Engineering Persistent and Rewritable Genetic Memory in Bacteria through Multiscale Plasmid Competition

Abstract

Genetic memory enables bacteria to record transient stimuli as persistent genetic states, providing a foundation for cellular information storage, sensing, and computation. Recombinase-based memories encode discrete states through rewritable DNA rearrangements, but these states can continue to evolve after stimulus removal as cells proliferate. Here, we show that the long-term stability of reversible recombinase-based memory can be engineered by controlling post-switching dynamics. We identify a temporal hierarchy in which heterogeneous states arising from imperfect recombinase orthogonality are first resolved through intracellular competition between incompatible plasmids and subsequently reshaped by fitness differences between cells carrying alternative states. Reducing DNA copy number accelerates intracellular state resolution and decreases expression-associated fitness costs, while tuning state-specific gene expression balances the relative fitness of alternative memory states. Using these principles, we engineer a reversible memory device that maintains its two states for up to two weeks while retaining rewritability after prolonged maintenance. Our results establish a framework for engineering persistent and rewritable genetic memory through control of intracellular competition and population-level fitness.