An on-chip capillary electrophoresis-based DNA collection was applied to the isolation of target DNA species from a DNA mixture generated by a polymerase chain reaction (PCR), whose starting material was a ligation mixture of an insert and an expression vector. The collected DNA was then amplified by PCR and properly worked as template DNA in a coupled cell-free transcription/translation system. These results demonstrateed that total operation in standard genetic engineering can be performed in a cell-free condition.