# SUPER: Upcycling Genetic Parts for Precise Gene Expression Control, Leakage Minimization, and Genetic Circuit Stability

## TL;DR

SUPER is a new platform that improves gene control by using small RNA to enhance biosensors and stabilize genetic circuits without altering target genes.

## Contribution

SUPER introduces a modular RNA-based system for upcycling genetic devices, enabling precise control and stability in synthetic circuits.

## Key findings

- SUPER increases biosensor dynamic range up to 22,018.9-fold and performance by 1011%.
- SUPER enables stable kill switch functionality for over 30 days in microbial co-culture.
- SUPER integrates with environmental sensors to create multi-input kill switches.

## Abstract

A long‐standing goal of synthetic biology is to reprogram cells by rewiring genetic parts. Despite the expanding library of genetic parts, construction of integrated synthetic circuits with desired specifications remains challenging in part due to intricate dependence on sequence contexts, where unexpected narrow dynamic ranges and leaky expression can plague system performance. To provide an alternative approach to the screening process of iterative design‐build‐test cycles, SUPER (
S
ynthetic
U
pcycling
P
latform for
E
ngineering
R
egulators), a modular platform for upcycling genetic devices is introduced. Inspired by antagonistic regulation mechanisms, SUPER employs small RNA as an add‐on controller to modulate gene expression patterns without genetic modification of target regulators. SUPER not only enhances the performance of RNA‐, chemical‐, temperature‐, and protein‐responsive regulators up to 1011%, but also allows to cover an expanded dynamic range up to 22 018.9‐fold. This enhanced control can provide genetic circuit stability, particularly under strong selective pressures, as demonstrated with a Holin‐expressing kill switch integrated with SUPER, maintaining stable functionality for over 30 days. Finally, SUPER combines with an environmental sensor, TlpA36, functioning as a chemical‐ and temperature‐responsive 2‐input kill switch. Featuring straightforward design, minimal cellular burden, and expanded tunability, SUPER provides a systematic upcycling framework for genetic circuit construction in biotechnology.

To precisely modulate natural and synthetic regulatory parts, SUPER is introduced as a sophisticated and tunable controller of genetic devices without requiring sequence modification on the target. SUPER enhances dynamic range of RNA‐, chemical‐, temperature‐, and protein‐responsive biosensors up to 1011%, and converts biosensors to cover an expanded dynamic range up to 22 018.9‐fold. This stringent control with SUPER results in long‐term genetic stability of synthetic kill switches up to 30 days in a microbial co‐culture setting, suggesting a broad range of potential applications in bioengineering and biotherapeutics.