An optimized technique for rapid genome modifications of Bacillus subtilis

Abstract

The transformation efficiency of naturally competent Bacillus subtilis cells can be significantly increased using β recombinase binding sequences, as revealed by the results of this study. Plasmids containing different variations of these so called six-site-marker-cassettes were investigated. Furthermore, an optimized protocol for knock-out or knock-in mutations combining the Cre-lox-system and the six-sites is presented, which can be used for multiple genome modifications of B. subtilis.

Keywords: Bacillus subtilis; CAA; CFU; Gene library; Natural competence; OD; PCR; SSS; Six sites; TM; Transformation efficiency; Wild type strain; base pairs; bp; casamino acids; colony forming units; optical density; polymerase chain reaction; six-site-spectinomycin resistance gene; transformation medium.