New mini-Tn5 derivatives for insertion mutagenesis and genetic engineering in Gram-negative bacteria

Abstract

Five mini-Tn5 derivatives encoding resistance to Km, Cm, Gm, Tc, and Sm, coupled with the polylinker of the pBluescriptII plasmid, were constructed. These derivatives are carried by an ampicillin-resistant plasmid that has a conditional origin of replication from plasmid R6K and origin of conjugal transfer from the broad host range plasmid RP4. The new vectors are smaller than those previously described and possess numerous unique restriction sites inside the minitransposons for gene cloning in addition to SfiI and NotI sites found in their predecessors.Key words: R6K γ-origin, suicide vectors, RP4 oriT, chromosomal insertion.