|
| Status |
Public on Mar 01, 2019 |
| Title |
6B4-Cap- replicate 3 |
| Sample type |
SRA |
| |
|
| Source name |
Microbial cell culture
|
| Organism |
Pseudomonas [fluorescens] SBW25 |
| Characteristics |
evolved strain: 6B4
morphotype: Switcher
|
| Growth protocol |
Method: 3x single colonies of each of SBW25, 6A4 and 6B4 were grown on KB agar (28˚C, 48 hours), precultured in shaken KB (28˚C, overnight), grown to mid-exponential phase in shaken KB flasks (28˚C, OD600 of 0.4-0.6). SBW25 and 6A4 cultures were used for total RNA extraction; 6B4 cultures were split into cap+ and cap- fractions by centrifugation before total RNA isolation (and are included as separate samples above).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA extraction and purification was performed using a RiboPureTM Bacteria Kit (Ambion®). RNA libraries were prepared for sequencing using standard Illumina protocols.
Normalized mRNA-seq library preparation was performed by the Australian Genome Research Facility (Brisbane, Australia). AGRF used a TruSeq kit to prep the RNA library, and perforemdd Ribo-Zero Bacteria rRNA Removal (https://sapac.illumina.com/products/scriptseq-bacteria.html). AGRF also performed the Illumina sequencing.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Data processing |
RNA-seq datasets were mapped to the SBW25 genome (downloaded from Genbank under the accession number NC_012660) via Bowtie2 with default settings
The coverage per gene of the genome mapping was determined with HTSeq. The gene annotation for HTSeq was also downloaded from the SBW25 Genbank entry.
Differentially expressed genes were identified by applying DESeq2. The standard workflow in https://bioconductor.org/packages/release/bioc/manuals/DESeq2/man/DESeq2.pdf was used, except that the alpha parameter was set to 0.3 to reduce the number of genes that were falsely classified as not significantly differentially expressed between the different morphotypes
Genome_build: ASM922v1
Supplementary_files_format_and_content: The processed data files contain information obtained by running "htseq-count" on the aligned sequence files. The files contain the sequence read coverage for each gene in the Pseudomonas fluorescens SBW25 genome. This data can be used to determine differentially expressed genes by for example DESeq2
|
| |
|
| Submission date |
Jul 02, 2018 |
| Last update date |
Mar 01, 2019 |
| Contact name |
Jenna Gallie |
| E-mail(s) |
[email protected]
|
| Organization name |
Max Planck Institute for Evolutionary Biology
|
| Street address |
August-Thienemann Strasse 2
|
| City |
Ploen |
| ZIP/Postal code |
24306 |
| Country |
Germany |
| |
|
| Platform ID |
GPL25322 |
| Series (1) |
| GSE116490 |
RNA-seq data showing the effects of rpoD mutation t1682c in Pseudomonas fluorescens |
|
| Relations |
| BioSample |
SAMN09531952 |
| SRA |
SRX4330077 |
