characteristics.text: ERC000011
characteristics.characteristic: ENA-CHECKLIST
characteristics.text: 2022-12-31T00:58:39Z
characteristics.characteristic: ENA-FIRST-PUBLIC
characteristics.text: 2022-12-31T00:58:39Z
characteristics.characteristic: ENA-LAST-UPDATE
characteristics.text: SAMEA111479729
characteristics.characteristic: External Id
characteristics.text: 1. Institut Pasteur, Universite Paris Cite, Plate-forme Technologique Biomics, F-75015 Paris, France 2. Institut Pasteur, Universite Paris Cite, Bioinformatics and Biostatistics Hub, F-75015 Paris, France
characteristics.characteristic: INSDC center name
characteristics.text: 2022-12-31T00:58:39Z
characteristics.characteristic: INSDC first public
characteristics.text: 2022-12-31T00:58:39Z
characteristics.characteristic: INSDC last update
characteristics.text: public
characteristics.characteristic: INSDC status
characteristics.text: ERS13573764
characteristics.characteristic: SRA accession
characteristics.text: E-MTAB-12307:34509_H2
characteristics.characteristic: Submitter Id
characteristics.text: ArrayExpress
characteristics.characteristic: broker name
characteristics.text: TMC 102
characteristics.characteristic: cell_line
characteristics.text: Protocols: To isolate resistant mutants, exponentially growing cultures were concentrated, and about 1010 cells were plated onto 7H10 agar plates, containing different concentrations of a given drug, ranging from 4X- to 20X-MIC. Plates were incubated at 37 °C for 4 weeks. The resistant phenotype was validated by assessing the MIC at least twice on the isolated colonies. Resistant colonies were reinoculated in the presence of the compound (8X-MIC) and 10 mL of exponentially growing cultures were used for genomic DNA isolation. In short, each cell pellet was heat-inactivated at 80 ˚C for 1, washed and resuspended in 450 µL of bug lysis solution (25 mM Tris-HCl pH 7.9; 10 mM EDTA 50 mM glucose). 50 µL of lysozyme (10 mg/mL) were added and the cell suspension was incubated 37 ˚C overnight. Next, 100 µL of sodium dodecyl sulfate (10 %) and 50 µL of proteinase K (10 mg/mL) were added, mixed gently by inversion, and incubated at 55 ˚C for 30 min. Next, 200 µL of sodium chloride (5 M) were added and mixed gently by inversion, followed by 160 µL of preheated cetrimide saline solution (4 %), mixed and incubated at 65˚C for 10 min. Genomic DNA was extracted twice, adding equal volume (1 mL) of chloroform: isoamyl alcohol (24:1), mixing by inversion, and centrifuging for 5 min at 12000 RPM. The aqueous layer was transferred to a clean tube (800 µL) and 560 µL (0.7X volumes) of isopropanol were added at room temperature, mixed by inversion until DNA precipitation. DNA was collected by centrifugation for 10 min and the pellet was washed with 70 % ethanol, dried and resuspended in 100 µL of TE buffer. Libraries were built using a TruSeq DNA PCR-Free Low Throughput Library Prep Kit (Illumina, USA) following the manufacturer’s protocol. Quality control was performed on an Agilent BioAnalyzer.
characteristics.characteristic: description
characteristics.text: wild type genotype
characteristics.characteristic: genotype
characteristics.text: logarithmic phase culture
characteristics.characteristic: growth condition
characteristics.text: Mycobacterium tuberculosis H37Rv
characteristics.characteristic: organism
characteristics.text: Mycobacterium tuberculosis H37Rv
characteristics.characteristic: organism
characteristics.text: whole organism
characteristics.characteristic: organism part
characteristics.text: Mycobacterium tuberculosis H37Rv
characteristics.characteristic: scientific_name
characteristics.text: 34509_H2
characteristics.characteristic: title