Institut de
Génomique
Fonctionnelle

 MGX – Montpellier GenomiX
 FPP – Plate-forme de Protéomique Fonctionnelle
 Small Animal Imaging Core Facility of Montpellier
 Plate-forme ARPEGE - Pharmacologie Criblage Interctome
 Animal facilities

MGX – Montpellier GenomiX

Technological highlights :
The MGX (Montpellier GenomiX) facility is labelled by the IBiSA committee, and is devoted to :
 The printing of microarrays from pangenomic oligonucleotide collections. Operon provided the human 21K, mouse 17K, drosophila 15K and grape 14K oligosets. We additionally use a drosophila 15K oligoset provided by the INternational Drosophila Array Consortium (INDAC ; www.indac.org) that selected the Montpellier facility to provide the French community with drosophila microarrays. We print microarrays from proprietary collections such as the CIRAD rice 21K cDNA collection, as well as from collections for various micro-organisms. The Montpellier facility was recently selected by a research consortium formed by Nestlé, CIRAD and INRA to provide the consortium with 12K coffee chips.
 The hybridization-scanning-quantification of various commercially available microarrays. These include Operon rat 27K, mouse 36K and human 35K microarrays, as well as high density (385K to 2.1M) NimbleGen microarrays.
 The operation of a next generation sequencer, i.e. the Illumina Genome Analyzer II. Available applications include de novo sequencing, re-sequencing as well as all tag census-based techniques, i.e. RNA-Seq, DGE, ChIP-Seq…
 The analysis of microarray and deep sequencing data. The facility helps users in designing their microarray experiments. It offers a full service for normalization and analysis of the data using the most appropriate statistical techniques. The facility is assisting users in publishing their microarray experiments through the BASE open-source LIMS that was implemented on the facility. The LIMS also provides an interface to various 2nd (basic statistics, differentially expressed genes, clustering) and 3rd (over-representation of GO terms, identification of TF binding sites in promoters of differentially expressed genes…) level analysis tools.

Identification of Zac1 target genes : 16,463 long oligonucleotides from the mouse Array-Ready Oligo Set (Operon) were spotted in duplicate onto Amersham type 7* slides, and hybridized with polyA+ RNA isolated from CAT- and Zac1-transfected Neuro2a neuroblastoma cells. Platform operative process and rules : The facility is operating in agreement with the IBiSA charter. Following project approval, the operating rule is ʻfirst in, first outʼ.

Technological developments The Montpellier facility is a partner of the AgriArray project funded through the Genoplante ANR. The project is coordinated by GeneWave, a biotech company located on the Polytechnique campus near Paris. The purpose of this proposal is to develop a platform using infrared fluorescent dyes, which display a highly improved signal to noise ratio compared to conventional cyanine dyes. GeneWave developed novel reflective slides based on their proprietary dielectric mirror technology, as well as a devoted infrared scanner. The Montpellier facility was selected to perform pilot experiments with various infrared fluorophores, and to perform proof-of-concept experiments using its expertise in plant microarrays. Funding : MGX is funded through service costs and competitive, project-focused grants from the ANR, IBiSA and the Languedoc-Roussillon district.

Publications :
The MGX platform contributed to the following five selected publications :
 Truffinet et al. (2007) The 3’ IgH locus control region is sufficient to deregulate a c-myc transgene and promote mature B cell malignancies with a predominant Burkitt-like phenotype. J Immunol. 179:6033-42
 Zeitouni et al. (2007) Signalling Pathways Involved in Adult Heart Formation Revealed by Gene Expression Profiling in Drosophila. PLoS Genet. 3:e174
 Fernandez et al.(2007) Identification of genes associated with flesh morphogenesis during grapevine fruit development. Plant Mol Biol 63:307-23
 Varrault et al. (2006) Zac1 belongs to an imprinted gene network critically involved in the control of embryonic growth. Dev Cell 11:711-22
 Negre et al. (2006) Chromosomal distribution of PcG proteins during Drosophila development. PLoS Biol 4:e170.

