GEO Accession viewer

NCBI > GEO > Accession Display

Not logged in | Login

GEO help: Mouse over screen elements for information.

Sample GSM4150224

Query DataSets for GSM4150224

Status

Public on Jul 09, 2020

Title

RMG Aero Stationary RNA-seq rep3

Sample type

SRA

Source name

cell culture

Organism

Zymomonas mobilis subsp. mobilis ZM4 = ATCC 31821

Characteristics

strain: ZM4
time point: stationary
rna-pretreatment: N/A
molecule subtype: rRNA-depleted RNA

Growth protocol

Zymomonas mobilis subsp. mobilis ZM4 ATCC31821 grown in rich medium with glucose (RMG) containing per L 10 g yeast extract, 2.6 g KH2PO4, 5 g K2HPO4, and 20 g glucose or minimal media with glucose (MMG) containing per L 20 g glucose, 2.6 g KH2PO4, 5 g K2HPO4, 0.5 g NaCl, 1 g (NH4)2SO4, 0.2 g MgSO4·7H2O, 25 mg Na2MoO4·2H2O, 10 mg CaCl2·2H2O, 1 mg calcium pantothenate, 25 mg FeSO4·7H2O, and 20 g glucose, and was adjusted to pH 6.4 with HCl. Cell growth was monitored in real-time by light scattering (termed OD) at 600 nM and extracellular glucose concentration was measurement by YSI 2700 Select. Overnight starter cultures of ZM4 were grown in RMG in an anaerobic chamber and used to inoculate 3L cultures of media in bioreactors (Applikon Biotechnology). Anaerobic cultures were sparged with N2/CO2 gas mix at a rate of 150mL/min and cells were agitated at 300 rpm. Aerobic cultures were sparged with atmospheric air at a rate of 700mL/min and agitated at 500 rpm. All multiomics samples were collected at 50% glucose depletion from the media (mid-glucose phase time point) and 1-hour post glucose depletion (stationary phase time point).

Extracted molecule

total RNA

Extraction protocol

Ten mL of culture was added to 1.25mL of ice-cold ethanol/phenol stop solution, and RNA was extracted using hot phenol method as described in PMID:21601083; total RNA was DNase I treated prior to all library construction.

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina HiSeq 2500

Data processing

RNA-seq: Reads were filtered for low quality and adapter readthrough using trimmomatic version 0.30 using the following parameters: ILLUMINACLIP:TruSeq3-PE.fa:2:22:10 SLIDINGWINDOW:4:28 MINLEN:75. Gene read counts were obtained using the revised protein-coding annotations from this study, using RSEM v1.2.3 and Bowtie v1.0.0 using the following parameters: --paired-end --calc-ci --estimate-rspd --forward-prob 0 --phred33-quals.
RSEM expected counts were used for downstream differential expression analysis. Pearson correlation between of gene counts between biological replicates was used to detect outlier libraries. All the retained libraries had inter-replicate Pearson correlation of at least 0.95. Features representing rRNA and tRNA, and genes with count sums less 5000 across all remaining samples were removed from the matrix. Gene count normalization and differential expression testing was performed using DESeq2 v1.14.1 run on R 3.3.0.
Genome_build: Genbank sequences CP023715.1, CP023716.1, CP023717.1, CP023718.1, and CP023719.1
Supplementary_files_format_and_content: 5p.complete.bedgraph files contain plus and minus strand, genome-wide read coverage for the corresponding sample where only the first base in the read is counted towards read coverage; coverage values are not normalized, negative values correspond reads mapped to the minus strand and positive coverage values correspond to reads mapped to plus strand. complete.bedgraph files contain plus and minus strand, genome-wide read coverage for the corresponding sample where every base in the read is counted towards read coverage; coverage values are not normalized, negative values correspond reads mapped to the minus strand and positive coverage values correspond to reads mapped to plus strand. compiledDESeq2.results.txt is a tab-delimited text file that contains the log2 fold change and adjusted p-value (aka q-value or FDR) for pair-wise differential expression analysis of RNA-seq gene counts. RNAseq.RSEM.expectedCounts.txt is a tab-delimited text file that contains the expected reads counts per gene as determined by rsem (used as described in corresponding data processing step description) compiled from all 19 RNA-seq data sets.

Submission date

Nov 05, 2019

Last update date

Jul 09, 2020

Contact name

Robert Landick

E-mail(s)

[email protected]

Organization name

University of Wisconsin

Street address

1550 Linden Dr