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Sample GSM2049267 Query DataSets for GSM2049267
Status Public on Feb 06, 2016
Title MG1655 transcriptome from batch growth_2
Sample type SRA
 
Source name MG1655 batch
Organism Escherichia coli
Characteristics strain: K12 (MG1655)
growth protocol: Bacteria liquid culture
treatment: batch growth
Growth protocol A minimal growth medium as described in the study by Ihssen and Egli (2004) was used for all experiments. Bacterial stock cultures were streaked onto agar plates and incubated overnight. One colony was then transferred into 20 ml minimal medium, grown at 37 °C (250 r.p.m.) over night culture (ONC) and served as the inoculum for experiments. For continuous culture experiments we designed and constructed 500 ml bioreactors according to the study by Huwiler et al., (2012) that were half-filled with medium (0.5 g glucose per l) and incubated at 37 °C in a temperature controlled water bath. Before continuous cultivation (dilution rate=0.25), 1–2 ml of the ONC was transferred and grown in batch-mode until reactors became visibly turbid. Subsequently, cells were grown to steady-state (defined as constant optical density over time) and harvested for experimentation. For starvation experiments the medium flow was stopped during steady-state and bacteria were collected after 4 h. To avoid gene-expression signatures of stationary cells from the ONC, batch cultures (1000 ml Erlenmeyer flasks containing 100 ml of pre-warmed medium (1 g glucose per l); 37 °C; 250 r.p.m.)) were inoculated with 5 ml of an exponentially growing pre-culture that derived from the ONC.
Extracted molecule total RNA
Extraction protocol Harvested cells (4 × 6 ml) were immediately combined with 6 ml RNAlater (Life Technologies, Grand Island, NY, USA), centrifuged for 15 min at 12 000 r.p.m., washed with 1 ml RNAlater (3 min at 15 000 r.p.m.), re-suspended in 0.5 ml RNAlater and stored at −20 °C. For RNA extraction the RiboPure-Bacteria Kit (Life Technologies) was used according to the manufacturer’s instructions. To achieve high RNA yields several reactions for each sample were done in parallel and pooled at the end of the procedure. An additional DNase treatment step with TURBO DNase (Life Technologies) was included to assure no genomic DNA contamination. Messenger RNA was enriched using the RiboMinus Bacteria Kit (Life Technologies) according to the manufacturer’s instructions.
RNA quality was confirmed with the Bioanalyzer system (Agilent Technologies, Santa Clara, CA, USA) and sequencing (50 cycles, pooling eight bar-coded samples per lane) was performed on the Illumina HiSeq platform (Illumina, San Diego, CA, USA) at the Research Technology Support Facility (RTSF) at Michigan State University.
 
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina HiSeq 2000
 
Description Processed data: raw_count.txt, normalized_count.txt, panGeneMap.txt, Gene_annotations.txt
MG1655b2
Data processing Quality trimming of 3' end at Read Segment Quality Control Indicator.
Map RNA-Seq reads to reference genomes using Bowtie (Langmead et al., 2009) with alignment parameters (-n 2-e 70-l 28 –best)
Summarize the read count to each annotated gene
Remove non-protein coding counts
Differential expression analysis with DESeq (Anders and Huber 2010) with the default setting
Genome_build: NC_011741 (E.coli strain IAI1), NC_000913.2 (E.coli strain MG1655), TW09308 (E.coli strain TW09308), and TW11588 (E.coli strain TW11588)
Supplementary_files_format_and_content: All in tab-delimited ASCII format. raw_count.txt (pre-normalized read count), normalized_count.txt, panGeneMap.txt (geneID map between the strains and pangene ID), Gene_annotations.txt (annotation of genes)
 
Submission date Jan 28, 2016
Last update date May 15, 2019
Contact name Benli Chai
E-mail(s) chaibenl@msu.edu
Phone 517-353-3842
Organization name Michigan State University
Department Center for Microbial Ecology
Street address 567 Wilson Road Rm 2225 A
City East Lansing
State/province Michigan
ZIP/Postal code 48824
Country USA
 
Platform ID GPL14548
Series (1)
GSE77325 Gene expression analysis of E. coli strains provides insights into the role of gene regulation in diversification
Relations
BioSample SAMN04446524
SRA SRX1552235

Supplementary data files not provided
SRA Run SelectorHelp
Raw data are available in SRA
Processed data are available on Series record

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