Plate-forme de Protéomique Fonctionnelle de Montpellier

La Plate-forme de Protéomique Fonctionnelle (FPP - www.fpp.cnrs.fr) est une plate-forme du Pôle Protéome de Montpellier (PPM), structuration régionale Languedoc Roussillon des capacités technologiques en analyse protéomique. La plate-forme est labellisée IbiSA et certifiée ISO 9001-2008. Elle participe dans le cadre de collaborations à des projets s’inscrivant dans des programmes nationaux (INCA, pôles de compétitivité EuroBioMed et MEDICEN, ANR) et internationaux (Programmes Européens), principalement dans les domaines de la cancérologie, la neurobiologie et la parasitologie.

La plate-forme est équipée des technologies les plus performantes en spectrométrie de masse, en particulier de spectromètres de masse à Transformée de Fourier (LTQ Orbitrap XL-ETD et VELOS) permettant des analyses à haute sensibilité et haute résolution. Au-delà de la caractérisation à large échelle des protéomes dans un contexte biologique donné, la FPP développe des approches de protéomique différentielle basées sur des analyses quantitatives de l’expression des protéines grâce à un ensemble de technologies récentes et performantes (analyse d’image de gels à deux dimensions, quantification par spectrométrie de masse par marquage métabolique - SILAC -, chimique - TMT, dimethylation – ou sans marquage - Label-free -). La FPP est engagée dans des programmes variés, incluant la caractérisation de réseaux multiprotéiques impliqués dans la signalisation cellulaire, l’analyse dynamique des modifications post-traductionnelles (phosphorylation, sumoylation…) et la recherche de biomarqueurs dans les maladies neurodégénératives (Programme Diatral - Diagnostic et Traitement de la Maladie d’Alzheimer).

Small Animal Imaging Core Facility of Montpellier

Technical highlights

IPAM (Imagerie du Petit Animal de Montpellier, http://ipam.igf.cnrs.fr/) was created in 2004 as the development section in small animal optical imaging of the Platform of Functional Exploration (National Network of Genopoles, RNG). IPAM has participated with the Royal College of Surgeons in Ireland in the build-up of the National Biophotonics & Imaging Platform Ireland (NBIPI, 2007-2011) (http://research1.rcsi.ie/BIP/index.asp), a 30million euro platform funded under the Higher Education Authority of Ireland (HEA). IPAM is also funded by IBiSA due to its links with MRI (http://www.mri.cnrs.fr/index.php?m=9). IPAM staff includes biologists (P. Mollard and M. Desarménien), engineers with strong background in cellular imaging and animal handling (C. Lafont and M. Schaeffer), engineers in biocomputing (P. Fontanaud, M. Cassou, 3 M2-Pro stud.), and one mathematician (F. Molino). IPAM objectives are to develop/adapt cellular in vivo imaging techniques with long-range objectives (working distance ≥ 1.3 cm) in order to monitor in real-time cell signals in organs/preparations (ventral side of brain, pancreas…), which remain inaccessible to most high resolution microscopy techniques because the latter work with short working distance objectives (e.g. x20 water-immersion objectives with a WD of a few mm only for e.g. in vivo 2-photon excitation microscopy). To circumvent this problem, IPAM recently adapted stereomicroscopes (M2-Bio and Discovery from Zeiss) with x20 air-transmission Mitutoyo objectives (one with n.a. 0.42 and wd 2.8 cm and the other with n.a. 0.6 and wd 1.3 cm, see set-up image below) so that organs could be imaged at cellular resolution in anasthetized animal models (transgenic mice, rats). Following their developments, these equipments became available to the scientific community and have been used by several teams from IGF and other labs (e.g. E. Kremer, IGMM Montpellier). Their uses have been extended to nonvertebrate species (e.g. flies by Y. Grau’s team). Complementary to technical developments, IPAM staff generated several custom-made software dedicated to cellular in vivo imaging analysis : (http://ipam.igf.cnrs.fr/index.php?p...).

