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Applied and Environmental Microbiology, February 2009, p. 637-642, Vol. 75, No. 3
0099-2240/09/$08.00+0     doi:10.1128/AEM.01684-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Construction and Enhancement of a Minimal Genetic AND Logic Gate{triangledown} ,{dagger}

Daniel J. Sayut, Yan Niu,{ddagger} and Lianhong Sun*

Department of Chemical Engineering, University of Massachusetts Amherst, 686 North Pleasant Street, Amherst, Massachusetts 01002

Received 21 July 2008/ Accepted 29 November 2008

The ability of genetic networks to integrate multiple inputs in the generation of cellular responses is critical for the adaptation of cellular phenotype to distinct environments and of great interest in the construction of complex artificial circuits. To develop artificial genetic circuits that can integrate intercellular signaling molecules and commonly used inducing agents, we have constructed an artificial genetic AND gate based on the PluxI quorum-sensing promoter and the lac repressor. The hybrid promoter exhibited reduced basal and induced expression levels but increased expression capacity, generating clear logical responses that could be described using a simple mathematical model. The model also predicted that the AND gate's logic could be improved by altering the properties of the LuxR transcriptional activator and, in particular, by increasing its rate of transcriptional activation. Following these predictions, we were able to improve the AND gate's logic by ~1.5-fold using a LuxR mutant library generated by directed evolution, providing the first example of the use of mutant transcriptional activators to improve the logic of a complex regulatory circuit. In addition, detailed characterizations of the AND gate's responses shed light on how LuxR, LacI, and RNA polymerase interact to activate gene expression.

* Corresponding author. Mailing address: Department of Chemical Engineering, University of Massachusetts Amherst, 686 North Pleasant Street, Amherst, MA 01002. Phone: (413) 545-6143. Fax: (413) 545-1647. E-mail: lsun{at}ecs.umass.edu

{triangledown} Published ahead of print on 5 December 2008.

{dagger} Supplemental material for this article may be found at http://aem.asm.org/.

{ddagger} Present address: Department of Biochemistry, 175 South University Street, Purdue University, West Lafayette, IN 47906.

Applied and Environmental Microbiology, February 2009, p. 637-642, Vol. 75, No. 3
0099-2240/09/$08.00+0     doi:10.1128/AEM.01684-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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