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Fourteen Ways to Reroute Cooperative Communication in the Lactose Repressor: Engineering Regulatory Proteins with Alternate Repressive Functions

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journal contribution
posted on 06.09.2016 by David H. Richards, Sarai Meyer, Corey J. Wilson
The lactose repressor (LacI) is a classic genetic switch that has been used as a fundamental component in a host of synthetic genetic networks. To expand the function of LacI for use in the development of novel networks and other biotechnological applications, we engineered alternate communication in the LacI scaffold via laboratory evolution. Here we produced 14 new regulatory elements based on the LacI topology that are responsive to isopropyl β-d-1-thiogalactopyranoside (IPTG) with variation in repression strengths and ligand sensitivitieson solid media. The new variants exhibit repressive as well as antilac (i.e., inverse–repression + IPTG) functions and variations in the control of gene output upon exposure to different concentrations of IPTG. In addition, examination of this collection of variants in solution results in the controlled output of a canonical florescent reporter, demonstrating the utility of this collection of new regulatory proteins under standard conditions.