Part:BBa_K1829001:Experience
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Applications of BBa_K1829001
pBar1 is an alpha factor responsive promoter that can be used to activate gene transcription in MATa S. Cerevisiae cells. It is activated by the presence of mating factor alpha, a signaling molecule utilized in the S. Cerevisiae mating pathway. Unfortunately it is a very weak promoter yielding very low levels of activation (close to zero).
Characterization
pBar1 is an alpha factor responsive promoter found endogenously in S. Cerevisiae, normally regulating the production of a protease called Bar1. However in order to measure the activity of pBar1, we built a construct in which pBar1 regulates the production of GFP (green fluorescence protein) and transformed it into S. Cerevisiae cells. We induced pBar1 with increasing concentrations of alpha factor then used flow cytometry to measure the fluorescent levels of GFP. Using GFP as a reporter we can therefore measure the activation of pBar1. As you can see on the graph to the right, pBar1 has a very low level of activation, especially when compared to another alpha factor responsive promoter, pAga1 (blue graph).
User Reviews
UNIQ9f9fe567d87be21e-partinfo-00000000-QINU UNIQ9f9fe567d87be21e-partinfo-00000001-QINU
UCSF iGEM 2015
The goal of the 2015 UCSF iGEM project was to model divergent community behavior in S. Cerevisiae using the endogenous mating pathway. Our engineered cells communicate but sensing and secreting a communication molecule called alpha factor. In order to create divergent populations within a community
However we suspect the low levels of activation may attribute to the endogenous use of pBar1. In S. Cerevisiae, pBar1 promotes the production of a protease called pBar1 which degrades mating factor alpha. This sharpens concentration gradients therefore creating a threshold of activation for alpha factor responsive promoters