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A portable regulatory RNA array design enables tunable and complex regulation across diverse bacteria

View ORCID ProfileBaiyang Liu, Christian Cuba Samaniego, Matthew R. Bennett, Elisa Franco, View ORCID ProfileJames Chappell
doi: https://doi.org/10.1101/2023.02.24.529951
Baiyang Liu
1Graduate Program in Systems, Synthetic, and Physical Biology, Rice University, Houston, TX, USA
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Christian Cuba Samaniego
2Department of Mechanical and Aerospace Engineering, Bioengineering, and Molecular Biology Institute, University of California at Los Angeles, CA, USA
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Matthew R. Bennett
3Department of Biosciences, Rice University, Houston, TX, USA
4Department of Bioengineering, Rice University, Houston, TX, USA
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Elisa Franco
2Department of Mechanical and Aerospace Engineering, Bioengineering, and Molecular Biology Institute, University of California at Los Angeles, CA, USA
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James Chappell
3Department of Biosciences, Rice University, Houston, TX, USA
4Department of Bioengineering, Rice University, Houston, TX, USA
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  • For correspondence: jc125@rice.edu
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ABSTRACT

A lack of composable and tunable gene regulators has hindered efforts to engineer non-model bacteria and consortia. To address this, we explore the broad-host potential of small transcription activating RNA (STAR) and propose a novel design strategy to achieve tunable gene control. First, we demonstrate that STARs optimized for E. coli function across different Gram-negative species and can actuate using phage RNA polymerase, suggesting that RNA systems acting at the level of transcription are portable. Second, we explore a novel RNA design strategy that uses arrays of tandem and transcriptionally fused RNA regulators to precisely alter regulator concentration from 1 to 8 copies. This provides a simple means to predictably tune output gain across species and does not require access to large regulatory part libraries. Finally, we show RNA arrays can be used to achieve tunable cascading and multiplexing circuits across species, analogous to the motifs used in artificial neural networks.

Competing Interest Statement

The authors have declared no competing interest.

Copyright 
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC-ND 4.0 International license.
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Posted February 25, 2023.
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A portable regulatory RNA array design enables tunable and complex regulation across diverse bacteria
Baiyang Liu, Christian Cuba Samaniego, Matthew R. Bennett, Elisa Franco, James Chappell
bioRxiv 2023.02.24.529951; doi: https://doi.org/10.1101/2023.02.24.529951
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A portable regulatory RNA array design enables tunable and complex regulation across diverse bacteria
Baiyang Liu, Christian Cuba Samaniego, Matthew R. Bennett, Elisa Franco, James Chappell
bioRxiv 2023.02.24.529951; doi: https://doi.org/10.1101/2023.02.24.529951

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