Wednesday, August 1, 2007 - 10:30 AM
S151
Metabolic engineering in filamentous fungi: Finding the optimum genetic modification
Vera Meyer1, Dirk Mueller1, Arthur Ram2, Cees AMJJ van den Hondel2, and Ulf Stahl1. (1) Institute of Biotechnology, Department Microbiology and Genetics, Berlin University of Technology, Gustav-Meyer-Allee 25, Berlin, Germany, (2) Institute of Biology Leiden, Fungal Genetics Research Group, Leiden University, Wassenaarseweg 64, Leiden, Netherlands
Filamentous fungi are widely used in biotechnology as cell factories for the production of chemicals, pharmaceuticals and enzymes. In order to achieve higher productivities and minimise unwanted by-product formation, metabolic engineering strategies can be powerful approaches. However, before engineering can become routine, introducing the desired genetic manipulation of the fungus of interest often represents a challenge.
Foremost, the establishment of a suitable transformation method is not trivial for many fungi. Moreover, the mode and frequency of individual integration events resulting from homologous or illegitimate recombination is not only dependent on the transformation host itself but also on the transformation technique applied. Thus, designing an engineering strategy first requires consideration of the most suitable transformation method.
Targeted deletion of key genes is often envisaged as strategy to reduce metabolic fluxes of side pathways. However, gene deletion in filamentous fungi is often hampered by low frequencies of homologous recombination. One approach to overcome this limitation is the use of strains defective for the nonhomologous end joining pathway. An alternative strategy circumventing the need for deleting fungal genes is to use RNA-based methods that silence gene expression post-transcriptionally. Among these, ribozyme technology has been shown to reduce gene expression up to 100%, suggesting that ribozymes represent a versatile tool for timed elimination of unwanted gene products in filamentous fungi.
Here, advantages and disadvantages of different options for genetic engineering such as transformation techniques, gene knock out or knock down approaches will be discussed.
Foremost, the establishment of a suitable transformation method is not trivial for many fungi. Moreover, the mode and frequency of individual integration events resulting from homologous or illegitimate recombination is not only dependent on the transformation host itself but also on the transformation technique applied. Thus, designing an engineering strategy first requires consideration of the most suitable transformation method.
Targeted deletion of key genes is often envisaged as strategy to reduce metabolic fluxes of side pathways. However, gene deletion in filamentous fungi is often hampered by low frequencies of homologous recombination. One approach to overcome this limitation is the use of strains defective for the nonhomologous end joining pathway. An alternative strategy circumventing the need for deleting fungal genes is to use RNA-based methods that silence gene expression post-transcriptionally. Among these, ribozyme technology has been shown to reduce gene expression up to 100%, suggesting that ribozymes represent a versatile tool for timed elimination of unwanted gene products in filamentous fungi.
Here, advantages and disadvantages of different options for genetic engineering such as transformation techniques, gene knock out or knock down approaches will be discussed.