Part:BBa_K1829005:Experience

Applications of BBa_K1829005

The pAga1 promoter is activated by presence of alpha factor and can therefore be used to activate genes in MATa S. Cerevisiae, which endogenously produce alpha factor receptors (Ste2).

Characterization

Figure1: pAga1 induced with alpha factor driving expression of GFP in S. Cerevisiae. Using GFP as a reporter we measured the fluorescence of our cells in order to read the activation levels of pAga1 at varying concentrations of alpha factor.

In order to characterize pAga1 we built a construct with pAga1 driving production of GFP (green fluorescent protein), and transformed it into S. Cerevisiae (budding yeast) cells. We induced these cells with increasing concentrations of mating factor alpha, or alpha factor, which activated the production of GFP. Using flow cytometry we measured the fluorescence levels of our cells which correlated with level of pAga1 activation, thereby using GFP as a reporter for pAga1 activation. The histogram on the right displays the fluorescence levels of our cells (y-axis) at increasing concentrations of alpha factor (x-axis). pAga1 plateaus at about 14-15 fold activation.

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