Mar 9, 2009

Research Projects

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Transgenic aspens: Testing the reliability of male sterility systems

(2008 – 2011) Johann Heinrich von Thünen-Institut, Institute for Forest Genetics, Großhansdorf

Topic

The use of genetically modified poplars in commercial plantations, as a renewable raw material for example, has been under discussion for a number of years. A key consideration in the safety assessment of GM crops is the potential spread of the newly inserted genes via pollen or seed. For this reason, scientists throughout the world are developing strategies aimed at the targeted prevention of gene transfer via pollen or seed (biological confinement).

The aim of this sub-project is to test the reliability of a confinement system developed for aspens. Transgenic aspens from a precursor project which contain gene constructs designed to cause male sterility will be investigated. The genes used contain information for specific cytotoxins (cell toxins). The selection of tissue-specific control elements (promoters) should ensure that the expression of these genes occurs only in the anthers.

The research will focus on the following questions:

  • Is any pollen produced?
  • Is any pollen that may be produced viable?
  • Are the cytotoxins expressed only in the anthers, or also in other tissue such as leaves, shoots or roots?

Precursor project:

Further information on methods:

Experiment description

Male flowers forming on an early-flowering poplar

Formation of male flowers in an early-flowering poplar.

Anthers (stained red) and pollen grains under the microscope

Anthers (dyed red) and pollen grains under the microscope.

Sterile, early-flowering poplar

Sterile early-flowering poplar

No pollen grains have formed in the anthers of the few single flowers.

Approximately two hundred transgenic aspens from a precursor project which have been cultivated in the greenhouse will be investigated. They are equipped with seven different gene constructs which use four different cytotoxin genes and four different promoters. Since poplars normally flower only after eight years, some of these strains also contain an ‘early-flowering’ gene.

Molecular-biological investigations

First the sites in the plant genome where a gene construct has been inserted will be investigated using PCR and Southern Blot.

Tests will also be carried out to determine whether the newly inserted cytotoxin genes are expressed only in the anthers, as intended, or in other tissues as well. This will involve a special type of PCR that measures the number of copies of specific RNA sequences, a procedure which enables conclusions to be drawn about whether and how strongly a gene is expressed.

Microscopic examinations and germination tests

Flower formation in the transgenic poplars will be studied throughout the year. Each flower will be bagged before pollen formation to prevent the spread of any fertile pollen. The presence of pollen will be investigated and quantified by dying the flowers and examining them under a microscope.

The germination capacity of the harvested pollen will be tested either by direct crossing with female stigma or by applying the pollen to agar-coated slides. Any seedlings obtained as a result of the crossing will then undergo molecular analysis.