A Continuous Cell-Free Protein Synthesis System for Coupled Transcription-Translation1

A continuous cell-free protein synthesis system [Spirin et al. (1988) Science 242, 1162-1164] was modified so as to be suitable for coupled transcription-translation, a process useful for obtaining products of cloned genes or cDNAs. A reaction chamber equipped with an ultrafiltration membrane was newly designed and an HPLC pump was used to supply a low molecular weight substrate solution at a constant rate to the viscous reaction mixture in the chamber. By using an Escherichia coli S30 extract in this modified flow system (1 ml), coupled transcription-translation could be continuously performed for 17 h, the synthesized chloramphenicol acetyltransferase (congruent to 0.1 mg) being subsequently eluted through the chamber's membrane and then purified.