Mycobacterial Strain Culturing Protocol

The Mtb H37Rv strain (obtained from C. Sassetti) was grown at 37 °C in a minimal medium (Difco Middlebrook 7H9 broth (BD, 271310) supplemented with 0.5% (v/v) glycerol, 0.05% (v/v) tyloxapol (Sigma, T8761), 0.2 g l⁻¹ casamino acids (BD, 223050), and 10% (v/v) OADC (oleic acid, albumin, dextrose and catalase; BD, 212351)). The double-auxotrophic Mtb mc²6206 strain (H37Rv ΔpanCDΔleuCD)^{42 (link)} (obtained from W. Jacobs Jr) was grown in the minimal medium with an additional 50 mg l⁻¹l-leucine (Sigma, L8000) and 24 mg l⁻¹ pantothenic acid (Sigma, P5155). The Msm mc²155 strain (obtained from S. Fortune) was grown in the Middlebrook 7H9 medium supplemented with 0.2% (v/v) glycerol, 0.05% (v/v) Tween-80 (VWR, M126), and 10% (v/v) albumin–dextrose–catalase. Liquid Mtb and Msm cultures were grown at 37 °C in Nalgene sterile square PETG medium bottles with constant agitation. The solid Mtb culture was grown on 7H11 agar (Sigma, M0428) supplemented as described above except for tyloxapol.