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Sample GSM4122832

Query DataSets for GSM4122832

Status

Public on Oct 09, 2020

Title

lomA mutant_RNA m2011-2

Sample type

SRA

Source name

lomA mutant

Organism

Leptospira interrogans serovar Manilae

Characteristics

strain: UP-MMC-NIID LP

Treatment protocol

no treatment

Growth protocol

Exponential-phase cultures at 30°C.

Extracted molecule

total RNA

Extraction protocol

Cells were collected by centrifugation and pelleted cells were resuspended in TRIzol. RNA was then purified according to the manufacturer's instructions (Invitrogen, Saint Aubin, Ile de France, France). DNA was degraded by using a Turbo DNA-free kit according to the manufacturer's instructions (Invitrogen).
From 0.5 μg of total RNA, we depleted ribosomal RNAs using the Ribo-Zero rRNA Removal Kit (Bacteria) from Illumina. On these rRNA-depleted RNAs, the sequencing libraries were constructed using the TruSeq Stranded mRNA Sample preparation kit following the manufacturer’s instructions (Illumina). The directional libraries were controlled on Bioanalyzer DNA1000 Chips (Agilent Technologies) and concentrations measured with the Qubit® dsDNA HS Assay Kit (ThermoFisher). Sequences of 65 bases were generated on the Illumina Hiseq 2500 sequencer.

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina HiSeq 2500

Description

lomA mutant_RNA

Data processing

Reads were cleaned of adapter sequences and low-quality sequences using cutadapt version 1.11
Bowtie version 1.2 with default parameters, was used for alignment on the reference genome (L. interrogans serovar Manilae strain UP-MMC-NIID LP, from MicroScope Platform).
Genes were counted using featureCounts version 1.4.6-p35from Subreads package (parameters: -t CDS -g locus_tag -s 1)
Count data were analyzed using R version 3.5.15and the Bioconductor packageDE Seq2 version 1.20.0f. The normalization and dispersion estimation were performed with DESeq2 using the default parameters and statistical tests for differential expression were performed applying the independent filtering algorithm. A generalized linear model was set in order to test for the differential expression between WT, complemented strain, and lomA. For each pairwise comparison, raw p-values were adjusted for multiple testing according to the Benjamini and Hochberg (BH) procedure and genes with an adjusted p-value lower than 0.05 were considered differentially expressed.
Genome_build: interrogans serovar Manilae strain UP-MMC-NIID LP, from MicroScope Platform
Supplementary_files_format_and_content: tabulated files from DESeq2

Submission date

Oct 15, 2019

Last update date

Oct 09, 2020

Contact name

Rachel Legendre

E-mail(s)

rachel.legendre@pasteur.fr

Organization name

Institut Pasteur

Department

Research and Resource Center for Scientific Informatics