Synthetic ratio computation for programming population composition and multicellular morphology

Abstract

Recent advancements in genetic engineering have provided diverse tools for artificially synthesizing population diversity in both prokaryotic and eukaryotic systems. However, achieving precise control over the ratios of multiple cell types within a population derived from a single founder remains a significant challenge. In this study, we introduce a suite of recombinase-mediated genetic devices designed to achieve accurate population ratio control, enabling the distribution of distinct functionalities across multiple cell types. We systematically evaluated key parameters influencing recombination efficiency and developed data-driven models to reliably predict binary differentiation outcomes. Using these devices, we implemented parallel and series circuit topologies to create user-defined, complex cell fate branching programs. These branching devices facilitated the autonomous differentiation of precision fermentation consortia from a single founder strain, optimizing cell-type ratios for applications such as pigmentation and cellulose degradation. Beyond biomanufacturing, we engineered multicellular aggregates with genetically encoded morphologies by coordinating self-organization through cell adhesion molecules (CAMs). Our work provides a comprehensive characterization of recombinase-based cell fate branching mechanisms and introduces a novel approach for the bottom-up, high-resolution construction of synthetic consortia and multicellular assemblies.