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Sample GSM7035080

Query DataSets for GSM7035080

Status

Public on Feb 08, 2023

Title

menF KO adapted 1, rep 2

Sample type

SRA

Source name

GMOS menF KO, adapted

Organism

Escherichia coli str. K-12 substr. MG1655

Characteristics

strain: GMOS menF KO, adapted
culture type: batch

Extracted molecule

total RNA

Extraction protocol

3 ml of cell broth (at an A600 ≈ 0.6) was immediately added to two volumes of Qiagen RNA-protect Bacteria Reagent (6 ml), vortexed for 5 s, incubated at room temperature for 5 min and immediately centrifuged for 10 min at 17,500 r.p.m. The supernatant was decanted, and the cell pellet was stored at −80 °C. Cell pellets were thawed and incubated with Ready-Lyse Lysozyme, SuperaseIn, protease K and 20% SDS for 20 min at 37 °C. Total RNA was isolated and purified using the Qiagen RNeasy Mini Kit columns and following vendor procedures. An on-column DNase treatment was performed for 30 min at room temperature. RNA was quantified using a Nano drop and quality assessed by running an RNA nano chip on a bioanalyser. The ribosomal RNA was removed using an Illumina Ribo-Zero rRNA removal kit for Gram-negative bacteria.
A KAPA Stranded RNA-Seq Kit (Kapa Biosystems KK8401) was used following the manufacturer’s protocol to create sequencing libraries with an average insert length of around 300 bp

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina HiSeq 4000

Description

quinone__menF_ale5__2
quinone_log_tpm.csv
quinone_counts.csv

Data processing

Raw read trimming was performed with Trim Galore using the default options, followed by FastQC on the trimmed reads.
Reads were aligned to the E. coli K-12 MG1655 reference genome (NC_000913.3) using bowtie with the following optional arguments: -X 1000, -3 3, -n 2
Read direction was inferred with RSEQC.
Read counts were generated with featureCounts with the following optional arguments: -p -B -C -P --fracOverlap 0.5
Quality control metrics were assembled with MultiQC and the final expression was reported in units of log2-transformed transcripts per million for each gene.
Assembly: NC_000913.3
Supplementary files format and content: quinone_log_tpm.csv contains log2[TPM] values for all samples and genes.
Supplementary files format and content: quinone_counts.csv contains raw read counts as computed by featureCounts for all samples and genes.

Submission date

Feb 08, 2023

Last update date

Feb 08, 2023

Contact name

Cameron Lamoureux

E-mail(s)

clamoure@eng.ucsd.edu

Organization name

University of California San Diego

Department

Bioengineering