compared to those of unmodified control. Results from HPLC on protein extracts demonstrated similar protein banding pattern in X81359 compared to the unmodified sunflower cultivars. The results of these investigations led to the conclusion that X81359 sunflower did not demonstrate any potential for toxicity and allergenicity compared to conventional unmodified sunflower.

Abstract Collapse

Clearfield™ sunflower hybrid X81359 was not subject to regulation in any jurisdiction except Canada since the development of this herbicide-tolerant line did not employ recombinant DNA technologies. In Canada, regulatory approval is required for use in human food and livestock feed, and for environmental release. X8159 is not intended for cultivation in Canada.

Sunflower (Helianthus annuus L.) was grown as a commercial crop in 67 countries, with a combined harvest 31 million metric tonnes in 2006. The major producers of sunflower in 2004 were Russia, Ukraine, Argentina, China, Romania and France. Sunflower is cultivated as an oilseed crop, as well as for the production of edible seeds for human consumption. Sunflower oil is used in cooking and in the production of margarine, salad dressings and baby formula. Industrial uses of sunflower oil include soaps, hydraulic fluids, paints and plastics. The byproduct of oil extraction, sunflower meal, is used as a livestock feed (CFIA, 2005a).

Weeds are a major production problem in sunflower cultivation. Sunflower is a poor competitor during the early growth stages until canopy closure and weeds during these growth stages compete for light, water and nutrients. Weeds can be managed using a combination of cultural practices (e.g., inter-row cultivation), integrated weed management (e.g., weed scouting, economic thresholds) and the use of herbicides. Herbicides can be applied before the crop emerges (e.g., ethalfluraline, trifluralin), or after (e.g., imazamethabenz, clethodim, fluazifop-p-butyl). The build-up of weed populations can be stemmed by applying herbicides on summer-fallowed fields, and by practicing crop rotation, which allows the use of different herbicides. Rotating among herbicide groups also prevents the development of herbicide-resistant biotypes.

Clearfield™ sunflower hybrid X81359 was developed to allow the use of imidazolinone herbicides as a post-emergence weed control option in sunflower production. The mode of action of imidazolinone herbicides consists of inhibiting the activity of acetohydroxyacid synthase (AHAS), an enzyme in plants active in glycolysis and in the biosynthesis of the branched-chain amino acids isoleucine, leucine and valine. The result of the inhibition of AHAS activity is a decrease in protein synthesis, and an accumulation of toxic levels of alpha-ketoglutarate, all of which causes the eventual death of the plant. While unmodified sunflower is not tolerant to imidazolinone, X81359 has been modified to survive an otherwise lethal application of this herbicide. The tolerance to imidazolinone herbicides in X81359 is due to a naturally occurring mutation in the AHAS gene in a wild population of Helianthus annus. This trait was introduced into X81359 using conventional plant breeding techniques.

The livestock safety of X81350 was based on the evaluation of the similarity of AHAS, in structure and function, to the enzyme naturally present in food and livestock feeds and the lack of toxicity or allergenicity of the modified AHAS. The nutritional equivalence and wholesomeness of X81350 compared to conventional sunflower was demonstrated by the analysis of key nutrients in the grain including proximates (e.g., crude protein, crude fat, crude fibre, ash, moisture), amino acids (valine, leucine and isoleucine), and fatty acids, as well as by the composition in anti-nutritional factors (phytic acid and trypsin inhibitor).

Links to Further Information Expand

Canadian Food Inspection Agency, Plant Biosafety Office Office of Food Biotechnology, Health Canada

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