A coupled in vitro transcription-translation system for the exclusive synthesis of polypeptides expressed from the T7 promoter

AbstractA coupled transcription—translation in vitro system has been developed in Escherichia coli specifically for the expression of genes under the exclusive control of the T7 promoter, This system consists of an E. coli crude extract (prepared from cells containing endogenous T7 RNA polymerase), rifampicin (an E. coli RNA polymerase inhibitor) and a labelled amino acid. When primed with a plasmid template encoding the target gene under exclusive control of the T7 promoter, this system has the capability to synthesize relatively large amounts of a unique, labelled polypeptide. This paper describes the characteristics and use of such a T7 RNA polymerase/T7-promoter specific in vitro system