TY - JOUR
T1 - Escherichia coli “Marionette” strains with 12 highly optimized small-molecule sensors
AU - Meyer, Adam J.
AU - Segall-Shapiro, Thomas H.
AU - Glassey, Emerson
AU - Zhang, Jing
AU - Voigt, Christopher A.
N1 - Funding Information:
This work was supported by the US Office of Naval Research Multidisciplinary University Research Initiative grant #N00014-16-1-2388 (A.J.M., T.H.S.-S., E.G., J.Z., and C.A.V.). This work was supported in part by the Koch Institute Support (core) Grant P30-CA14051 from the National Cancer Institute. We would like to thank A.M. Kunjapur and K.L.J. Prather (Department of Chemical Engineering, Massachusetts Institute of Technology) for providing DNA templates for the amplification of PVan, P3B5, vanR, and pcaU and performing the initial characterization of VanR in E. coli. We would also like to thank S. Liu at the MIT-Broad Foundry for assisting in the RNA-seq and ribosome profiling sequencing run.
Publisher Copyright:
© 2018, The Author(s), under exclusive licence to Springer Nature America, Inc.
PY - 2019/2/1
Y1 - 2019/2/1
N2 - Cellular processes are carried out by many genes, and their study and optimization requires multiple levers by which they can be independently controlled. The most common method is via a genetically encoded sensor that responds to a small molecule. However, these sensors are often suboptimal, exhibiting high background expression and low dynamic range. Further, using multiple sensors in one cell is limited by cross-talk and the taxing of cellular resources. Here, we have developed a directed evolution strategy to simultaneously select for lower background, high dynamic range, increased sensitivity, and low cross-talk. This is applied to generate a set of 12 high-performance sensors that exhibit >100-fold induction with low background and cross-reactivity. These are combined to build a single “sensor array” in the genomes of E. coli MG1655 (wild-type), DH10B (cloning), and BL21 (protein expression). These “Marionette” strains allow for the independent control of gene expression using 12 small-molecule inducers.
AB - Cellular processes are carried out by many genes, and their study and optimization requires multiple levers by which they can be independently controlled. The most common method is via a genetically encoded sensor that responds to a small molecule. However, these sensors are often suboptimal, exhibiting high background expression and low dynamic range. Further, using multiple sensors in one cell is limited by cross-talk and the taxing of cellular resources. Here, we have developed a directed evolution strategy to simultaneously select for lower background, high dynamic range, increased sensitivity, and low cross-talk. This is applied to generate a set of 12 high-performance sensors that exhibit >100-fold induction with low background and cross-reactivity. These are combined to build a single “sensor array” in the genomes of E. coli MG1655 (wild-type), DH10B (cloning), and BL21 (protein expression). These “Marionette” strains allow for the independent control of gene expression using 12 small-molecule inducers.
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U2 - 10.1038/s41589-018-0168-3
DO - 10.1038/s41589-018-0168-3
M3 - Article
C2 - 30478458
AN - SCOPUS:85057602225
SN - 1552-4450
VL - 15
SP - 196
EP - 204
JO - Nature Chemical Biology
JF - Nature Chemical Biology
IS - 2
ER -