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Sample GSM1331412 Query DataSets for GSM1331412
Status Public on Oct 18, 2014
Title LB 0.4 B2 TEX neg L1 HS2
Sample type SRA
 
Source name Escherichia coli MG1655 cells
Organism Escherichia coli str. K-12 substr. MG1655
Characteristics strain: MG1655
medium: LB
growth phase: exponential
Treatment protocol Samples were mixed, incubated on ice for 10 min after which cells were collected by centrifugation at 4150 rpm at 4˚C for 10 min. Cell pellets were snap frozen in an ethanol/dry ice slurry and stored at -80°C until total RNA could be extracted.
Growth protocol To harvest total RNA samples, overnight cultures of wild type MG1655 grown in LB at 37˚C were diluted back 1:500 in either fresh LB or M63 minimal glucose medium and allowed to grow until the cultures reached an OD600 of ~ 0.4 and 2.0 for cultures grown in LB and an OD600 of ~0.4 for cultures grown in M63. For samples grown to OD600 of ~0.4 a total volume of 25 ml of cells were harvested and combined with 5 ml of stop solution (95% Ethanol, 5% acid phenol). For samples grown to OD600 of 2.0 a total volume of 5 ml of cells were harvested and combined with 1 ml of stop solution.
Extracted molecule total RNA
Extraction protocol Hot phenol (Sharma et al. 2010, Bloomberg 1990, EMBO J)
The libraries were generated by Vertis Biotechnologie AG (Munich, Germany). The samples were poly(A)-tailed by using poly(A) polymerase. The 5'-PPP were removed using tobacco acid pyrophosphatase (TAP) followed by the ligation of the RNA adapter to the 5'-monophosphate of the RNA. First-strand cDNA synthesis was performed with an oligo(dT)-adapter primer and the M-MLV reverse transcriptase. The resulting cDNA was PCR-amplified to reach a concentration of 20-30 ng/μl using a high fidelity DNA polymerase. The cDNA was purified using the Agencourt AMPure XP kit (Beckman Coulter Genomics) and was analyzed by capillary electrophoresis. The primers used for PCR amplification were designed for TruSeq sequencing according to the instructions of Illumina
 
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina HiSeq 2000
 
Data processing Demultiplexing
Fastq quality trimming using FastX and a cut-off value of 20
Fastq to fasta conversion using FastX
Size filtering: discarding reads shorter than 12 nt (TRAPL)
Read mapping using segemehl version 0.13 (TRAPL)
Coverage calculation and normalisation (TRAPL)
Genome_build: NC_000913.2
Supplementary_files_format_and_content: wiggle
 
Submission date Feb 20, 2014
Last update date May 15, 2019
Contact name Konrad U. Förstner
E-mail(s) foerstner@zbmed.de
Organization name ZB MED - Information Centre for Life Sciences
Department Information Services
Lab Förstner Lab
Street address Gleueler Str. 60
City Cologne
State/province North Rhine-Westphalia
ZIP/Postal code 50931
Country Germany
 
Platform ID GPL15010
Series (1)
GSE55199 Identification and validation of antisense RNAs in Escherichia coli
Relations
BioSample SAMN02646724
SRA SRX474168

Supplementary file Size Download File type/resource
GSM1331412_LB_0.4_B2_TEX_neg_L1_HS2_forward.wig.gz 8.8 Mb (ftp)(http) WIG
GSM1331412_LB_0.4_B2_TEX_neg_L1_HS2_reverse.wig.gz 9.1 Mb (ftp)(http) WIG
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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