P8: Microscale Fermentation Makes Isotope Labeled Metabolic Flux Analysis an Economical High-Throughput Process

Monday, November 7, 2011
Capri Ballroom (Marriott Marco Island)
Matthew F. Conway, Ashish Misra, Joseph Johnnie and Ganesh Sriram, Chemical and Biomolecular Engineering, University of Maryland, College Park, MD
Metabolic engineering, the manipulation of a cell’s biochemical pathways for a beneficial purpose, extensively benefits from organism-wide metabolic flux quantification. Metabolic fluxes enable visualization of carbon traffic through metabolic pathways and thereby provide nontrivial information on which genes or proteins should be engineered. Fluxes can be estimated by isotope-assisted metabolic flux analysis (MFA), a procedure in which cells are grown on carbon-13 or nitrogen-15 labeled substrates and then analysed to find the labeling patterns in their amino acids.  This data is used to map the traffic through the cell's central metabolism.   MFA is experimentally and computationally laborious and employs very expensive labeled carbon sources. We perform MFA on three Saccharomyces cerevisiae (baker’s yeast) strains grown in various conditions in 1 mL culture in microplates. We investigate simple ways to overcome the oxygen transfer limitations at this scale and how the microplate data compares to the same strains grown in 50 mL shake flasks. We also investigate the ease of which different strains can be compared and fermentation parameters, such as induction timing, can be investigated cheaply.
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