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Sample GSM4456084 Query DataSets for GSM4456084
Status Public on Dec 31, 2022
Title R0199 rep3
Sample type SRA
 
Source name whole bacterial cells
Organism Caulobacter vibrioides
Characteristics strain: NA1000
genotype: pKF482 (pBVMCS-6-R0199)
growth phase: OD660=0.8
treatment: 0.5 mM vanillate
Treatment protocol Cells were harvested in tubes containing 1/5 vol. "stop-mix" (95% ethananol, 5% phenol) and snap frozen in liquid nitrogen until further processing.
Growth protocol C. crescentus NA1000 carrying either pBVMCS-6 (empty vector control) or pKF482 (pBVMCS-6 with R0199) were grown in M2 medium (+ 0.2% glucose; supplemented with chloramphenicol) to OD660=0.8, when expression from the vanillate-inducible promoter was activated by addition of 0.5 mM vanillate. Cells were harvested 15 minutes post induction.
Extracted molecule total RNA
Extraction protocol RNA was isolated using the HotPhenol method according to standard protocols. Prepared RNA was digested with DNaseI and cleaned up by phenol-chloroform extraction. Ribosomal RNA was depleted with the Ribo-Zero kit (Epicentre) for Gram-negative bacteria. Integrity of the prepared RNA was tested using a Bioanalyzer.
Directional cDNA libraries were prepared using the NEBNext Ultra II Directional RNA Libary Prep Kit for Illumina (NEB E7760) according to the manufacturer's instructions. Quality of the prepared libraries was tested using a Bioanalyzer.
 
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina HiSeq 1500
 
Description R199_3_ATTCCT
Data processing Sequencing of the cDNA libraries was performed by the LAFUGA service unit (Gene Center, munich, Blum group)
cDNA reads were imported into CLC Genomics Workbench v11.0 and analysed using mostly standard parameters.
3'adaptor- and quality trimming was performed using the trim reads command with the following settings: Sequence = TGACTGGAGTTCAGACGTGTGCTCTTCCGATCT, Strand = Minus, Action = Remove adapter, Scores = Mismatch: 2, Gapcost: 3, Cutoff: 10, Cutoff at end: 4
The trimmed reads were checked for quality and mapped to the C. crescentus NA1000 reference genome; NCBI Acc.number: NC_011916 using the following settings: Mismatch cost = 2, Insertion cost = 3, Deletion cost = 3, Length fraction = 0.8, Similarity fraction = 0.8, Global alignment = Yes, Strand specific = Reverse, Maximum number of hits for a read = 1
Reads mapping in CDS were counted, and genes with a total count cutoff of >10 in all samples were considered for analysis. Read counts were normalized (CPM), and transformed (log2). Differential expression was tested using the built in tool corresponding to edgeR in exact mode with tagwise dispersions. Genes with a fold-change of > 2.0 in either condition and a FDR adjusted p-value < = 0.05 were considered to be differentially expressed.
Genome_build: NC_011916
Supplementary_files_format_and_content: Read count table and edgeR (exact mode) analysis
 
Submission date Apr 07, 2020
Last update date Dec 31, 2022
Contact name Kathrin Fröhlich
E-mail(s) kathrin.froehlich@uni-jena.de
Organization name Friedrich-Schiller-Universität Jena
Department Fakultät für Biowissenschaften, Bacterial RNA Biology
Lab AG Fröhlich
Street address Winzerlaer Straße 2
City Jena
State/province Thuringia
ZIP/Postal code 07745
Country Germany
 
Platform ID GPL28356
Series (2)
GSE148208 Target identification for R0199
GSE148211 To be updated
Relations
BioSample SAMN14548610
SRA SRX8073089

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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