Sculpting DNA-based synthetic cells through phase separation and phase-targeted activity

Synthetic cells, like their biological counterparts, require internal compartments with distinct chemical and physical properties where different functionalities can be localized. Inspired by membrane -less compartmentalization in biological cells, here, we demonstrate how microphase separation can be used to engineer heteroge-neous cell-like architectures with programmable morphology and compartment-targeted activity. The synthetic cells self-assemble from amphiphilic DNA nanostructures, producing core-shell con-densates due to size-induced de-mixing. Lipid deposition and phase-selective etching are then used to generate a porous pseudo-membrane, a cytoplasm analog, and membrane-less organ-elles. The synthetic cells can sustain RNA synthesis via in vitro tran-scription, leading to cytoplasm and pseudo-membrane expansion caused by an accumulation of the transcript. Our approach exem-plifies how architectural and functional complexity can emerge from a limited number of distinct building blocks, if molecular-scale programmability, emergent biophysical phenomena, and biochem-ical activity are coupled to mimic those observed in live cells.