Jul 4, 2005

Research Projects

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Transgenic fructan potatoes – does genetic modification lead to unexpected metabolic changes and does this have an impact on drought tolerance?

(2001 – 2004) Max Planck Institute (MPI) of Molecular Plant Physiology, Potsdam

Topic

As a result of genetic modification, the fructan potato stores a group of carbohydrates that is typically found in artichoke plants. The aim of the project was to check whether unpredictable metabolic changes take place in addition to the expected and desired modification.

Since fructan formation is assumed to improve drought resistance (fructans can protect cell components against dehydration damage), drought tolerance was chosen as the trait to be tested for. The behaviour of genetically modified strains was compared with that of the parent strain under drought stress.

It was hoped that the results of these tests would provide information about whether it is possible to discover unexpected effects of genetic modifications with a high degree of certainty using methods currently available.

Summary

As a result of genetic modification, the transgenic potatoes accumulate fructans in tubers and, with strong seasonal fluctuations, in leaves. The tests showed that the fructan-accumulating potato plants do not display higher drought tolerance than their unmodified parent variety. To look for unexpected transgenic effects, RNA expression patterns were also analysed. Here too it was ascertained that the transgenic effect is much smaller than the naturally occurring drought stress effects and is therefore negligible.

Experiment description

The aim of the project was to detect unexpected metabolic changes caused by the genetic modification. As a working hypothesis it was assumed that the expression of the fructan synthesis gene influences metabolism in such a way that the plant’s tolerance to drought is modified. For this reason, genetically modified strains and their control partners in the experiment were subjected to different levels of drought stress. These experiments took place under controlled conditions in climate chambers. The following tests were carried out at the harvest date:

• Analysis of the growth and water metabolism parameters (relative water content, saturation water content, osmotic potential)
• Analysis of biochemical parameters (including fructan content, content of soluble sugars in leaf and tuber)
• Analysis of RNA expression patterns . To create the RNA expression pattern, the RNA concentrations of several thousand genes are measured. In this way, the activity of several thousand genes can be assessed simultaneously. Checks are carried out to assess whether this method is suitable for detecting unexpected metabolic changes caused by the transgene.

Results

In research years 2001 to 2003 experiments were carried out in climate chambers, and plant material of the transgenic strains and the parent variety was produced for the field trials.

System for the reproducible adjustment of drought stress. Plant pots stand on a water-conductive sponge standing in a basin with a constant water level (h). The greater the distance h, the lower the water content in the pots.

Growth of fructan potatoes (SST, SST/FFT) and their parent variety (wild type) under control conditions (12 cm) and drought stress (18 and 20 cm). There are no indications of increased drought stress tolerance in the fructan potatoes.

Fructan content in the leaves of fructan potatoes (SST, SST/FFT) by season.

Analysis of the growth and water metabolism parameters

A crop system was established in which plants were reproducibly subjected to drought stress under field-like conditions. Stress conditions can be simulated in order to measure the reaction of the plants using selected growth and water metabolism parameters.

So far, the tests have not provided any indication of increased drought tolerance in the transgenic potatoes compared with the parent variety.

Analysis of biochemical parameters

To accompany the field trials and the greenhouse experiments being run by our project partners, concentrations of fructans and other sugars were measured in several hundred samples. This gave the project partners background data for the interpretation of the results. The field sample measurements confirmed the expected fructan accumulation in the tubers. In the roots, the fructan content is on the detection limit of the measuring system. The fructan content of the leaves showed an interesting seasonality (illus. 3), although the leaves’ saccharose content, from which fructans are synthesised, remained constant. This distinct seasonality was observed in 2001 and 2003 but could not be reproduced under greenhouse conditions. For biosafety research this means that investigations into environmental effects must be carried out on field material from the same vegetation section in which they are expected to occur. Otherwise the findings cannot be transferred to the field situation.

Analysis of the RNA expression patterns

The measurements of RNA expression profiles were initially carried out on material from plants of the unmodified parent variety that had been grown under different water supply conditions. The comparison of leaf samples of drought-stressed potatoes with control plants (grown without drought stress) showed that, as expected, stress-regulated genes are activated in the former. The different propagation systems produced very different expression profiles.

The comparison of these measurements with those from transgenic plants that had been cultivated in the same way showed only very slight differences between the transgenic potatoes and the unmodified parent variety (genotype effect). Only two genes displayed a significant genotype effect. By contrast, the expression of 179 genes was strongly influenced by the water supply. According to these results, the differences between transgenic plants and the unmodified parent variety are much smaller than between control plants and plants that were subjected to the kind of drought stress that regularly occurs at the location. The transgenic effect is therefore negligible. However, appropriate research needs to be carried out on field material before a generalisation can be made.