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GENOMYCA
Genes and genetic engineering for arbuscular mycorrhiza technology and applications in sustainable agriculture

Over-fertilising, agricultural additives (like sewage wastes) and atmospheric pollution cause a decrease in soil fertility and plant production. Arbuscular mycorrhizal fungi are symbiotic microorganisms central to plant development in natural conditions and can be used as biotools to avoid heavy chemical inputs. An important amount of research has clearly shown the role of non-culturable, symbiotic arbuscular mycorrhizal fungi as biofertilisers and bioprotectors in plant production systems. However, their sustainable use in agriculture is still unpredictable and emerging SMEs producing inoculum for plant growers are demanding strains that are better adapted to this purpose.

Objectives
The overall objectives of the GENOMYCA project are to improve genetically arbuscular mycorrhizal fungi for exploitation in sustainable agriculture or heavy metal/organically polluted agro-ecosystems. This will be achieved by:
1) using genes from arbuscular mycorrhizal fungi and their endosymbiotic bacteria with key functions in phosphate and nitrogen nutrition or heavy metal tolerance
2) manipulating novel genes to broaden the spectrum of activity of these fungi to the breakdown of organic pollutants
3) producing high quality mycorrhizal inoculum and mycorrhizal plants to be used in confined plant production systems.

Progress to Date
Much of the progress achieved so far has targeted the isolation and characterisation of novel genes with key functions in mineral nutrition and heavy metal tolerance in the arbuscular mycorrhizal fungi, and of nitrogen fixation genes in endosymbiotic bacteria.
Work has also progressed towards the genetic transformation of arbuscular mycorrhizal fungi with the development of an efficient biolistic methodology to introduce genes into spores, and a reporter gene has been selected to monitor vector incorporation into fungal structures.
The transmission mode of endosymbiotic bacteria has been determined, endobacterial cells have been isolated and bacteria-free spores obtained in order to study the role of the endosymbionts in mycorrhiza effectiveness.
In order to evaluate the fungal products obtained within the project, microcosm and mesocosm protocols have been established to produce fungal inocula efficiently and their biological properties tested.

Results
1) Genomic libraries were established for three arbuscular mycorrhizal fungi (Glomus mosseae, Gigaspora rosea, Gigaspora margarita) and for the endosymbiotic bacteria of Gigaspora margarita. Full-length metallothionein and zinc transporter genes, which are upregulated by heavy metals, were cloned from several fungi and their incorporation into transformation vectors was started.
2) Research for new biological properties to be introduced into arbuscular mycorrhizal fungi focused on bacterial genes, which code for an enzyme degrading petrol derivatives (benzene, ethylebenzene, toluene and xylene).
3) Parameters for efficient biolistic methodology, based on firing gold particles coated with DNA into fungal spores, were defined for genetic transformation of all three arbuscular mycorrhizal fungi.
4) First evidence has been obtained showing the existence of prokaryotic genes involved in nitrogen fixation (nif H, K, D) in spores of Gigaspora margarita, representing a notable step towards identifying this function in the endosymbiotic bacteria.
5) Spore supply was optimised for the three arbuscular mycorrhizal fungi, and microcosm (leek, sorghum) or mesocosm (olive microplants) protocols were established to test the biological properties of fungal products obtained within the project.
6) The project has been extended to include two additional participants from NAS institutes in order to cover aspects on cell physiology and the stability of arbuscular mycorrhizal fungi modified for heavy metal tolerance.

Classified in SOIL

Scientist responsible for the project

Dr VIVIENNE GIANINAZZI-PEARSON
LPA, INRA-CMSE, UMR 1088 PME, Box BP 86510
21065 Dijon CEDEX Cedex
France - FR

Phone: +33 3 80 69 31 55
Fax: +33 3 80 69 37 53
E-mail: Vivienne.Gianinazzi-Pearson@epoisses.inra.fr

References

Project ID QLRT-1999-31319
Organisation Institut National de la Recherche Agronomique
Area 5.1.1
Start date 01 February 2001
Duration (months) 36
Total cost 3 184 096 €
Total EC contribution   1 850 458 €
Status Ongoing
Web address of the project   http://www.dijon.inra.fr/genomyca/

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