Information Technology Reference
In-Depth Information
1,000.00
pINV-102
100.00
pINV-112-R1
10.00
1.00
0.1
1.0
10.0
100.0
1,000.0
10,000.0
IPTG (uM)
Figure 7.4
Controlling signal levels using external induction with isopropylthio-
β
-
galactoside (IPTG).
concentrations. The graph shows how to control an
in vivo
signal using ex-
ternal induction with IPTG. The relationship between the ECFP and EYFP
fluorescence intensities in Figure 7.4 is used to normalize between simulta-
neous ECFP/EYFP readings in subsequent experiments in this chapter. This
genetic setup is used in the following sections to set the levels of input mRNA
to the inverters under study.
The lacI/p(lac) Inverter
Figure 7.5 shows the genetic circuit used to measure the device physics of an
inverter based on the
lacI
repressor and the p(lac) promoter. The first two logic
gates set the level of the input signal to the inverter in a mechanism similar to one
used in the circuit described in the previous subsection. Here, the
λ
P(R
−
O
12
)
inverter functions as a constitutive promoter (no
cI
in the system) to set a
constant high level of the Tet repressor (
tetR
). Then, through the
tetR
/P(LtetO-
1) IMPLIES gate, the concentration of the aTc (anhydrotetracycline) inducer
molecule controls the level of the lac repressor (
lacI
).
lacI
is the input protein
to the inverter gate under study. The ECFP transcribed along with
lacI
reports
the level of the input signal. Finally, EYFP reports the output signal expressed
from the lacI/p(lac) inverter.
The output of this circuit is the logic NOT of the aTc input signal. Figure
7.6a shows FACS cell population data of the EYFP output signal in seperate
experiments where the cells were exposed to different aTc inducer input con-
centrations. For a low input concentration of 3 ng/ml aTc, the output of the
circuit is appropriately high. For a high input concentration of 30 ng/ml aTc,
the output of the circuit is correctly low. The figure also illustrates the good
