We recently developed a cell-free system for propagating circular DNAs by reconstituting the whole replication cycle of the Escherichia coli chromosome using 26 purified proteins. The propagation proceeds exponentially, even from a single DNA molecule, with extremely high fidelity. We further combined a multiple-DNA fragment assembly technique to generate large circular DNA, which provides a powerful tool for the cell-free synthesis of genome-length DNA without biological cloning. In this symposium, I‘ll talk about our cell-free genome synthesis technology as a tool for drug discovery and deployment.