A versatile plasmid vector system for the regulated expression of genes in Escherichia coli

A series of plasmid expression vectors, which support the regulated and efficient expression of genes in Escherichia coli, have been constructed. The vectors consist of a DNA replication origin cassette, a promoter cassette, an efficient ribosome binding site together with a polylinker region and a lacZ gene. Several types of replication origins and promoter sequences are each available on cassettes. Fusion of the 5' TG dinucleotide of the gene under consideration to the A nucleotide, present on the vector, results in an ATG start codon and allows, in combination with the plasmid-borne ribosome binding site, the efficient expression of the cloned gene. Additionally, a second fusion of the gene at its 3' end with the lacZ gene, which is available in all three reading frames relative to the polylinker region, allows rapid selection of the correctly fused genes. As an example of the cloning of a regulatory gene, this vector system was used for the expression of the dnaA gene, of Escherichia coli, the initiator protein for DNA replication.