GEO Accession viewer

NCBI > GEO > Accession Display

Not logged in | Login

GEO help: Mouse over screen elements for information.

Sample GSM8647163

Query DataSets for GSM8647163

Status

Public on Nov 27, 2024

Title

WT_45 min

Sample type

SRA

Source name

bacterial cultures

Organism

Escherichia coli K-12

Characteristics

tissue: bacterial cultures
strain: K-12
genotype: WT
treatment: 3.5 mg/ml phenol for 45 min

Treatment protocol

These log-phase cultures were treated with 3.5 mg/ml phenol. After treatment for 45 min, cells were collected by centrifugation, frozen using liquid nitrogen, and then stored at -80 °C for subsequent total RNA extraction. The cultures immediately before phenol treatment were used as a control (time 0).

Growth protocol

Overnight cultures of the wildtype and pheS F158C strains were diluted 100-fold into fresh LB medium, and were grown to mid-log phase (OD600 = 0.3) at 37 °C with shaking.

Extracted molecule

total RNA

Extraction protocol

Total RNAs were prepared using an RNA extraction kit following the manufacturer protocol (Tiangen, China).
A total amount of 3 µg RNA per sample was used as input material for the RNA sample preparations. Sequencing libraries were generated using NEBNext Ultra Directional RNA Library Prep Kit for Illumina (NEB, USA) following manufacturer’s recommendations. Briefly, rRNAs were removed by the Ribo-zero kit to enrich mRNAs, which were then digested into short fragments. The fragmented mRNAs were used as template to generate one-strand DNA using a six-base random primer and the corresponding double-stranded cDNA. After purification, the cDNA was firstly end-repaired, A-tailed and ligated to a sequencing linker. The size of fragments was selected using AMPure XP beads. PCR amplification was performed, and the amplified products were then purified using AMPure XP beads to obtain the final library. The cDNA library was sequenced (RNA-seq) on an Illumina novaseq platform, and paired-end reads were generated.

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina NovaSeq 6000

Description

Library name: WT_phenol

Data processing

Clean reads were obtained by removing reads containing adapter, reads containing ploy-N and low quality reads from raw data.
Both building index of reference genome and aligning clean reads to reference genome were used Bowtie2-2.2.3 (Langmead, B. and S. L. Salzberg, 2012). HTSeq v0.6.1 was used to count the reads numbers mapped to each gene.Then FPKM of each gene was calculated based on the length of the gene and reads count mapped to this gene.
Differential expression analysis of two conditions was performed using the DEGSeq R package (1.20.0).
Gene Ontology (GO) enrichment analysis of differentially expressed genes was implemented by the GOseq R package, in which gene length bias was corrected.
Assembly: Genome_build: NZ_CP009273.1
Supplementary files format and content: tab-delimited text file includes raw counts for each Sample
Supplementary files format and content: csv files include FPKM values for each Sample

Submission date

Nov 22, 2024

Last update date

Nov 27, 2024

Contact name

shouqiang hong

E-mail(s)

[email protected]

Phone

+8613358397835

Organization name

Xiamen university