title: Tribromsalan 3
organism: Mycobacterium tuberculosis
description: Protocols: Mycobacterium tuberculosis H37Rv was cultured in Middlebrook 7H9 broth (with 0.05% Tween 80 and 10% albumin-dextrose-catalase). Drugs were prepared as 10 mM stock solutions in DMSO and stored at -20â€™C in aliquots to avoid freeze-thawing. Log phase M. tuberculosis cultures (OD 0.2-0.4, 4-day shaking at 150 rpm) were treated with 10x MIC niclosamide (final concentration 30.9 uM) or tribromsalan (final concentration 681.6 uM) for 4 hours, or vehicle control only (sterile DMSO, final concentration 0.75%), from three independent biological replicates on different days. Following drug exposure, mycobacterial RNA was extracted using an established GTC/TRIzol method (Waddell et al., Tuberculosis 2004; 84:263-274). The resulting mycobacterial RNA was DNase-treated and purified using the RNeasy spin columns (Qiagen, Hilden, Germany). RNA yield and quality were assessed by NanoDrop One (Thermo Scientific, Waltham, MA, USA) and Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA, USA). RNA yields from all samples were similar and of good quality. Mycobacterial ribosomal RNA was depleted using RiboCop META kit (Lexogen, Vienna, Austria) resulting in mRNA suitable for RNA sequencing. Strand-specific RNAseq libraries were prepared using the NEBNext Ultra II directional RNA library prep kit (New England Biolabs, Ipswitch, MA, USA) and pooled in-house.
attributes.tag: strain
attributes.value: H37Rv
attributes.tag: isolate
attributes.value: not applicable
attributes.tag: replicate
attributes.value: Biological Replicate 3
attributes.tag: organism
attributes.value: Mycobacterium tuberculosis
attributes.tag: organism
attributes.value: Mycobacterium tuberculosis
attributes.tag: growth condition
attributes.value: Log phase in vitro
attributes.tag: collection date
attributes.value: not collected
attributes.tag: scientific_name
attributes.value: Mycobacterium tuberculosis
attributes.tag: organism part
attributes.value: whole organism
attributes.tag: geographic location (country and/or sea)
attributes.value: not collected
attributes.tag: ENA-LAST-UPDATE
attributes.value: 2024-02-02