Development and Optimisation of a Defined High Cell Density Yeast Medium

Saccharomyces cerevisiae cells grown in a small volume of a defined media neither reach the desired cell density nor grow at a fast enough rate to scale down the volume and increase the sample number of classical biochemical assays, as the detection limit of the readout often requires a high number of cells as an input. To ameliorate this problem, we developed and optimised a new high cell density (HCD) medium for S. cerevisiae. Starting from a widely-used synthetic medium composition, we systematically varied the concentrations of all components without the addition of other compounds. We used response surface methodology (RSM) to develop and optimise the five components of the medium: glucose, yeast nitrogen base, amino acids, mono-sodium glutamate and inositol. We monitored growth, cell number and cell size to ensure that the optimisation was towards a greater density of cells rather than just towards an increase in biomass (i.e larger cells). Cells grown in the final medium, HCD, exhibit growth more similar to the complex medium YPD than to the synthetic medium SD, while the final cell density prior to the diauxic shift is increased about three- and tenfold, respectively. We found normal cell-cycle behaviour throughout the growth phases by monitoring DNA content and protein expression using fluorescent reporters. We also ensured that HCD media could be used with a variety of strains and that they allow selection for all common yeast auxotrophic markers.