Abstract
BackgroundPositive feedback is a common mechanism used in the regulation of many gene circuits as it can amplify the response to inducers and also generate binary outputs and hysteresis. In the context of electrical circuit design, positive feedback is often considered in the design of amplifiers. Similar approaches, therefore, may be used for the design of amplifiers in synthetic gene circuits with applications, for example, in cell-based sensors.ResultsWe developed a modular positive feedback circuit that can function as a genetic signal amplifier, heightening the sensitivity to inducer signals as well as increasing maximum expression levels without the need for an external cofactor. The design utilizes a constitutively active, autoinducer-independent variant of the quorum-sensing regulator LuxR. We experimentally tested the ability of the positive feedback module to separately amplify the output of a one-component tetracycline sensor and a two-component aspartate sensor. In each case, the positive feedback module amplified the response to the respective inducers, both with regards to the dynamic range and sensitivity.ConclusionsThe advantage of our design is that the actual feedback mechanism depends only on a single gene and does not require any other modulation. Furthermore, this circuit can amplify any transcriptional signal, not just one encoded within the circuit or tuned by an external inducer. As our design is modular, it can potentially be used as a component in the design of more complex synthetic gene circuits.
Highlights
Positive feedback is a common mechanism used in the regulation of many gene circuits as it can amplify the response to inducers and generate binary outputs and hysteresis
Subsequent experiments involving anhydrotetracycline induction were conducted in E. coli strain GN100 (F- ilvG rfb-50 rph-1 ΔenvZ::FRT attBl::[PN25tetR lacIq spcR]) and those involving aspartate induction were performed in GN101 (F- ilvG rfb-50 rph-1 ΔenvZ:: FRT)
Design of positive-feedback amplifier In order to construct a positive feedback circuit, we required a transcriptional activator that did not interfere with native gene regulation in E. coli
Summary
Positive feedback is a common mechanism used in the regulation of many gene circuits as it can amplify the response to inducers and generate binary outputs and hysteresis. In the context of electrical circuit design, positive feedback is often considered in the design of amplifiers. May be used for the design of amplifiers in synthetic gene circuits with applications, for example, in cell-based sensors. Positive feedback is a common mechanism involved in the regulation of genetic circuits [1]. Any time a gene product has the capacity to enhance its own production, either directly or indirectly, the circuit is said to involve positive feedback. A number of behaviors can be attributed to positive feedback loops. Positive feedback is an integral element in many oscillatory, pattern-formation, and intracellular polarization processes [2,3]
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