This study presents a novel method for lysing coryneform bacteria to facilitate DNA extraction. The researcher tested eight glutamic acid-producing bacterial species that were resistant to standard lysis procedures. The method involves treating bacteria in late-exponential phase with cell wall biosynthesis inhibitors—penicillin G or sencephalin—followed by enzymatic digestion and detergent treatment. Most organisms required 0.5-2 hours of antibiotic pretreatment before becoming sensitive to lytic enzymes. This combination approach proved more effective and convenient than previous methods including freeze-thaw cycles, high glycine concentrations, and sonication. The antibiotic pretreatment weakened cell walls, making subsequent lysis by Kyowa lytic enzyme and lysozyme highly efficient. Complete lysis was confirmed by microscopic observation and clearing of bacterial suspensions. This technique successfully enabled DNA isolation suitable for homology experiments from multiple coryneform species, providing a practical improvement over existing time-consuming and inefficient procedures.

Key findings

• Pretreatment with β-lactam antibiotics (penicillin G or sencephalin) that inhibit cell wall biosynthesis significantly increased the sensitivity of coryneform bacteria to enzymatic lysis
• The combination of antibiotic pretreatment followed by lytic enzymes and SDS detergent treatment produced complete bacterial lysis more efficiently than previous methods
• This method successfully facilitated DNA extraction from eight different glutamic acid-producing bacterial species, including Corynebacterium glutamicum, Brevibacterium species, and Nocardia erythropolis
• Optimal antibiotic pretreatment duration was 0.5 hours for most organisms, with longer exposure sometimes causing unwanted spontaneous lysis