Abstract
One challenge for synthetic biologists is the predictable tuning of genetic circuit regulatory components to elicit desired outputs. Gene expression driven by ligand-inducible transcription factor systems must exhibit the correct ON and OFF characteristics: appropriate activation and leakiness in the presence and absence of inducer, respectively. However, the dynamic range of a promoter (i.e., absolute difference between ON and OFF states) is difficult to control. We report a method that tunes the dynamic range of ligand-inducible promoters to achieve desired ON and OFF characteristics. We build combinatorial sets of AraC-and LasR-regulated promoters containing −10 and −35 sites from synthetic and Escherichia coli promoters. Four sequence combinations with diverse dynamic ranges were chosen to build multi-input transcriptional logic gates regulated by two and three ligand-inducible transcription factors (LacI, TetR, AraC, XylS, RhlR, LasR, and LuxR). This work enables predictable control over the dynamic range of regulatory components.
Highlights
One challenge for synthetic biologists is the predictable tuning of genetic circuit regulatory components to elicit desired outputs
In growing E. coli cells, most promoters are regulated by σ70, a housekeeping transcription factor that binds the −10 and −35 sites of a promoter and enables RNA polymerase to bind and initiate transcription[36]
Transcription rates have been shown to be highly dependent on the sequences of the −10 and −35 sites for promoters regulated by σE and σ70, and certain −10 and −35 site combinations are associated with known transcription rates[19,37,38]
Summary
One challenge for synthetic biologists is the predictable tuning of genetic circuit regulatory components to elicit desired outputs. Many attempts have been made to engineer transcriptional systems that are better suited for use in synthetic gene circuits[13,14,15,16,17,18] Tuning these regulatory pathways involves building and testing circuits with precharacterized −35 and −10 sites[19] and Shine–Dalgarno sequences[20] of varying strengths until the desired properties (e.g., leakiness, fold induction) are empirically achieved; this approach can be extremely laborintensive and costly. These methods can help to control gene expression, they universally alter protein output regardless of inducer concentration and the dynamic range of gene expression may change in unpredictable ways. If one were to increase the copy number of a gene to increase protein production, the rates of production increase for both the OFF state (in the absence of inducer) and the ON state (in the presence of inducer)
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
