New suicide vector for gene replacement in yersiniae and other gram-negative bacteria

A suicide vector named pUK4134 was constructed to enlarge the repertoire of vectors available for allelic exchange of mutated sequences in gram-negative bacteria. This plasmid combines the properties of two previously described plasmid vectors, pJM703.1 and pRTP1. pUK4134 is a suicide vector, carrying the origin of replication of the plasmid R6K and thus requiring the product of the pir gene for its stable maintenance. The rpsL gene encoding Escherichia coli ribosomal protein S12 confers streptomycin sensitivity on streptomycin-resistant strains and provides a positive selection for bacteria that have lost the plasmid following allelic exchange. The bla gene provides for selection by Apr. Other features are a unique EcoRV cloning site, oriT of plasmid RK2, and the bacteriophage lambda cos sequence. This vector was successfully used several times to carry out allelic exchange in Yersinia pestis.