The aim of the platform is to develop biochip technology for the study of model microorganisms and infectious diseases (comparative genomics and gene expression profiling). We have the expertise in the making and the analysis of microarrays (glass slides hybridized with fluorescent probes) and macroarrays (nylon membranes hybridized with radioactive probes).

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). The platform has at its disposal robotic stations for the arraying of nylon membranes and glass slides. It has the expertise in the labeling and hybridization of the probes (fluorescent and radioactive), in the analysis of hybridization signals and in the interpretation of the data. The platform is in charge of projects selected by internal calls for proposals. Most of the selected projects have national or european frameworks.

Following the first call for proposals, 5 projects have been completed and two others are in progress:

For all these projects based on the macroarray technology (arraying of PCR products on nylon membranes and hybridization with radioactive probes), the platform performed the PCR amplifications (primers design, amplification reactions, PCR controls by agarose electrophoresis, random sequencing of 10 % of the PCR products), the arraying and the quality controls of the macroarrays (hybridization with control probes). These macroarrays are now in use by the teams of the Pasteur Institute involved in the projects.

Important developments of the microarray technology have also been performed.

These include :

• Glass slides chemistry (three types of slides are used : polylysines, amines, aldehydes)
• Labeling of the probes (direct and indirect labeling of RNA and DNA)
• Arraying of the slides (PCR products and oligonucleotides)
• Quality controls (RNA, labeling efficiency, arraying quality)
• Reproducibilty of the so obtained gene expression profiles

Most of these improvements have been obtained in the frame of a project aiming at the global gene expression analysis in Plasmodium falciparum (collaboration with Peter H. David, Unité d'Immunologie Moléculaire des Parasites). A complete set of 6000 oligonucleotides (70mer) covering most of the P. falciparum identified genes has been spotted on polylysine glass slides. These slides are now in use for the comparison of the gene expression profiles associated with the different development stages of the parasite.

Maitournam Aboubakar

Sismeiro Odile, osisme@pasteur.fr