An efficient gene deletion system for Bacillus thuringiensis

Abstract: It is not easy to manipulate biosynthetic genes of Bacillus thuringiensis since there is a powerful methyl-specific restriction system in this microorganism. In this study, a PCR-based system was used to delete polyphosphate kinase gene (ppk) of Bacillus thuringiensis israelensis (Bti) by replacing the wild-type gene with a cassette containing the apramycin resistance gene as selectable marker. λ-Red was used to promote recombination in Escherichia coli between a PCR-amplified apramycin resistance cassette (linear deletion cassette selectable in E. coli and Bti) and Bti DNA on a plasmid. The isolated mutant plasmid was transferred to Bti by conjugation. Double cross-over transformants were screened for their antibiotic resistance and the mutation was proven by PCR, southern blot hybridization and RT-PCR. The described method, which uses the advantage of quick plasmid construction in E. coli and simple transformation of linear deletion cassette, is very useful to delete entire gene/genes of Bti without any polar effects on genes transcriptionally downstream.

Angaben zum Objekt

• Standort: Deutsche Nationalbibliothek Frankfurt am Main
• Umfang: Online-Ressource
• Sprache: Englisch

Verweise und Beziehungen

• Erschienen in: An efficient gene deletion system for Bacillus thuringiensis ; volume:68 ; number:3 ; year:2013 ; pages:358-364 ; extent:7 ; Biologia ; 68, Heft 3 (2013), 358-364 (gesamt 7)

Beteiligte, Orts- und Zeitangaben

• Urheber: Doruk, Tugrul ; Gedik, Sedef

Weitere Informationen

• DOI: 10.2478/s11756-013-0184-4
• URN: urn:nbn:de:101:1-2409231603495.650085287758
• Rechteinformation: Open Access; Der Zugriff auf das Objekt ist unbeschränkt möglich.
• Letzte Aktualisierung: 15.08.2025, 07:36 MESZ