Unit: DNA Micro-arrays
Director: Coppée, Jean-Yves
The aim of the platform is to develop biochip technology for the study (compartive genomics and gene expression profiling) of model microorganisms and infectious diseases (pathogens and response of the host to the infection).
We have the equipment for making and using DNA chips on various supports (glass slides, nylon membranes, Affymetrix chips) . We are also developing the bioinformatical and statistical tools necessary for the analysis, interpretation and handling of the results.
The availability of the complete genome sequences of a growing number of microorganisms and the emergence of new technologies for the study of gene expression lead the Pasteur Institute to set up a platform dedicated to the fabrication and analysis of biochips for the study of model microorganisms and infectious diseases. These studies involve both the agents responsible for these pathologies (comparative genomics and gene expression profiling) and the response of the host (gene expression profiling).
At present, three technologies are used at the platform :
Microarrays : spotting of PCR products or long oligos onto glass slides and hybridization with fluorescent probes
Macroarrays : spotting of PCR products onto nylon membranes and hybridization with radioactive probes
Affymetrix : use of commercial chips containing in situ synthesized short oligos
The platform is in charge of projects selected by internal calls for proposals. Most of the selected projects have national or european frameworks. The selected projects receive fundings from the Genopole and are made as collaborations.
For the selected projects, the platfom is in charge of :
the design of the oligos (primers for PCR amplification and long oligos for spotting)
the PCR amplifications (including quantification and quality control)
the spotting onto the nylon membranes or the glass slides
The platform also has training activities for :
The target preparation (fluorescent and radioactive) and the hybridizations
The detection and quantification of the hybridization signals
The statistical analysis (normalization, differential analysis), the interpretation (data clustering) and the handling (databases) of the results.
The platform has also some services activities. This implies a limited participation to some projects (which receive no fundings from the Genopole) according to the availability of the equipement and the people of the platform.
Projects centered on pathogens
In the year 2003, three projects aiming at gene expression profiling of pathogen microorganisms have been started :
Comparison of the transcriptomes of Yersinia pestis and Y. pseudotuberculosis
Elisabeth Carniel, Laboratoire des Yersinia
Use of DNA chips to sudy the virulence of the human parasite Entamoeba histolytica
Nancy Guillen-Aghion, Unité de Biologie Cellulaire du Parasitisme
Search for small regulatory RNAs in Listeria monocytogenes
Pasacle Cossart, Unité des Interactions Bactéries Cellules
The platform keeps on working on several projects which had been selected during previous calls for proposals and which are still pending. It s for example the case of the analysis of the transcriptome of Plasmodium falciparum (we are interested in the effects of various environmental, immune and drug pressures), Bacillus anthracis and Aspergillus fumigatus.
Projects centered on the host response
Initially, most of the activities of the platform were centered on microbiology (we were mainly interested in model and pathogen microorganisms). However, there is indeed an important demand on the campus for an accesse to human and murine chips for the analysis of the response of the host to the infection. In this goal, the Genopole has acquired an Affymetrix station. This is accessible to the whole Institute via calls for proposals. The projects selected by the steering comittee receive fundings (corresponding to 2/3 of the global cost of the study). In 2003, six projects have been selected and are under progress. Most of them deal with the analysis of the gene expression in human or mouse in response to infection by pathogens like Shigella flexneri, Klebsiella pneumoniae, Helicobacter pylori et Mycobacterium tuberculosis.
Keywords: Transcriptomics, comparative genomics, infectious diseases, macroarrays, microarrays, Affymetrix