E. coli-based in vitro transcription/translation: in vivo-specific synthesis rates and high yields in a batch system.
A highly efficient Escherichia coli-based, batch in vitro protein synthesis system using circular plasmid DNA is described. Compared to a presently available commercial kit, this improved system produced several hundredfold greater yields of the rDNA human protein thrombopoietin (ca. 450 micrograms/mL). The system is capable of obtaining specific synthesis rates similar to those in vivo, approximately a 1000-fold increase compared to the original methods previously described. It compares favorably in rates and yields to the recently published semicontinuous methods but with the convenience of a true batch system.
Patnaik
R
R
Genentech, South San Francisco, CA, USA.
Swartz
J R
JR
D001426:Bacterial Proteins / Q000096:biosynthesis; D002458:Cell Fractionation; D004269:DNA, Bacterial / Q000302:isolation & purification; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics*; D006801:Humans; D010957:Plasmids / Q000235:genetics; D014176:Protein Biosynthesis; D011994:Recombinant Proteins / Q000096:biosynthesis*; D012967:Sodium Dodecyl Sulfate; D013698:Templates, Genetic; D013926:Thrombopoietin / Q000096:biosynthesis / Q000235:genetics; D014158:Transcription, Genetic
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D011994:Recombinant Proteins; D012967:Sodium Dodecyl Sulfate; D013926:Thrombopoietin