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: SAMEA111479758
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: ERS13573793
characteristics.characteristic: SRA accession
characteristics.text: E-MTAB-12306:34509_3
characteristics.characteristic: Submitter Id
characteristics.text: ArrayExpress
characteristics.characteristic: broker name
characteristics.text: TMC 102
characteristics.characteristic: cell_line
characteristics.text: Protocols: Primary cultures were grown up to OD600nm 0.5/0.6 and diluted to OD600nm 0.025 using pre-warmed Middlebrook 7H9 broth. Secondary cultures were grown up to OD600nm 0.25 and drugs were added at a concentration equal to 10 fold the minimal inhibitory concentration for 6 hours. Cells were collected by centrifugation for 10 minutes and frozen at -80 °C, before RNA isolation. RNA was extracted by the hot phenol method. In detail, bacterial cell pellets were processed in 400 µL of hot lysis buffer (50 mM Tris pH 8.0, 1 mM EDTA, 1% SDS) and then mixed in another screw cap tube containing 400 µL of hot phenol (pH 6.6) and 400 µL of acidified glass beads (425-600-µm diameter). Cells were incubated at 65 °C for 15 minutes with intermittent vortexing every 5 minutes. Samples were centrifuged at 10,000 rpm for 15 minutes at 4 °C. The aqueous phase was separated into another fresh screw-capped microcentrifuge tube, an equal amount of Trizol and 100 µl of chloroform were added and mixed gently by inversion. The samples were again centrifuged at 10,000 rpm for 10 minutes at 4 °C, the aqueous phase separated into a fresh tube, and the RNA was precipitated with 0.5 M LiCl and three volumes of cold 100% ethanol. Samples were mixed gently by inversion, and then precipitated for 2 hours at -20 °C. Samples were centrifuged at 13,500 rpm for 35 minutes at 4 °C and RNA was washed with 80% ethanol. RNA samples were air-dried for 30 minutes and resuspended in 20 µL of nuclease-free water. Finally, DNA was depleted by TURBO DNase treatment (Ambion), according to the manufacturer's instructions. The RNA integrity index was examined on the Agilent Bioanalyzer RNA 6000 Pico kit to confirm the quality and integrity of the RNA for RNAseq purposes. For RTq-PCR, the concentration was measured via NanoDrop 2000/2000c spectrophotometer. rRNA was depleted using the QIAseq FastSelect 5S/16S/23S kit for bacteria (Qiagen),Ribosoma vailableaccording to manufacturer’s recommendations. The libraries were prepared using the TruSeq Stranded mRNA Library Preparation kit (Illumina) according to manufacturer’s protocol https://support.illumina.com/content/dam/illumina-support/documents/documentation/chemistry_documentation/samplepreps_truseq/truseq-stranded-mrna-workflow/truseq-stranded-mrna-workflow-checklist-1000000040600-00.pdf. The quality, integrity and amount of the libraries were assayed with the Agilent 2100 Bioanalyzer and Qubit fluorimeter.
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_3
characteristics.characteristic: title