Journal
NATURE BIOTECHNOLOGY
Volume 38, Issue 8, Pages 962-+Publisher
NATURE PORTFOLIO
DOI: 10.1038/s41587-020-0468-5
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Funding
- National Institute of Health P50 grant [P50-GM098792]
- Office of Naval Research Multidisciplinary University Research Initiatives Program [N00014-13-1-0074]
- Defense Agency Research Projects Agency Synergistic Discovery and Design (SD2) [FA8750-17-C-0229]
- National Science Foundation Semiconductor Synthetic Biology for Information Processing and Storage Technologies (SemiSynBio) program [CCF-1807575]
- National Institutes of Health (NIH) [R01EB021908]
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Bacteroides thetaiotaomicron is a human-associated bacterium that holds promise for delivery of therapies in the gut microbiome(1). Therapeutic bacteria would benefit from the ability to turn on different programs of gene expression in response to conditions inside and outside of the gut; however, the availability of regulatory parts, and methods to combine them, have been limited in B. thetaiotaomicron(2-5). We report implementation of Cello circuit design automation software(6) for this species. First, we characterize a set of genome-integrated NOT/NOR gates based on single guide RNAs (CRISPR-dCas9) to inform a Bt user constraint file (UCF) for Cello. Then, logic circuits are designed to integrate sensors that respond to bile acid and anhydrotetracycline (aTc), including one created to distinguish between environments associated with bioproduction, the human gut, and after release. This circuit was found to be stable under laboratory conditions for at least 12 days and to function in bacteria associated with a primary colonic epithelial monolayer in an in vitro human gut model system. A platform for genetic circuit design in a human gut commensal bacterium brings microbiome therapies that respond to gastrointestinal signals a step closer.
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