Recent advancements in genetic engineering have provided diverse tools for artificially synthesizing population diversity in both prokaryotic and eukaryotic systems. However, accurately controlling the ratios of multiple cell types within a population derived from a single founder remains a significant challenge. In this study, we present a suite of recombinase-mediated genetic devices designed to achieve accurate population ratio control, enabling distinct functionalities to be distributed across multiple cell types. Key parameters influencing recombination efficiency were systematically evaluated, and data-driven models were developed 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 enabled 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 constructing synthetic consortia and multicellular assemblies.Competing Interest StatementT-C.T. is a cofounder and equity holder of Anthology. T.K.L. is a cofounder of Senti Biosciences, Synlogic, Engine Biosciences, Tango Therapeutics, Corvium, BiomX, Eligo Biosciences, Bota.Bio, Avendesora and NE47Bio. T.K.L. also holds financial interests in nest.bio, Armata, IndieBio, MedicusTek, Quark Biosciences, Personal Genomics, Thryve, Lexent Bio, March Therapeutics, Serotiny, Avendesora and Pulmobiotics. G.M.C disclosures can be found on the following webpage: https://arep.med.harvard.edu/gmc/tech.html. C.Z. is a cofounder and equity holder of Shenzhen PAM2L Biotechnologies Co., Ltd....

Synthetic ratio computation for programming population composition and multicellular morphology

10.1101/2024.11.26.624747