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Optimisation of a protocol for the extraction and purification of DNA from nodules of leguminous plants

Moudjahid DEMBA DIALLO student laureate


Benin
Master in Natural Sciences, Université Cheikh Anta Diop, Dakar, Senegal, 1997

Optimisation d'un protocole d'extraction et de purification d'ADN de nodules de légumineuses

Nitrogen is an important nutrient for all plants. Most plants obtain the nitrogen they require for their growth by absorbing ammonium or nitrate from the soil by their roots. In addition, leguminous plants can also use atmospheric nitrogen gas (N2). Their roots produce nodules containing the bacterium Rhizobium. These bacteria can use nitrogen gas from the air as a nitrogen source and secrete part of the fixed nitrogen, which is used by the plants. After intensive research, this complex process is now relatively well understood. In principle, one can thus select Rhizobium strains with improved symbiotic properties and supply them to plants to achieve improved nitrogen fixation. For developing countries this offers the advantage that the use of expensive nitrogen fertilisers can be limited or avoided. Moreover, fertilizers are an important source of pollution of subsoil and surface water. In the field, inoculation of crop plants with selected Rhizobium strains turned out to be difficult, because these often compete poorly with the wild Rhizobium strains that are already present in the soil. Therefore, further research on the ecology of Rhizobium is of great importance. In his prize-wining thesis Mr. Moujahid Demba Diallo describes the development of efficient methods, based on the genetic material of the bacteria, allowing a rapid and reliable identification of the Rhizobium strains present in nodules. Total DNA, which contains both bacterial DNA and DNA of the host plant, is isolated from the nodules. Subsequently, a well-defined fragment of the bacterial DNA is selectively amplified by PCR (Polymerase Chain Reaction) to obtain a sufficient quantity of material for further characterisation using restriction enzymes. The laureate demonstrates that this method can be applied to nodules of a range of wild and cultivated leguminous plants. Using this method, he subsequently studied the competition between bacteria during the artificial nodulation of different leguminous plants by a mixture of Rhizobium strains. Such methods can contribute significantly to improving the efficiency of symbiotic nitrogen fixation in crops.

Report by Prof. Dr. Jean-Pierre Hernalsteens, Viral Genetics, Vrije Universiteit Brussel, Belgium