Transcription termination signal for the cat-86 indicator gene in a Bacillus subtilis promoter-cloning plasmid

Abstract

Plasmid pPL703 is a promoter-cloning plasmid for Bacillus subtilis consisting of the promoter-less cat-86 gene inserted between the EcoRI and BamHI sites of pUB110. The orientation of cat-86 in pPL703 is opposite to that of two major transcript species that occur within the pUB110 vector portion of pPL703. Therefore, transcripts initiated in cloned promoters which activate cat-86 expression presumably must terminate prior to entering the vector portion of pPL703 to permit stable maintenance of promoter-containing derivatives in host cells. We have identified an apparent Rho-independent transcription terminator 35 bp 3' to the cat-86 coding sequence. A restriction fragment spanning the terminator is 90% efficient in terminating transcription in both B. subtilis and Escherichia coli. The structure of the cat-86 transcription termination site is similar to Rho-independent termination sites identified in E. coli.