{"database":"biostudies-arrayexpress","file_versions":[],"scores":null,"additional":{"omics_type":["Metabolomics","Unknown","Transcriptomics","Genomics","Proteomics"],"submitter":["Jarmila Hnilicova"],"instrument_platform":["NextSeq 500"],"study_type":["RNA-seq of coding RNA"],"organism":["Mycolicibacterium smegmatis"],"species":["Mycolicibacterium smegmatis"],"full_dataset_link":["https://www.ebi.ac.uk/biostudies/studies/E-MTAB-13812"],"description":["CarD is an essential transcription factor for Mycobacterium smegmatis. We identified a new interaction partner of CarD and named it CrsL (MSMEG_5890). To address the genome-wide effect of CrsL, we generated knockdown strains for both carD (MSMEG_6077) and crsL (MSMEG_5890) genes in Mycobacterium smegmatis. We aimed to compare the differentially expressed genes of crsL knockdown and correlate them with the ones of carD knockdown to describe the role of CrsL in transcription."],"repository":["biostudies-arrayexpress"],"sample_protocol":["Sequencing - Pooled barcoded libraries were sequenced in single lane with Illumina NextSeq® 500/550 High Output Kit v2 in 75 bp single end regime at Institute of Molecular Genetics AS CR, Prague, Czech Republic.","Nucleic Acid Extraction - For unbiased quantification, RNA spike-in were added to each sample according to the following rule: 15 ng of RNA spike-in were added per 30 ml culture of OD600~0.5. Cell pellets were vortexed in 240 μl TE (10 mM Tris, pH 8.0; 1 mM EDTA), 60 μl LETS (50 mM Tris–HCl pH 8.0, 500 mM LiCl, 50 mM EDTA pH 8.0, 5% SDS), 600 μl acidic phenol: chloroform (1:1) were added, sonicated for 30 seconds, centrifuged and the aqueous phase was transferred. 600 μl acidic phenol: chloroform (1:1) was added, the samples were vortexed and again, the aqueous phase was transferred. This step was repeated once more. Then, the RNA was precipitated with ethanol, dissolved in double distilled water, DNase treated (TURBO DNA-free Kit, Ambion) and approximately 1 μg of DNase treated RNA was ribodepleted (riboPOOL Kit Pan-Actinobacteria, siTOOLs). The sample integrity was checked using Agilent 2100 Bioanalyzer Pico Chip.","Library Construction - The ribodepleted RNA sample (20-100 ng) was used for library construction according to the NEXTFLEX® Rapid Directional RNA-Seq Kit. The libraries were checked using Agilent 2100 Bioanalyzer Nano Chip.","Growth Protocol - Mycobacterium smegmatis mc2 155 cells (negative control (LK2261), carD/MSMEG_6077 knockdown strain (LK3301) and crsL/MSMEG_5890 knockdown strain (LK3302)) were inoculated to OD600 0.1 and grown at 37°C in Middlebrook 7H9 medium with 0.2% glycerol and 0.05% Tween 80. For exponential phase cells, knockdowns were induced with anhydrotetracycline (100 ng/mL) after 3 h of OD600 0.1 and the strains were cultivated for another 3 h and harvested (OD600 ~0.5). For stationary phase cells, knockdowns were induced with anhydrotetracycline (100 ng/mL) after 8 h of OD600 0.1 and the strains were cultivated for another 16 h to reach early stationary phase (OD600 ~2.5).","Sample Collection - 50 ml exponential phase cells and 25 ml stationary phase cells were pelleted by centrifugation and frozen at -70°C."],"figure_sub":["Organization","MINSEQE Score","Assays and Data","MAGE-TAB Files"],"pubmed_authors":["Mahmoud Shoman","Jarmila Hnilicova","Martin Převorovský"],"additional_accession":[]},"is_claimable":false,"name":"RNA-seq of Mycobacterium smegmatis mc2 155 negative control strain (NC), carD (MSMEG_6077) and crsL (MSMEG_5890) knockdown strains, from exponential and stationary phase cells.","description":"CarD is an essential transcription factor for Mycobacterium smegmatis. We identified a new interaction partner of CarD and named it CrsL (MSMEG_5890). To address the genome-wide effect of CrsL, we generated knockdown strains for both carD (MSMEG_6077) and crsL (MSMEG_5890) genes in Mycobacterium smegmatis. We aimed to compare the differentially expressed genes of crsL knockdown and correlate them with the ones of carD knockdown to describe the role of CrsL in transcription.","dates":{"release":"2025-11-01T00:00:00Z","modification":"2025-11-01T02:01:47.57Z","creation":"2024-02-28T21:18:37.223Z"},"accession":"E-MTAB-13812","cross_references":{"ENA":["ERP157491"],"EFO":["EFO_0002944","EFO_0004170","EFO_0003789","EFO_0005518","EFO_0003738","EFO_0004184"]}}