Spectramax plate reader

All L-form strains were cultured in liquid L phase broth (LPB) and solid L phase medium agar (LPMA). LPB consists of a 1:1 mixture of yeast extract malt extract (YEME) and tryptic soy broth supplemented with 10% sucrose (TSBS) and 25 mM MgCl₂. LPMA consists of LPB supplemented with 1.5% agar, 5% horse serum, and 25 mM MgCl₂ (9 ). P-buffer containing sucrose, K₂SO₄, MgCl₂, trace elements, KH₂PO₄, CaCl₂, and N-tris (hydroxymethyl)-methyl-2-aminoethanesulfonic acid (TES) (9 ) was used for transformation and all fusion experiments and was supplemented with 1 mg/mL DNase I (Roche Diagnostics GmbH). The antibiotics apramycin (Duchefa Biochemie) and hygromycin (Duchefa Biochemie) were used for selection and were added at final concentrations of 50 μg/mL and 100 μg/mL, respectively. Growth conditions for all cultures were 30°C in an orbital shaker (New Brunswick Scientific Innova) with 100 rpm for the liquid cultures. Centrifugation (Eppendorf centrifuge 5424) conditions were always 1,000 × g for 10 min (<1 mL) or 30 min (>10 mL), depending on culture volume. The above-mentioned culture conditions and centrifugation settings were applied throughout the study unless mentioned otherwise. All measurements for optical densities (ODs) of samples were done with 200-μL aliquots of culture in a 96-well flat-bottom plate (Sarstedt) using the Tecan Spectramax plate reader.

All L-form strains were cultured in liquid L phase broth (LPB) and solid L phase medium agar (LPMA). LPB consists of a 1:1 mixture of yeast extract malt extract (YEME) and tryptic soy broth supplemented with 10% sucrose (TSBS) and 25 mM MgCl₂. LPMA consists of LPB supplemented with 1.5% agar, 5% horse serum, and 25 mM MgCl₂ (9 ). P-buffer containing sucrose, K₂SO₄, MgCl₂, trace elements, KH₂PO₄, CaCl₂, and N-tris (hydroxymethyl)-methyl-2-aminoethanesulfonic acid (TES) (9 ) was used for transformation and all fusion experiments and was supplemented with 1 mg/mL DNase I (Roche Diagnostics GmbH). The antibiotics apramycin (Duchefa Biochemie) and hygromycin (Duchefa Biochemie) were used for selection and were added at final concentrations of 50 μg/mL and 100 μg/mL, respectively. Growth conditions for all cultures were 30°C in an orbital shaker (New Brunswick Scientific Innova) with 100 rpm for the liquid cultures. Centrifugation (Eppendorf centrifuge 5424) conditions were always 1,000 × g for 10 min (<1 mL) or 30 min (>10 mL), depending on culture volume. The above-mentioned culture conditions and centrifugation settings were applied throughout the study unless mentioned otherwise. All measurements for optical densities (ODs) of samples were done with 200-μL aliquots of culture in a 96-well flat-bottom plate (Sarstedt) using the Tecan Spectramax plate reader.