Strain and culture conditions

The following bacterial strains were used in this study: Escherichia coli Top10 (Invitrogen, Carlsbad, CA, USA), DH5α (laboratory stock), M. smegmatis mc²155 (laboratory stock), and M. tuberculosis H37Rv (laboratory stock). E. coli strains were grown at 37 °C in Luria-Bertani (LB) broth or on LB agar plates. Mycobacterial strains were grown at 37 °C in Middlebrook 7H9 broth (Difco, Franklin Lakes, NJ, USA) in 150 ml roller bottles with slow rotation (3 rpm), in 10 ml screw-cap tubes without agitation, or Middlebrook 7H10 agar plates (Difco) supplemented with 0.2% glycerol and 0.05% Tween-80. For growth of M. tuberculosis, the medium was supplemented with 10% albumin-dextrose-sodium chloride complex (ADN)¹⁴. When needed, antibiotics (Sigma) were added to the media at the following concentrations: streptomycin (Sm) 20 µg ml⁻¹, kanamycin (Km) 50 µg ml⁻¹ (E. coli) or 20 µg ml⁻¹ (M. smegmatis and M. tuberculosis), hygromycin (Hyg) 150 µg ml⁻¹ (E. coli) or 50 µg ml⁻¹ (M. smegmatis and M. tuberculosis), anhydrotetracycline (ATc) 50 ng ml⁻¹ (M. smegmatis) or 500 ng ml⁻¹ (M. tuberculosis). Preparation of electrocompetent cells for electroporation and preparation of mycobacterial genomic DNA were performed as previously described¹⁵. M. tuberculosis strains were handled and cultivated in a Biosafety Level 3 Laboratory (BLS3).