Cesium-Resistant Mutant Generation

Cs⁺-sensitive strains and their Cs⁺-resistant revertants were generated via chemical mutagenesis with ethyl methane sulfonate (EMA), as previously reported (Koretsune et al., 2022 (link)). The Cs⁺-sensitive mutants obtained here were designated Mut5 and Mut7. Single colony isolation was performed for the isolated Mut5 and Mut7 on NC agar medium (pH 8). The colonies were inoculated into 2 ml NC medium (pH 8.0) and reciprocally shake-cultured at 200 rpm at 30°C for 18 h. The culture (100 μl) was independently plated on NC agar medium (pH 8) containing 200 mM or 400 mM CsCl to obtain spontaneous mutants whose cesium resistance was restored. Their Cs⁺-resistant revertants obtained here were designated Mut5R and Mut7R.