The past 20 years have witnessed enormous progress in synthetic biology in the development of engineered cells for diverse applications, including biomanufacturing, materials fabrication, and potential therapeutics and diagnostics. However, it still remains a major challenge to maintain long-term performance of synthetic gene circuits, due to the emergence of mutants that lose circuit function. Here, we highlight major vulnerabilities of synthetic gene circuits resulting in circuit failure and mutant escape. We also discuss engineering strategies to enhance long-term circuit stability and performance. These approaches can be divided into two strategies: the suppression of the emergence of mutants and the suppression of their relative fitness if mutants do emerge. We anticipate that mechanistic understanding of the modes of circuit failure will facilitate future efforts to design evolutionarily robust synthetic biology-inspired applications.
Keywords: design principles; gene circuits; metabolic burden.