Plate-forme ARPEGE - Pharmacologie Criblage Interctome

Objectifs :

La plate-forme ARPEGE Pharmacologie-Criblage-Interactome (www.arpege.cnrs.fr) a été créée pour mettre à la disposition de la communauté scientifique (académique ou privée) des outils adaptés aux études pharmacologiques et des techniques de pointes les plus récentes. Ces techniques permettent notamment :
 le criblage de banques de petites molécules, le criblage de protéines mutées comme des récepteurs mutants,
 la caractérisation de ligands sur des banques de récepteurs,
 l’analyse fonctionnelle de récepteurs couplés aux protéines G et de leurs voies de signalisation intracellulaires,
 l’analyse de la composition et de la dynamique des complexes protéiques, comme ceux associés aux récepteurs couplés aux protéines G ou des complexes de signalisation intracellulaires, des dimères, oligomères et hétéro-oligomères protéiques. Ces techniques peuvent être utilisées pour des analyses à moyen débit, dans des plaques 96 et/ou 384 puits, et sont adaptées pour des analyses rapides et de routine.

Un des caractères innovants de la plate-forme ARPEGE Pharmacologie-Criblage-Interactome est l’utilisation de techniques basées sur des principes physiques de fluorescence et le développement de tests utilisant le transfert d’énergie par résonance en particulier le BRET (Bioluminescence Resonance Energy Transfer), le TR-FRET (Time-Resolved Fluorescence Resonance Energy Transfer (FRET)), et l’HTRF (Homogeneous Time Resolved Fluorescence, propriété de Cisbio Int., Bagnols sur Cèze, France) en plus des techniques classiques de scintillation et de fluorescence. Le TR-FRET et l’HTRF ont plusieurs avantages — temps d’émission de la fluorescence élevé, compatibilité spectrale, ratio signal /bruit de fond élevé et peu de contrainte d’orientation — qui en font des méthodes très puissantes pour étudier les interactions protéine-protéine et pour créer des tests de mesure de seconds messagers ou d’activation de voies de signalisation.

Animal facilities

Presentation :

Our animal facility is a common platform that belongs to the IFR3. The IGF has been highly involved in running this facility since most of technicians presently working on the platform were originally IGF staff. The animal facility is currently composed of 3 specific zones, according to their microbiological status and hosted species :
 140 m2 dedicated to breeding and housing of genetically-modified mice, under a specific pathogen-free (SPF) status. The entry into this SPF zone is strictly limited to zootechnicians who take care of the animals. It is located in the IGH building and hosts circa 6 000 mice permanently. 15 000 new animals per year are tagged for a total of 22 user teams. The various mouse models hosted in this facility encompass the interest of the 5 departments of the IGF (e.g. mutated GPCRs, fluorescent reporters of synaptic plasticity or of hormonal release, cancer-prone mice...). An engineer (AI, INSERM) has been specially hired by the IGF since January 2009 for genotyping new animals by real-time quantitative PCR. Such a facility spares tedious and time-consuming bench work for researchers, and ensures timed delivery of genotype identification for animal caretakers.
 30 m2 for housing of rabbits and xenopus under a conventional status. This zone, located in the IGH building, hosts rabbits used for the production of antibodies against specific epitopes, and xenopus that produce ovocytes for developmental biology or for the study of transfected ionic channels.
 60 m2 for rodents under a conventional status, in the IGF building. This facility hosts wildtype mice and rats, and is also dedicated to short-time housing of class I genetically-modified animals in view of quick testing of well-defined scientific hypotheses (promising mouse lines are then decontaminated and transferred into the SPF zone for long-term programs). Moreover, the platform also provides help to researchers with injection protocols or small surgery (orchydectomy, ovariectomy...). We recently established a committee for ethics in animal experimentation. Affiliated with the regional committee of Languedoc -Roussillon (CEEA-LR), this local committee is devoted to provide advices in designing experiments with animals and fill in protocol forms submitted to the CEEA-LR.