Separation of a brewing yeast strain of Saccharomyces cerevisiae based on cellular age.

摘要

In yeast, aging appears to be marked by a progressive impairment in cellular mechanisms, resulting in irreversible changes in physiology and morphology. To date, very little has been reported about the biochemical changes that occur in yeast as a function of individual cell aging. To investigate this further, six generations of a brewing yeast strain of Saccharomyces cerevisiae (NCYC 1239) were separated according to cellular age using continuous phased culturing and biotin-streptavidin magnetic cell sorting.;To obtain cells with no bud scars (virgin cells), a concentrated yeast slurry was layered onto sucrose density gradients and centrifuged. The uppermost band from the gradients was collected and cells were biotinylated with biotinamidocaproate- N-hydroxysuccinimide ester, that covalently binds to lysine residues on the yeast cell wall. For continuous phased culturing, biotinylated cells were added to a carbon-limited nutrient medium and growth was synchronized using the doubling time of the cells. Harvested cells were incubated with streptavidin superparamagnetic beads and sorted with a strong permanent magnet. In total, approximately 75% of the biotinylated cells were recovered. Viability testing was conducted using vital staining and plate counts, with >98% viability reported with the vital stain and 37% viability with the agar plates.;In conclusion, continuous phased culture, together with magnetic cell sorting has the potential to become a powerful tool for the study of age-related biochemical changes in yeast. Further studies will focus on ensuring the reproducibility of the method and using the recovered cells to study biochemical changes occurring during yeasts' replicative lifespan.