DNA affects the phenotype of fuel-dependent coacervate droplets

Synthetic cells emulate fundamental biological behaviors, such as growth, metabolism, and evolution, under non-equilibrium conditions, but have lacked genotype-driven selection, which is essential for Darwinian evolution. Here, we introduce short DNA sequences as genotypes into fuel-dependent, peptide-RNA-based coacervate droplets. By sequencing random DNA libraries, we identify sequence-dependent partitioning into droplets, revealing strong preferences for guanine-rich and adenine-rich motifs. Adenine-rich sequences shortened the droplet lifetimes through RNA hybridization, whereas guanine-rich sequences kinetically trapped droplets via peptide interactions, altering dissolution rates and morphology. This study demonstrates genotype-phenotype coupling in synthetic cells, establishing essential design principles for achieving Darwinian evolution in minimal protocellular systems.