A continuous cell-free protein synthesis system for coupled transcription-translation

J Biochem. 1991 Aug;110(2):166-8. doi: 10.1093/oxfordjournals.jbchem.a123551.

Abstract

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.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Cell-Free System
  • Chloramphenicol O-Acetyltransferase / metabolism
  • Chromatography, Affinity
  • Chromatography, High Pressure Liquid
  • DNA / genetics
  • Escherichia coli / genetics
  • Genes, Bacterial
  • Protein Biosynthesis*
  • Transcription, Genetic*
  • Ultrafiltration

Substances

  • DNA
  • Chloramphenicol O-Acetyltransferase