Construction of plasmid vectors bearing a NotI-expression cassette based on the lac promoter

Abstract

We have constructed two plasmid vectors for cloning and expression of DNA fragments controlled by the lac promoter as a NotI-expression cassette. Whereas plasmid pSJ33 allows mobilization of the expression cassette into a wide variety of Gram-negative bacteria by RP4-mediated conjugation, the low-copy-number plasmid pSJP18Not facilitates cloning and expression in Escherichia coli when high gene dosage may be detrimental. In addition to their suitable cloning features (e.g. multiple cloning site, lacZa fragment, compatible with ColE1-derived vectors), these plasmids are particularly useful as auxiliary vectors for cloning of the expression cassettes at the NotI site of mini-transposon elements [1, 2] and their eventual stable insertion into the host chromosome.