A suppressor tRNA-mediated feedforward loop eliminates leaky gene expression in bacteria.
Nucleic Acids Res
; 49(5): e25, 2021 03 18.
Article
en En
| MEDLINE
| ID: mdl-33290521
ABSTRACT
Ligand-inducible genetic systems are the mainstay of synthetic biology, allowing gene expression to be controlled by the presence of a small molecule. However, 'leaky' gene expression in the absence of inducer remains a persistent problem. We developed a leak dampener tool that drastically reduces the leak of inducible genetic systems while retaining signal in Escherichia coli. Our system relies on a coherent feedforward loop featuring a suppressor tRNA that enables conditional readthrough of silent non-sense mutations in a regulated gene, and this approach can be applied to any ligand-inducible transcription factor. We demonstrate proof-of-principle of our system with the lactate biosensor LldR and the arabinose biosensor AraC, which displayed a 70-fold and 630-fold change in output after induction of a fluorescence reporter, respectively, without any background subtraction. Application of the tool to an arabinose-inducible mutagenesis plasmid led to a 540-fold change in its output after induction, with leak decreasing to the level of background mutagenesis. This study provides a modular tool for reducing leak and improving the fold-induction within genetic circuits, demonstrated here using two types of biosensors relevant to cancer detection and genetic engineering.
Texto completo:
1
Bases de datos:
MEDLINE
Asunto principal:
ARN de Transferencia
/
Regulación Bacteriana de la Expresión Génica
Idioma:
En
Revista:
Nucleic Acids Res
Año:
2021
Tipo del documento:
Article
País de afiliación:
Estados Unidos