A versatile plasmid system for the study of prokaryotic transcription signals in Escherichia coli

For comparing the relative efficiencies of Escherichia coli promoters, a modified plasmid system, pKO-2 and pKM-2, has been constructed using short synthetic DNA fragments. The new vectors were derived from the plasmids pKO-1 and pKM-1. The plasmids contain seven clustered unique restriction sites which can be used for promoter insertions. Also, three adjacent stop codons were introduced to abort any undesired translational initiation from various upstream origins. The DNA sequence of any insert in pKO-2 and pKM-2 can be determined rapidly by the supercoiled plasmid DNA sequencing method using a single oligonucleotide primer. The plasmid pKM-2 is especially suitable for the cloning and sequence determination of strong promoters.