Biology Reference
In-Depth Information
expressed and the cell dies off. However, the prey
E. coli
strain synthesizes and releases the
quorum sensing molecule, 3OC6HSL, which activates the expression of the antidote gene
ccdA
, which blocks the activity of
ccdB
, thereby saving the predator
E. coli
. As the predator
E. coli
population increases, the concentration of another quorum sensing molecule,
3OC12HSL, increases. This molecule induces the expression of
ccdB
in the prey
E. coli
,
resulting in the death of prey cells. Consequently, the predator
E. coli
cannot survive without
its
which is then killed off by the predator when the predator population achieves a
certain cell density. Eventually steady-states in the populations of both strains are achieved
with predator
E. coli
killing off the prey, and the prey
E. coli
saving the predator.
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prey,
'
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Genetic Oscillators
Oscillators are important characteristics of biological systems as many cellular functions
display oscillating behaviors, such as cell cycle regulation and circadian rhythms. These
oscillators can be constructed using simple components already present in the cell, with
additional features to confirm and evaluate the synthetic oscillatory behavior. One such
oscillating network was developed in
E. coli
using a three transcriptional repressor system
(repressilator) that is not found in any natural biological clocks.
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This oscillating network
was used to monitor the state of the individual
E. coli
cells through the periodic synthesis of
GFP.
23
Three different types of transcriptional oscillator have also been constructed and evaluated
in vitro: (1) a negative feedback oscillator regulated by excitatory and inhibitory RNA
signals; (2) a positive-feedback loop to extend and modulate the oscillatory regime of the
negative feedback oscillator; and (3) a three-switch ring oscillator that is analogous to the
repressilator.
8
Mathematical modeling was employed using kinetic models in the design and
analysis of the three oscillators. Using the mathematical model, the robustness of the
regulatory oscillator to different conditions was analyzed, which allowed better
understanding of oscillator
145
s principles of operation within the kinetic models.
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Implementing Synthetic Biology Components Through Engineering of
Cellular Components
The implementation of these genetic circuits is achieved through the engineering of different
native components to display the numerous decision nodes needed in constructing the
genetic circuits. One such tool that allows for the implementation of synthetic biology
genetic circuits is RNA programming.
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RNA has become an important tool in synthetic
biology with increased knowledge of their functional roles in cellular physiology, beyond
being a bridge between DNA and proteins. Small RNA molecules have been found to
perform regulatory functions through attenuating genetic expression. Through the design of
functional synthetic RNA molecules that exhibit complex functions, cellular circuits can be
engineered to perform high-level biological functions.
18
Small hairpin RNAs (shRNA) are small molecules of RNA with tight hairpins that have been
used to silence gene expression through ligand control of RNA interferences (RNAi).
18
One
of the limitations of employing shRNA as a regulatory control element is the lack of
predictive tools in optimizing the design of shRNA sequences in order to not only alter and
tune the system
s response to perturbations, but also to broaden the implementation of
shRNA to a wide range of systems. The shRNA switch platform was developed to provide
ligand control of RNAi in mammalian cell lines, which provides a fine-tuning, multi-input
control, and model-guided design of regulatory systems.
18
The platform utilizes a strand
displacement strategy where ligand binding, RNAi activation, and translation of the binding
interaction are isolated into individual domains within the platform. Standard RNA folding
algorithms were used to establish quantitative sequence-to-function relationships. By
demonstrating combinatorial tuning strategies and multi-input control, the switch
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