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QLK3-2002-02097
Developing a genomic toolbox for exploring and exploiting bacterial biodiversity (BACDIVERS)
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Quality of Life - 3. The Cell Factory
| Type of Project |
Shared Cost Research |
| Contract No |
QLK3-2002-02097 |
| Total Cost |
|
| EC Contribution |
1,700,000 EUR |
| Start Date |
01-01-2003 |
| Duration |
36 Months |
Abstract
Array based tools for the characterisation and exploitation of all aspects of bacterial diversity
will be developed: microarrays using oligonucleotides and PCR products, as well as macro-arrays
using PCR products. The technologies will be compared and evaluated. Work will focus on
Sinorhizobium, bacteria capable of nitrogen fixation and formation of root-nodules
on leguminous plants, but the methods developed will be applicable to other bacterial groups
and other genes / biological functions of interest. Initially, a total S. meliloti genome
microarray will be used to study the diversity of housekeeping and accessory genes. Custom
arrays will be developed for: species identification (the key to other data on an organism),
screening for stress-tolerance genes (drought and salt stress are important factors affecting the
efficiency of inoculant Sinorhizobium, strains) and screening for genes involved
in rhizobium-legume symbiosis.
Objectives
The main aims are to:
- develop array-based tools to characterise bacteria, in particular isolates
of Sinorhizobium, for identification of species, screening for new stress-tolerance
genes and for genes involved in rhizobium-legume symbiosis,
- obtain a better insight to
bacterial classification and in what constitutes a bacterial species at the genome level.
- identify genes for drought or salt tolerance and thus make them available for genetic
improvement of other strains that are particularly effective nitrogen-fixers.
- select and test particularly salt tolerant and effective symbiotic strains.
- develop appropriate bioinformatics tools to analyse microarray data.
Activities
- WP1 a collection of reference strains from Sinorhizobium, species and
related bacteria will be assembled, characterised and distributed to the partners. It will be
extended with salt and drought tolerant strains from the Mediterranean area and Russia, and
commercial innoculant strains.
- WP2. will provide bioinformatics support to the other work packages, in particular in searching
sequence databases, primer design and analysis of microarray data.
- WP3. will study the genome composition of Sinorhizobium,. By hybridising a total
S. meliloti genome microarray with strains from the collection (WP1), housekeeping genes
will be identified. Using subtractive hybridisation, accessory genes will be identified. Selected
housekeeping and accessory genes will be sequenced. Using microplate hybridisations, the
distribution and the sequence diversity of genes will be studied.
- WP4. using the sequences of selected housekeeping genes (WP2 and WP3), an
oligonucleotide microarray for identification of Sinorhizobium species will be constructed.
By hybridising with labelled DNA from strains with known sequences, the system will be
optimised and calibrated. It will then be used to screen known and new rhizobial isolates.
- WP5. using the sequences of genes involved in osmotic protection (WP2 and WP3), a
microarray of PCR products will be produced, optimised and calibrated. It will be used to
screen for stress-tolerance genes in strains from WP1.
- WP6. using an extensive sequence database of nif/nod genes, a macroarray, representing
the total known diversity of these genes will be constructed, optimised and calibrated. It will be
used to screen the strain collection and other isolates for new nif/nod genes.
Deliverables
- a set of Sinorhizobium, reference strains, stress tolerant and inoculant strains.
- an inventory of housekeeping and accessory genes in Sinorhizobium.
- sequence databases for housekeeping genes, stress tolerance genes and genes involved
in plant-bacterium symbiosis.
- micro- and macroarrays for identification and for screening of stress tolerance and symbiotic
genes in Sinorhizobium, and other soil and rhizosphere bacteria.
- new stress tolerant, potential inoculant isolates.
Contacts
Coordinator
Participant
