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Sample GSM8146996

Query DataSets for GSM8146996

Status

Public on May 31, 2024

Title

WT, Daptomycin, repl 3

Sample type

SRA

Source name

bacterial cell

Organism

Enterococcus faecalis

Characteristics

mode of growth: planktonic bacteria
strain: OG1RF
cell type: bacterial cell
genotype: WT
growth phase: Exponential phase
treatment: Daptomycin

Treatment protocol

For antibiotic treated cells, WT cells were grown in BHI to mid-exponential growth phase (OD 0.4) and then treated with 1µg/ml daptomycin for 15 min.

Growth protocol

Strains WT, ∆srtA, ∆htrA, ∆srtA∆htrA, croR::tn∆srtA∆htrA, ∆srtA∆htrA∆ebpABC cells were grown in TSB supplemented with 0.25% glucose and incubated static at 37°C until mid-exponential growth phase (OD600 0.5).

Extracted molecule

total RNA

Extraction protocol

Total RNA was extracted using the UltraClean® Microbial RNA Isolation Kit (MO BIO Laboratories Inc., Singapore). Extracted RNA samples were subjected to rigorous DNase treatment using TURBO DNA-free™ kit (Ambion®, Singapore) and purified DNA-free RNA samples were subjected to ribosomal depletion with Ribo-Zero™ Magnetic Kits (Epicentre®, Singapore), all according to manufacturer’s protocols. Quantification of RNA and DNA were performed using Qubit™ RNA Assay Kits and Qubit™ dsDNA HS Assay Kits (Invitrogen, Singapore), respectively. The integrity of RNA was analyzed by gel electrophoresis using Agilent RNA ScreenTape (Agilent Technologies, Singapore).
mRNA libraries for RNA sequencing were prepared using TruSeq Stranded mRNA Library Prep Kit (Illumina, USA), the quality of the library analyzed via Bioanalyzer (Agilent, USA).

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina HiSeq 2000

Description

WT-DAP1-E2
WT DAP vs untreated.xlsx

Data processing

Sequencing reads (Accession number CP002621.1) mapping to predicted open reading frames (ORFs) were quantified using HTSeq
Counts for ribosomal and transfer RNA sequences were filtered out of the data set and differential expression analyses were performed in R (version 2.15.1) using the Bioconductor package, edgeR
Significantly differentially expressed genes were determined using a P-value and false discovery rate (FDR) cut-off of 0.05.
We annotated differentially expressed genes using a combination of KEGG annotations, as well as manual annotation using operon and other functional data from the literature and E. faecalis OG1RF database available on BioCyc
Assembly: RNA sequencing reads were mapped to the E. faecalis OG1RF reference genome (NCBI accession: NC_017316.1) using BWA (v0.5.9) with default parameters
Supplementary files format and content: tab-delimited text file includes raw counts, log2-fold change, and p-value and FDR for each strain/condition, as well as gene name and annotation if applicable.

Submission date

Mar 14, 2024

Last update date

May 31, 2024

Contact name

Kimberly Kline

Organization name

University of Geneva

Street address

1 Rue Michel Servet

City

Genève