Modelisation de la croissance d'eschscholtzia Californica a l'aide d'une plateforme de culture a haute capacite analytique.

摘要

Key words: Eschscholtzia californica, minibioreactors, high analytical capability, kinetic model, cell growth, optimal concentrations, intracellular.;Over the past 30 years, industrial technology from plant cells was ineffective. This is partly explained by the fact that the conditions of growth of these cells are not optimal. To determine the optimal culture conditions, it is necessary to model the growth kinetics. However, the existing models are not developed to adequately explain the complexity of cell growth. In addition, multivariate analysis is needed to understand this complexity.;The first objective of this work is to develop a platform with a high analytical capability to follow multiple cell cultures. This platform is composed of 9 minibioreactors incorporated into a pipetting robot Freedom Evo (c) (TECAN). In addition to aseptically and automatic cell cultures, this robot cans replicate 7 growth variables analysis: total sugar, glucose, phosphate, nitrate, ammonium, biomass and cell count. The selected analytical methods are simple, rapid, inexpensive and require low sample volume. For each sample, only 400 microl are required for the analysis of seven variables in duplicate. For nine cultures, this gives a total of 126 analyzes. In addition, the sequence analysis only takes an hour to perform. This platform will test and monitor different culture conditions in a short time, with little equipment and labor. This will get more followed culture used to develop a kinetic model of cell growth quickly.;The development of a growth model corresponds to the second objective of this work. The model is segregated type and unstructured. Rather than being based on a constant yield (substrate/biomass), this model uses the stoichiometric ratios from mass balances to estimate nutrient consumption and product biosynthesis. The selected test platform cell model is a line of cells Eschscholtzia californica (California poppy) was dedifferentiated and suspended. This plant produces sanguinarine, an alkaloid promising for the treatment of several cancers. This model showed that the optimal concentrations of absorption of the different substrates are higher than 19 mM glucose and nitrate, 1.9 mM phosphate and 8 mM ammonium. These values represent the optimal concentrations for which it obtains more than 95% of the maximum speed (mu max) reactions entered nutrients into the cell.