An on-chip capillary electrophoresis-based DNA collection was applied to isolate desired DNA construct from a reaction mixture of DNA-assembly instead of traditional living cell-based screening. The isolated construct was amplified with polymerase chain reaction and subjected to a coupled cell-free transcription/translation system as a template, and the template-dependent protein expression was observed. It was demonstrated that total process of standard recombinant DNA technology was performed in a cell-free condition.