|
| Status |
Public on Dec 07, 2018 |
| Title |
M. tuberculosis H37Rv, replicate 2 |
| Sample type |
SRA |
| |
|
| Source name |
cell culture
|
| Organism |
Mycobacterium tuberculosis |
| Characteristics |
strain: H37Rv
|
| Treatment protocol |
No treatment was perfomed.
|
| Growth protocol |
Mycobacterium tuberculosis strains were grown at 37°C in 7H9 medium (Difco) supplemented with 0.2% glycerol, 0.05% Tween 80 and 10% albumin-dextrose-catalase (ADC, Middlebrook) or on 7H10 plates supplemented with 0.5% glycerol and 10% oleic acid-albumin-dextrose-catalase (OADC, Middlebrook).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total mRNA was extracted with 300 μl of chloroform : isoamyl alcohol (24:1) and precipitated at −20°C overnight with isopropyl alcohol prior to washing with 70% EtOH, then dried and resuspended in 40 μl DEPC‐treated water. RQ1 RNase‐Free DNase (Promega) and TURBO DNase (Ambion) were used to remove contaminating DNA, as recommended by the manufacturers. DNase treatment was followed by RNA purification using one volume of phenol : chloroform : isoamyl‐alcohol (25:24:1) and precipitation using 0.1 volume of 3 M sodium acetate (pH = 5.2) and 0.7 volume of isopropanol. RNA was resuspended in DEPC‐treated water and stored at −80°C. Integrity of RNA was checked by agarose gel electrophoresis, the amount and purity of RNA were assessed using a Nanodrop instrument.
RNA-seq libraries were prepared from 1 g of total RNA. The RNA samples were depleted of r-RNAs with the Illumina Ribo-Zero rRNA Removal Kit (Gram-Positive Bacteria) then used to generate sequencing libraries with the Illumina TruSeq Stranded mRNA reagents, omitting the polyA selection step (Illumina, San Diego, California, USA). Cluster generation was performed with the resulting libraries using the Illumina TruSeq SR Cluster Kit v4 reagents and sequenced on the Illumina HiSeq 2500 using TruSeq SBS Kit v4 reagents.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Data processing |
Reads were adapter- and quality-trimmed with Trimmomatic v0.33 and mapped onto the M. tuberculosis H37Rv reference genome (RefSeq NC_000962.3) using Bowtie2 v2.2.5.
Counting reads over features was done with featureCounts from the Subread package v1.4.6.
DESeq2 was used to infer differentially expressed genes.
Genome_build: NC_000962.3
|
| |
|
| Submission date |
Aug 23, 2018 |
| Last update date |
Dec 07, 2018 |
| Contact name |
Andrej Benjak |
| E-mail(s) |
[email protected]
|
| Organization name |
University of Bern
|
| Department |
Department for BioMedical Research
|
| Street address |
Murtenstrasse 24-28
|
| City |
Bern |
| ZIP/Postal code |
3008 |
| Country |
Switzerland |
| |
|
| Platform ID |
GPL17879 |
| Series (1) |
| GSE118994 |
Transcriptional profile of Mycobacterium tuberculosis espL deletion mutant |
|
| Relations |
| BioSample |
SAMN09907129 |
| SRA |
SRX4605772 |
