Release Date: 26 June 2018

• Martin Převorovský

  Martin Převorovský

  Email: martin.prevorovsky@natur.cuni.cz
  Phone: (+420) 221 95 1769
  Role: consultant;data analyst;data coder
  Affiliation: Department of Cell Biology, Charles University, Faculty of Science
  ¹
• Olga Ramaniuk

  Olga Ramaniuk

  Email: ramaniuk@biomed.cas.cz
  Phone: (+420) 296 443 406
  Role: data analyst;experiment performer;investigator;submitter
  Affiliation: Laboratory of Microbial Genetics and Gene Expression, Institute of Microbiology, v.v.i. AS CR
  ²
• Libor Krásný

  Libor Krásný

  Email: krasny@biomed.cas.cz
  Phone: (+420) 296 443 208
  Role: consultant;curator;submitter
  Affiliation: Laboratory of Microbial Genetics and Gene Expression, Institute of Microbiology, v.v.i. AS CR
  ³

• ¹ Department of Cell Biology, Charles University, Faculty of Science
  Address: Viničná 7, 128 43 Praha 2
  
• ² Laboratory of Microbial Genetics and Gene Expression, Institute of Microbiology, v.v.i. AS CR
  Address: Videnska 1083, 14220 Prague, Czech Republic
  
• ³ Laboratory of Microbial Genetics and Gene Expression, Institute of Microbiology, v.v.i. AS CR
  Address: Videnska 1083, 14220 Prague, Czech Republic
  

Protocols show table

Name	Type	Description	Hardware	Software
P-MTAB-70319	growth protocol	Bacillus subtilis wt, ∆sigI-rsgI and ∆rsgI strains were inoculated from single colony to 10 ml of LB medium. Cultures were grown overnight at 37°C. Next day, the cultures were inoculated into 20 ml of LB medium (RT) to OD600∼0.01 and grown at 37°C and 52°C, respectively. Cells were harvested in exponential phase (OD600∼0.5). Three ml of culture were immediately treated with a double volume of RNAprotect Bacteria reagent (QIAGEN) for 5 min at RT to prevent degradation of RNA. Cells were pelleted and frozen immediately. In all steps of cultivation (with the exception of the last step of cultivation for total RNA isolation) medium for ∆sigI-rsgI strain was supplemented with spectinomycin 100 μg/ml; medium for ∆rsgI strain was supplemented with linkomycin 12.5 μg/ml and erythromycin 0.5 μg/ml. The experiment was repeated three times.
P-MTAB-70320	nucleic acid extraction protocol	RNA was extracted using RNeasy Mini Kit 50 (QIAGEN) according to the manufacturer’s protocol. Finally, RNA was treated twice with DNase (TURBO DNA-free Kit, Ambion).
P-MTAB-70321	nucleic acid library construction protocol	Two micrograms of total RNA were rRNA-depleted with RiboMinus Transcriptome Isolation Kit, bacteria (Invitrogen). A strand-specific library was prepared for each sample with Illumina compatible NEXTflex Rapid Directional RNA-Seq Kit (Bioo Scientific) according to manufacturer’s protocol.
P-MTAB-70322	nucleic acid sequencing protocol	Transcriptome profiling with RNA-seq was performed at EMBL Genomics Core Facility (Heidelberg, Germany). Pooled barcoded library (6 samples in biological triplicates) was sequenced in a single lane at Illumina HiSeq 2000 (50 bp single-end; ~8-16 million reads per sample regime) The quality of sequencing reads was checked with fastQC 0.11.2 (www.bioinformatics.babraham.ac.uk/projects/fastqc).	Illumina HiSeq 2000
P-MTAB-70323	high throughput sequence alignment protocol	Reads were aligned to the Bacillus subtilis subps. subtilis strain 168 genome (NCBI Nucleotide acc. no. NC_000964) using BWA 0.7.9a-r786 (Li and Durbin, 2009) and samtools 0.1.19 (Li et al., 2009). Alignment quality was checked using QualiMap 2.1.3 (Okonechnikov et al., 2015) and IGV 2.3 (Thorvaldsdóttir et al., 2013). Bacillus subtilis genome annotation was obtained from NCBI Assembly (acc. no. GCF_000009045.1). Analysis of differential gene expression was performed using R (www.r-project.org) and the Bioconductor package DESeq2 (Huber et al., 2015; Love et al., 2014) at 5% false discovery rate.		BWA 0.7.9a-r786

Samples

Sample count18

Experimental Designsgrowth condition design, replicate design, stimulus or stress design, genetic modification design

Experimental Factorsgenotype, temperature

Experimental Factors

Factors Table show table

genotype	temperature	No. of Samples
rsgI mutant	37 degree celsius	3
rsgI mutant	52 degree celsius	3
sigI-rsgI knockout	37 degree celsius	3
sigI-rsgI knockout	52 degree celsius	3
wild type genotype	37 degree celsius	3
wild type genotype	52 degree celsius	3

GenotypesigI-rsgI knockout, wild type genotype, rsgI mutant

Temperature37 degree celsius, 52 degree celsius

Source Characteristics

Characteristics Table show table

growth condition	genetic modification	genotype	replicate	No. of Samples
37°C LB broth	gene disruption	rsgI mutant	3	1
37°C LB broth	gene disruption	rsgI mutant	1	1
37°C LB broth	gene disruption	rsgI mutant	2	1
37°C LB broth	deletion of the whole operon	sigI-rsgI knockout	3	1
37°C LB broth	deletion of the whole operon	sigI-rsgI knockout	1	1
37°C LB broth	deletion of the whole operon	sigI-rsgI knockout	2	1
37°C LB broth	none	wild type genotype	3	1
37°C LB broth	none	wild type genotype	2	1
37°C LB broth	none	wild type genotype	1	1
52°C LB broth	gene disruption	rsgI mutant	2	1
52°C LB broth	gene disruption	rsgI mutant	1	1
52°C LB broth	deletion of the whole operon	sigI-rsgI knockout	2	1
52°C LB broth	deletion of the whole operon	sigI-rsgI knockout	3	1
52°C LB broth	none	wild type genotype	2	1
52°C LB broth	none	wild type genotype	3	1
52°C LB broth	none	wild type genotype	1	1
52°C LB broth	gene disruption	rsgI mutant	3	1
52°C LB broth	deletion of the whole operon	sigI-rsgI knockout	1	1

OrganismBacillus subtilis subsp. subtilis str. 168

Growth condition52°C LB broth, 52°C LB broth, 37°C LB broth, 37°C LB broth

Genetic modificationdeletion of the whole operon, gene disruption, none

GenotypesigI-rsgI knockout, wild type genotype, rsgI mutant

Replicate3, 1, 2

Assays and Data

Assay count18

TechnologySequencing assay

Assay by MoleculeRNA assay

MAGE-TAB Files

MINSEQE Score

Exp. Design*

Raw*

Processed-

Protocols*

Variables*