An optimized technique for rapid genome modifications of Bacillus subtilis |
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| Authors: | Jana Kumpfmü ller,Johannes Kabisch,Thomas Schweder |
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| Affiliation: | 1. Institute of Pharmacy, Dept. of Pharmaceutical Biotechnology, Ernst-Moritz-Arndt-University, 17487 Greifswald, Germany;2. Institute of Biochemistry, Dept. of Biotechnology & Enzyme Catalysis, Ernst-Moritz-Arndt-University, 17487 Greifswald, Germany |
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| 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. |
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| Keywords: | PCR, polymerase chain reaction bp, base pairs SSS, six-site-spectinomycin resistance gene TM, transformation medium CAA, casamino acids CFU, colony forming units OD, optical density |
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