Quorum Sensing: Week 6 Results - PowerPoint PPT Presentation

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Quorum Sensing: Week 6 Results

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Proc Natl Acad Sci U S A. 2002 June 11; 99(12): 7877 7882. doi: 10.1073/pnas.082243699. FACS - Inspiration Anderson, J. C., Clarke, E. J., Arkin, A. P., ... – PowerPoint PPT presentation

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Title: Quorum Sensing: Week 6 Results


1
Quorum Sensing Week 6 Results
  • Two Cell vs. One Cell Systems

2
Switched Fluorophores
Original Constructs
Senders
lacI regulated RFP Sender (S08I07)
tetR regulated RFP Sender (S23I07)
Receiver
GFP Receiver (T02)
3
Switched Fluorophores
Flipped Constructs
Senders
lacI regulated GFP Sender (S08E40)
tetR regulated GFP Sender (S23E40)
Receiver
RFP Receiver (F20I07)
4
Switched Fluorophores
5
Switched Fluorophores
6
Switched Fluorophores
  • Protein Folding/Maturation issue?
  • GFP (E0040)
  • gfpmut3b
  • The expression of GFP is observable minutes
    after induction1
  • RFP (E1010)
  • mRFP1 - highly engineered mutant of red
    fluorescent protein from Discosoma striata
    (coral)
  • Based on our data at 37C, we estimate the t0.5
    for maturation of mRFP1 and T1 is less than 1 h,
    consistent with the 0.7 h previously reported for
    T12

1 FACS-optimized mutants of the green
fluorescent protein (GFP).Gene 0378-1119
Cormack yr1996 vol173 iss1 Spec No pg33 -8 2
A monomeric red fluorescent protein. Proc Natl
Acad Sci U S A. 2002 June 11 99(12) 78777882.
doi 10.1073/pnas.082243699.
7
FACS - Inspiration
Anderson, J. C., Clarke, E. J., Arkin, A. P.,
Voigt, C. A. (2006) Environmentally Controlled
Invasion of Cancer Cells by Engineered Bacteria,
Journal of Molecular Biology, 355 (4) 619-627.
8
FACS - Results
110 Dilution Final OD 0.02 Mean GFP 370
1100 Dilution Final OD 0.141 Mean GFP 315
11000 Dilution Final OD 0.555 Mean GFP 426
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