Digital switching in a biosensor circuit via programmable timing of gene availability

Area of Science:

• Synthetic biology
• Molecular and cell biology
• Biotechnology

Background:

• Transient gene delivery is crucial for synthetic biology applications.
• Pre-steady-state dynamics challenge robust gene circuit function.
• Controlling gene availability is key for predictable circuit behavior.

Purpose of the Study:

• To demonstrate site-specific recombinases for programmable temporal control in multigene circuits.
• To enhance the performance of microRNA (miRNA) sensors.
• To enable cell type classification using miRNA expression profiles.

Main Methods:

• Utilized site-specific recombinases to program gene availability timing.
• Developed a proportional sensor for endogenous microRNA (miRNA).
• Assessed sensor performance, including dynamic range and specificity, and applied it for cell classification.

Main Results:

• Achieved significant reduction in sensor ground state leakage through component desynchronization.
• Demonstrated a 1,000-fold dynamic range for miRNA sensors, a substantial improvement over standard configurations.
• Successfully classified cell types based on miRNA profiles with >200-fold output differential and improved specificity for cytotoxic outputs.

Conclusions:

• Site-specific recombinases offer a powerful strategy for temporal control in gene circuit design.
• This method enhances sensor performance and enables novel applications like cell type classification.
• Judicious temporal control of genetic makeup opens new possibilities in synthetic biology.