An improved plasmid for the isolation and analysis of Streptomyces promoters

Abstract

A Streptomyces promoter-probe plasmid vector was developed to isolate and characterize nucleotide sequences involved in transcriptional regulation. This pCLL34 vector was derived from plasmid pARC1 carrying the SLP1.2 origin of replication [Horinouchi and Beppu, J. Bacteriol. 162 (1985) 406-412]. Important features of the new vector include moderate host range, low copy number, simple identification of promoter activity by brown pigment production, an upstream transcriptional terminator, and a polylinker sequence containing a unique BamHI site for flexible cloning and fragment reisolation. The vector also provides the capability to directly determine the sequence of promoter-active inserts by dideoxy chain-termination sequencing methods using double-stranded plasmid templates and the appropriate oligodeoxynucleotide primers.