Mycobacterium smegmatis Growth Media and Reagents

As previously described13 (link), the 7H9 liquid culture medium for M. smegmatis strains consisted of Middlebrook 7H9 medium (Becton Dickinson, Franklin Lakes, New Jersey, U.S.) supplemented with 10% ADS (5% (w/v) bovine serum albumin fraction V, 2% (w/v) dextrose and 8.1% (w/v) NaCl), 0.5% (v/v) glycerol, and 0.05% (v/v) Tween80. 7H10 media containing Middlebrook 7H10 medium (Becton Dickinson, Franklin Lakes, New Jersey, U.S.), 10% ADS, and 0.5% (v/v) glycerol was used for solid culture to examine growth status. Hygromycin (Hyg) was purchased from GenView, erythromycin (EM) from Merck, hydrogen peroxide (H₂O₂) and kanamycin (Kan) from Sigma. Restriction enzymes such as Van91I, AlwNI, MfeI, and PacI were purchased from Fermentas. T4 DNA ligase and Q5 DNA polymerase were purchased from New England Biolabs.