title: Digital switching in a biosensor circuit via programmable timing of gene availability.
abstract: Transient delivery of gene circuits is required in many potential applications of synthetic biology, yet the pre-steady-state processes that dominate this delivery route pose major challenges for robust circuit deployment. Here we show that site-specific recombinases can rectify undesired effects by programmable timing of gene availability in multigene circuits. We exemplify the concept with a proportional sensor for endogenous microRNA (miRNA) and show a marked reduction in its ground state leakage due to desynchronization of the circuit's repressor components and their repression target. The new sensors display a dynamic range of up to 1,000-fold compared to 20-fold in the standard configuration. We applied the approach to classify cell types on the basis of miRNA expression profile and measured >200-fold output differential between positively and negatively identified cells. We also showed major improvements in specificity with cytotoxic output. Our study opens new venues in gene circuit design via judicious temporal control of circuits' genetic makeup.
authors.affiliation: Department of Biosystems Science and Engineering, Swiss Federal Institute of Technology Zurich (ETHZ), Basel, Switzerland.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Lapique
authors.affiliation: Department of Biosystems Science and Engineering, Swiss Federal Institute of Technology Zurich (ETHZ), Basel, Switzerland.
authors.forename: Yaakov
authors.initials: Y
authors.lastname: Benenson
mesh_terms: D015374:Biosensing Techniques / Q000379:methods*; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D053263:Gene Regulatory Networks; D005818:Genetic Engineering; D057809:HEK293 Cells; D006801:Humans; D056968:Lac Repressors / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D008164:Luminescent Proteins; D035683:MicroRNAs / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D008957:Models, Genetic; D010957:Plasmids / Q000737:chemistry; D011401:Promoter Regions, Genetic; D012330:RNA, Double-Stranded / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D015398:Signal Transduction; D058615:Synthetic Biology; D013997:Time Factors; D015534:Trans-Activators; D015533:Transcriptional Activation; D014162:Transfection
chemical_list: D029968:Escherichia coli Proteins; D056968:Lac Repressors; C539064:LacI protein, E coli; D008164:Luminescent Proteins; C524940:MIRN21 microRNA, human; D035683:MicroRNAs; D012330:RNA, Double-Stranded; D012333:RNA, Messenger; D015534:Trans-Activators; C413663:fluorescent protein 583