Expansion of Insecticidal Host Range of Bacillus Thuringiensis by in vivo Genetic Recombination

We describe a novel approach for the insertion of an insecticidal toxin gene into a resident plasmid in Bacillus thuringiensis (Bt). A gene encoding a coleopteran–specific toxin was cloned within a fragment of IS232 and inserted into a plasmid thermosensitive for replication in Bt. The plasmid was used to transform a Bt strain toxic to lepidoptera, and the transformants were then selected at non–permissive temperature for clones in which the vector had integrated into a copy of IS232 present on a resident plasmid. A second recombination event was selected such that the vector was eliminated and the newly introduced toxin gene was conserved. The resulting strain contained only DNA of Bt origin, and displayed insecticidal activity against both lepidoptera and coleoptera.