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Sample GSM7720021

Query DataSets for GSM7720021

Status

Public on Oct 01, 2023

Title

B. cereus, LF10, Rep a

Sample type

SRA

Source name

INRA C3

Organism

Bacillus cereus

Characteristics

strain: INRA C3
treatment: 10 mg/ml lactoferrin

Growth protocol

100 ml mMOD medium were inoculated with strain INRA C3 to OD600 = 0.1 and incubated at 37 °C for five hours under continuous agitation. The culture was centrifuged (10 min, 4,000 x g, 4 °C) and washed twice in cold mMOD-Fe. 50 ml mMOD, mMOD-Fe and mMOD with 10 mg/ml lactoferrin-based food supplement no. 1 were inoculated to OD600 = 1 and further incubated for two hours before centrifugation (2 min, 10,000 x g, 4 °C).

Extracted molecule

total RNA

Extraction protocol

Total RNA preparation and on-column DNase digestion were performed using RNeasy® Mini Kit (Qiagen) according to the instructions of the manufacturer. RNA quality was tested via spectrophotometer and agarose gel electrophoresis. RNA purity was confirmed via PCR for 16 S rRNA.
Random primed cDNA libraries were constructed according to the Illumina Genome Analyzer II protocol.

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina HiSeq 2500

Data processing

Samples were sequenced via Illumina HiSeq. 2500 Genome Sequencer with 2 x 150 bp paired-end mode. Data analysis included mapping of the samples against a reference genome (B. cereus F837/76; CP003187) and determination of gene expression.
Mapping of reads to reference sequences was performed using BWA-MEM (version 0.7.12-r1039, http: //bio-bwa.sourceforge.net/). The reads and their associated alignment positions are provided as BAM formatted files.
Raw read counts were created using featureCounts (version 1.5.1, http://subread.sourceforge. net/, Liao et al. (2014)). Only reads overlapping "CDS" features were counted. All reads mapping to features with the same meta-feature attribute were summed. Hereby, the "META_FEATURE_ID" attribute was used as a meta-feature attribute. Please see the provided .gff file for used annotations. Only reads with unique mapping positions and a mapping quality score of at least 10 were considered for read counting. Supplememtary alignments were ignored for read counting. Paired-end reads that mapped to different chromosomes or with unexpected strandedness were ignored.
Instead of counting paired-end reads twice, they were counted only once, i.e. as single fragment. Reads mapping to multiple features were assigned to the feature that has the largest number of overlapping bases. A Trimmed Mean of M-values (TMM) normalization was performed using the edgeR package (version 3.16.5, Robinson and Oshlack, 2010. Robinson and McCarthy, 2010. http://bioconductor.org/ packages/release/bioc/html/edgeR.html).
The basic assumption of this normalization technique is that most features (e.g. genes) should not be differentially expressed between samples. Roughly, for each sample a normalization factor is calculated as the weighted mean of feature-wise log ratios between this sample and a reference sample. For this calculation, the most expressed features and the features with the largest log ratios are excluded. The counts-per-million values of the normalized read counts are provided in the *.profiling-table.txt file. Length-normalization was not performed.
Assembly: CP003187.fasta
Supplementary files format and content: tsv format

Submission date

Aug 21, 2023

Last update date

Oct 01, 2023

Contact name

Nadja Jessberger

E-mail(s)

Nadja.Jessberger@tiho-hannover.de

Organization name

University of Veterinary Medicine Hannover, Foundation

Department

Institute for Food Quality and Food Safety