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. 1986 Nov;168(2):982–989. doi: 10.1128/jb.168.2.982-989.1986

Tn5-mediated integration of the delta-endotoxin gene from Bacillus thuringiensis into the chromosome of root-colonizing pseudomonads.

M G Obukowicz, F J Perlak, K Kusano-Kretzmer, E J Mayer, S L Bolten, L S Watrud
PMCID: PMC213580  PMID: 3023294

Abstract

Gene replacement mediated by Tn5 sequences was used to integrate the Bacillus thuringiensis subsp. kurstaki HD-1 delta-endotoxin gene (tox) into the chromosome of two corn root-colonizing strains of Pseudomonas fluorescens. A Tn5 transposase deletion element containing the tox gene (delta Tn5-tox) was substituted for a Tn5 element previously present in the P. fluorescens chromosome. Two classes of delta Tn5-tox elements were made. The first class encodes kanamycin resistance in addition to the Tox protein, whereas the second class encodes only the Tox protein. Both classes of delta Tn5-tox elements can no longer transpose, owing to a 324-base-pair deletion in the transposase gene of IS50R, minimizing the potential for horizontal gene transfer of the tox gene to other bacterial species. A frameshift mutation in the transposase gene of IS50L was also constructed to eliminate the possibility of suppression or of a spontaneous reversion at the ochre termination codon that would create an active transposase. Expression of the Tox protein in P. fluorescens strains 112-12 and Ps3732-3-7 was demonstrated by an immunological assay (Western blot) and toxicity against larvae of the tobacco hornworm (Manduca sexta).

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

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