|
| Status |
Public on Sep 25, 2020 |
| Title |
RNASeq of planctonic culture of Pseudomonas aeruginosa UCBPP-PA14 delta fliC 2 |
| Sample type |
SRA |
| |
|
| Source name |
planctonic culture of Pseudomonas aeruginosa UCBPP-PA14 delta fliC
|
| Organism |
Pseudomonas aeruginosa UCBPP-PA14 |
| Characteristics |
p. aeruginosa genotype: PA14 delta fliC
growth condition: Bacterial mid-log phase culture (4 h) in LB
|
| Treatment protocol |
DualSeq experiment: The bacterial concentration was adjusted to MOI 1 (5*10^5 per 100 µl) and 100 µl were used for the infection. The bacteria were centrifuged on top of the cells using 1200 rpm for 5 min. The infection was performed for 3 h at 37°C, 5% CO2 and 75% humidity.
Spermidine experiment: Cells were treated for 1 h with 10 µM spermidine at 37°C, 5% CO2 and 75% humidity. Triplicates were pooled to obtain one biological replicate.
|
| Growth protocol |
DualSeq experiment: Bacteria: Bacterial subcultures were grown to mid-log phase (3.5 h) in liquid LB and the infection dose was adjusted to 5*10^5 bacteria (MOI 1). RAW264.7: Cells were cultured in DMEM Medium at 37°C , 5% CO2 and 75% Humidity. One day before infection, cells were isolated, counted and split into 24-well plates at a concentration of 2.5*10^5 cells per well.
Spermidine experiment: Cells were maintained as during the Dual-Seq Experiment.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
DualSeq experiment: Medium was discarded. Cells were scratched off and resuspend in RNAprotect. RNA extraction was done as previously described (Kordes et al., 2018).
Spermidine experiment: Medium was discarded. Cells were scratched off and resuspend in RNA buffer. RNA extraction was done as previously described (Kordes et al., 2018).
DualSeq experiment: NEBNext Ultra II Directional RNA Library Prep.
Spermidine experiment: NEBNext® Single Cell/Low Input RNA Kit.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Description |
PA14_dfliC_2
C_2
Processed data file: rpg_table_PA14_planctonic.csv
|
| Data processing |
Mus musculus:
Reads were mapped using tophat2 version 2.1.0 (configurations: --no-novel-juncs --library-type fr-secondstrand. Attention, data from spermidine experiment are unstranded: fr-unstranded)
Files were converted into bam format using samtools version 1.9.
Reads per gene for RAW267.7 cells were extracted using the R package Rsubreads version 1.34.7.
Pseudomonas aeruginosa UCBPP-PA14:
Reads were mapped using bowtie2 version 2.2.6 (--no-1mm-upfront).
Files were converted into bam format using samtools version 1.9.
Reads per gene from Pseudomonas aeruginosa UCBPP-PA14 were extracted bedtools v2.29.0.
Supplementary_files_format_and_content: Semicolon separated csv file including reads per gene. Row names: gene Ids; column names: Treatment/bacterial strain (S/C: Spermidine/control; Wt: PA14 wild type, C: delta fliC, M: delta motABCD, K: delta flgK, U: uninfected).
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| |
|
| Submission date |
Dec 10, 2019 |
| Last update date |
Sep 25, 2020 |
| Contact name |
Matthias Preusse |
| Organization name |
Helmholtz Centre for Infection Research
|
| Street address |
Inhoffenstr. 7
|
| City |
Braunschweig |
| ZIP/Postal code |
38124 |
| Country |
Germany |
| |
|
| Platform ID |
GPL27892 |
| Series (1) |
| GSE141757 |
Host-induced spermidine production in motile Pseudomonas aeruginosa triggers phagocytic uptake |
|
| Relations |
| BioSample |
SAMN13523780 |
| SRA |
SRX7299392 |
