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Poster De Conférence Année : 2023

Development of high throughput miniaturized robotic platforms for metabolism investigations of microorganisms

Cécilia Bergès
Edern Cahoreau
Stephanie Heux
Jean-Charles Portais

Résumé

During the past few years, we have developed a fully integrated solution to analyze the fluxome (i.e. in vivo reaction rate associated with a cellular network of an organism), which combines two robotic cultivation and sampling workstation for 13C-labelling experiments with MS and NMR-based isotopic profiling, and tools for processing and interpreting isotopic data. First of all, we developed in collaboration with TECAN, a platform for automated HT-fluxome profiling of metabolic variants. Controlled by two software, this robotic system prepares, runs, monitors and controls the fermentations, and adjusts the pH, temperature and stirrer speed for 48 micro-scale (10 ml) fermentations in parallel. The robotic system also features fully automated metabolite harvesting and extraction (e.g. cell pellets, supernatants and intracellular metabolites) for downstream analysis. Drastically increasing the quantity of biological samples produced, a second robotic platform has been acquired to fully automatize the preparation of biological samples for analysis by liquid chromatography coupled to mass spectrometry (LC-MS) and nuclear magnetic resonance (NMR). The device is able to handle a complete range of sample racks as well as carrying out various types of manipulation such as dilution, aliquoting, addition of buffer and internal standard, filtration, centrifugation, sample pre-treatment (i.e SPE) all while ensuring traceability of all samples. Recently, we focus our development on the handling of low-volume or small-quantity samples in order to analyze matrices for which we have few amounts available or to concentrate classical samples and have access to minority metabolites. We have adapted our robotic platform to pipette a few microliters (2µL) of sample while maintaining excellent precision (CV<10%), and equipped the pipetting arm with capillary needles to fill 1.7mm internal diameter NMR tubes. This opens up new prospects, including the possibility of studying the metabolism of new sample niches such as spheroids or others extracts with very low volumes. We have now a complete high-throughput, miniaturized workflow for microorganisms cultivations, preparing and analyzing miniaturized samples, as well as bioinformatics tools for processing the large datasets generated. This entire workflow is now available to scientists wishing to carry out targeted or non-targeted metabolomics and fluxomic experiments.

Domaines

Biotechnologies
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Dates et versions

hal-04207380 , version 1 (14-09-2023)

Identifiants

  • HAL Id : hal-04207380 , version 1

Citer

Cécilia Bergès, Edern Cahoreau, Lindsay Peyriga, Stephanie Heux, Floriant Bellvert, et al.. Development of high throughput miniaturized robotic platforms for metabolism investigations of microorganisms. Biosynsys, Jul 2023, Toulouse, France. ⟨hal-04207380⟩
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