Datensatz

Zusammenfassung

The suicide plasmid pfdA31−Tn5 was constructed to mutagenize Erwinia amylovora and Escherichia coli strains by electorporation. This vector carries the bacteriophage fd replication origin, a beta−lactamase gene and the transposon Tn5. For propagation the plasmid depends on host cells producing fd gene−2 protein. Electroporation of E.amylovora or E.coli cells with plasmid pfdA31−Tn5 yielded more than 10(4) transposition events per micrograms DNA. We have produced and characterized transposon mutants of E.amylovora affecting either galactose metabolism or the synthesis of the phytotoxin (L)−2,5−dihydrophenylalanine. A Tn5−insertion in a gene, involved in exopolysaccharide synthesis of E.amylovora strain Ea7/74, was subcloned into vector pfdA31 and used to mutagenize E.amylovora strain Ea1/79 by site−directed recombination