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Cloning and expression in Escherichia coli of the insecticidal δ-endotoxin gene of Bacillus thuringiensis var. israelensis

Cloning and expression in Escherichia coli of the insecticidal δ-endotoxin gene of Bacillus thuringiensis var. israelensis
Cloning and expression in Escherichia coli of the insecticidal δ-endotoxin gene of Bacillus thuringiensis var. israelensis

Recombinant plasmids containing the mosquitocidal δ-endotoxin gene were constructed by inserting HindIII fragments of the Bacillus thuringiensis var. israelensis 72-75 Md plasmid in to the Escherichia coli vector pUC12. Two recombinants producing the 26000 Da δ-endotoxin (pIP173 and pIP174) were identified by screening clones in an E. coli in vitro transcription-translation system. Both recombinants were 12.4 kb chimaeric plasmids comprising pUC12 and a common 9.7 kb HindIII fragment of the B. thuringiensis plasmid. The 26000 Da polypeptide synthesis in vivo from pIP174 transformed into E. coli JM101 was lethal to mosquito larvae and cytotoxic to mosquito cells in vitro. The biological authenticity of the cloned product was further confirmed by demonstrating that the cytotoxicity of the polypeptide was neutralised by antiserum to the authentic δ-endotoxin or by preincubation with excess toxin receptor. Transcription of the recombinant δ-endotoxin gene in E. coli appears to utilise a Bacillus promotor sequence(s) rather than the pUC12 β-galactosidase promotor.

Bacillus thuringiensis var. israelensis, Gene cloning, Insecticide, Mosquito, Plasmid, δ-Endotoxin gene
0014-5793
377-382
Ward, E. S.
b31c0877-8abe-485f-b800-244a9d3cd6cc
Ellar, D. J.
afb465a4-5ccc-4ac8-8dc1-38789156951d
Todd, J. A.
9dd4d014-af5c-4aaf-97ad-d61bc65c84da
Ward, E. S.
b31c0877-8abe-485f-b800-244a9d3cd6cc
Ellar, D. J.
afb465a4-5ccc-4ac8-8dc1-38789156951d
Todd, J. A.
9dd4d014-af5c-4aaf-97ad-d61bc65c84da

Ward, E. S., Ellar, D. J. and Todd, J. A. (1984) Cloning and expression in Escherichia coli of the insecticidal δ-endotoxin gene of Bacillus thuringiensis var. israelensis. FEBS Letters, 175 (2), 377-382. (doi:10.1016/0014-5793(84)80772-3).

Record type: Article

Abstract

Recombinant plasmids containing the mosquitocidal δ-endotoxin gene were constructed by inserting HindIII fragments of the Bacillus thuringiensis var. israelensis 72-75 Md plasmid in to the Escherichia coli vector pUC12. Two recombinants producing the 26000 Da δ-endotoxin (pIP173 and pIP174) were identified by screening clones in an E. coli in vitro transcription-translation system. Both recombinants were 12.4 kb chimaeric plasmids comprising pUC12 and a common 9.7 kb HindIII fragment of the B. thuringiensis plasmid. The 26000 Da polypeptide synthesis in vivo from pIP174 transformed into E. coli JM101 was lethal to mosquito larvae and cytotoxic to mosquito cells in vitro. The biological authenticity of the cloned product was further confirmed by demonstrating that the cytotoxicity of the polypeptide was neutralised by antiserum to the authentic δ-endotoxin or by preincubation with excess toxin receptor. Transcription of the recombinant δ-endotoxin gene in E. coli appears to utilise a Bacillus promotor sequence(s) rather than the pUC12 β-galactosidase promotor.

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More information

Published date: 1 October 1984
Keywords: Bacillus thuringiensis var. israelensis, Gene cloning, Insecticide, Mosquito, Plasmid, δ-Endotoxin gene

Identifiers

Local EPrints ID: 425244
URI: http://eprints.soton.ac.uk/id/eprint/425244
ISSN: 0014-5793
PURE UUID: 49320f98-439f-4e32-bb71-10c4c74f133c
ORCID for E. S. Ward: ORCID iD orcid.org/0000-0003-3232-7238

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Date deposited: 12 Oct 2018 16:30
Last modified: 18 Mar 2024 03:48

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Contributors

Author: E. S. Ward ORCID iD
Author: D. J. Ellar
Author: J. A. Todd

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