Public concerns about the permanent introduction of foreign DNA into food crops
The creation of food crops that sustain cultivated life is one of the greatest accomplishments of mankind. For thousands of years, plant breeders carefully recombined the genetic material available within species barriers and selected for combinations that provided the highest local yields. Just over the last forty years, intensified breeding efforts supported a doubling of these yields for some of the most important crops. There is concern, however, about the potential for much further yield increases. Furthermore, it may be difficult to apply conventional breeding methods to rapidly address increasing demands for more nutritious and healthy foods.

One of the most promising approaches for accelerated plant breeding may be based on genetic engineering. However, rather than engineering the plant’s own genetic material, initial applications of this technology have been directed towards the stable integration of foreign DNA into plant genomes. During the last two decades, hundreds of thousands of transgenic plants have been generated that contain foreign DNA, either created synthetically or derived from bacteria, viruses, fungi, animals, and unrelated plants. An example of such a transgenic plant is Monsanto’s NewLeaf Plus^® potato variety, which contains a total of eleven different foreign genetic elements (Fig. 1). Processor and consumer rejection of this variety resulted in market withdrawal within a year after launch.

Although the representatives of large agricultural biotechnology companies insist that "DNA is DNA, regardless of source," it is not surprising that the large-scale planting of transgenic crops triggered widespread public concerns. A recent market survey in Mississippi showed that 81% of respondents would eat a vegetable with an extra gene from the same vegetable, whereas only 14% would eat that vegetable if it had an extra gene from a virus¹. Furthermore, about 70% of Northern European consumers surveyed in a second poll agreed that it is more acceptable to transfer DNA within than across species boundaries². In the face of this public perception issue, only the acreage of transgenic crops destined for feed, oil, fibers, and processed ingredients has increased over the past decade, whereas products closer to the table, such as fruits and vegetables, have been hindered in their transgenic development.

Public concerns were addressed by Nielsen³, who proposed to diversify genetically modified crops based on the genetic distance between the source of new genetic material and target organism. According to this proposal, the introduction of foreign DNA creates ‘transgenic’ plants, whereas ‘xenogenic’ plants result from the insertion of synthetic DNA for which no naturally evolved genetic counterpart can be found or expected. Some members in these two groups of plants deviate substantially from what has been achieved through conventional breeding. In contrast, rearrangements of genomic material from within the same sexual compatibility group would create ‘intragenic’ plants. Such modifications would often alter traits in a similar but more efficient and precise manner than that of plant breeding.

Improving crops by unleashing their own potential
In an attempt to incorporate public responses into new and friendlier approaches for biotechnology, we developed methods to produce intragenic plants⁴. These methods were first applied by transforming potato plants with genetic elements isolated from either potato itself or wild potato species that are sexually compatible with potato (Fig. 2).

Instead of an Agrobacterium-derived T-DNA, a plant (P-) DNA fragment was used to support the transfer of genetic material from Agrobacterium to individual plant cell nuclei. This P-DNA is delineated by regions that share homology with the left border of Agrobacterium nopaline strains and the right border of octopine strains. It carried a potato-derived expression cassette designed to reduce expression of the bruise-related polyphenol oxidase (PPO) gene in tubers. A positive selectable marker gene was placed on an accompanying T-DNA that also carried a negative selectable marker gene. Upon co-transfer of the P-DNA and T-DNA, potato cells were first temporarily selected for transient T-DNA-based marker gene expression and then temporarily selected against stable integration of the T-DNA. Analysis of hundreds of plants only containing stable P-DNA insertions resulted in the identification of about seventy lines that displayed the anticipated strongly reduced expression of the PPO gene in potato tubers. These tubers were tolerant to black spot bruise, one of the most important traits for processing potatoes.

P-DNAs have now also been isolated from a variety of other plant species including Arabidopsis, tomato, pepper, alfalfa, and barley. In addition to application of the marker-free transformation system described above, an alternative method was developed for recalcitrant crops such as Kentucky bluegrass⁵. This second method relies on the vortexing of germinated seedlings in Agrobacterium suspensions carrying desired DNA. Efficient germline transmission of the (re-) introduced DNA makes it possible to omit a selection step. Treated seedlings are transplanted to soil, allowed to self-fertilize, and screened for desirable genotypes in the next generation.

Some of the traits that can be modified in crops by using their own genetic material include increased yield, disease tolerance, drought and cold tolerance, herbicide tolerance, improved shelf life, reduced processing-induced acrylamide accumulation, reduced allergen production, increased levels of antioxidants such as carotenoids, flavonols, vitamin C and vitamin E, enhanced flavor, and optimized taste⁶. By employing the same genetic material that is also available to plant breeders, genetic engineering approaches may be more readily integrated into existing plant breeding programs than engineering approaches that override the breeding approach by incorporating foreign DNA and developing super traits. All-native DNA transformation efforts delivering intragenic crops can be expected to offer consumer products within the next five years.

1. Lusk JL & Sullivan P (2002) Consumer acceptance of genetically modified foods. Food Technology 56: 32-37

2. Reviewed in: Schaart J (2004) Towards consumer-friendly cisgenic strawberries that are less susceptible to Botrytis cinerea. PhD-thesis, Wageningen University

3. Nielsen KM (2003) Transgenic organisms: time for conceptual diversification? Nat. Biotechnol. 21: 227-228

4. Rommens CM et al. (2004) Crop improvement through modification of the plant’s own genome. Plant Physiol. 135: 421-31

5. Weeks T & Rommens CM (2003) Refined plant transformation. World patent application 2003/ 079765A2

6. Rommens CM (2004) All-Native DNA transformation: a new approach to plant genetic engineering. Trends Plant Sci. 9: 457-464

Caius Rommens
Simplot Plant Sciences
J. R. Simplot Company
Boise, ID
crommens@simplot.com