Tuning genetic control through promoter engineering

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Flow Cytometry
Flow Cytometry is a technique for counting, examining, and sorting microscopic particles suspended in a stream of fluid. It allows simultaneous multiparametric analysis of the physical and/or chemical characteristics of single cells flowing through an optical and/or electronic detection apparatus. (from Wikipedia)

Davidson FACS protocol
Helpful Flow Cytometry Link

In molecular biology, reverse transcription polymerase chain reaction (RT-PCR) is a laboratory technique for amplifying a defined piece of a ribonucleic acid (RNA) molecule. The RNA strand is first reverse transcribed into its DNA complement or complementary DNA, followed by amplification of the resulting DNA using polymerase chain reaction. (from Wikipedia)

Davidson RT-PCR Animation
Wikipedai quantitative PCR

Application of Promoter Library

Growth Yield Pathway
Phosphoenolpyruvate carboxylase on growth yield

Product Formation Pathway
Deoxy-xylulose P synthase on lycopene production


Promoter Strength Metric

Online Supplemental Material

Supporting Figure 5 Fig. 5. The functional promoter library was analyzed by using flow cytometry. The relative average geometric mean fluorescence of the members of the library is illustrated above and exhibited a nearly 3-log fold range after 14 h in a minimal media with 0.1% casamino acids.
Supporting Figure 6 Fig. 6. Above are representative light-field (right) and false-color dark-field (Left) photomicrographs of clones with highly heterogeneous expression of GFP. These promoters were not further analyzed and are not included in the functional promoter library.
Supporting Figure 7 Fig. 7. In contrast, representative clones chosen for further analysis had much more homogenous distributions of fluorescence. Below, two representative sets of photomicrographs are shown. The top two images correspond to a promoter with a relative promoter strength metric of 0.124 and the bottom two to 0.417.