|
| Status |
Public on Jul 03, 2024 |
| Title |
Bacteria, Flow, rep3 |
| Sample type |
SRA |
| |
|
| Source name |
Planktonic bacteria
|
| Organism |
Pseudomonas aeruginosa PAO1 |
| Characteristics |
strain: PAO1
cell type: Planktonic bacteria
genotype: WT
treatment: Flow
|
| Treatment protocol |
Planktonic bacteria was flowed in a microfluidic channel. A branched channel section near the device outlet was used to inject 37% Formaldehyde at a flow rate equal to 1/8th of the main channel to fix planktonic bacteria in a 4% Formaldehyde (final concentration) media solution after they flowed for 55 min within the device and prior to exiting the channel. No-flow control cells were incubated in the channel for the same duration as cells that were subject to flow.
|
| Growth protocol |
P. aeruginosa PAO1 was grown in LB media at 25C on a floor shaker. Prior to experiments, overnight-grown cultures were diluted 1:200 in fresh media and grown to mid-exponential phase (OD of 0.4-0.5).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from fixed cells using the Qiagen RNeasy Mini kit with the following modifications to the protocol. Fixed cells were spun down at 6000 ×g for 5 min and resuspended in an 80 µL volume containing 30 mM Tris-HCL (pH 8.0), 2 mM EDTA, and 0.1% Triton-X 100, and 20 mg/mL lysozyme. Lysis was performed at 37C for 20 min, followed by Proteinase K digestion at 65C for 1 h. In addition, on-column DNase digestion was performed during RNA extraction as per the Qiagen RNeasy protocol.
Libraries were prepared by SeqCenter (Pennsylvania) using llumina Stranded Total RNA library preparation and RiboZero Plus rRNA depletion
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq X Plus |
| |
|
| Description |
See column Rc1 for counts per million
|
| Data processing |
RNA-Seq analysis was performed by SeqCenter (Pennsylvania). Quality control and adapter trimming was performed with bcl-convert. Read mapping was performed with HISAT2. Read quantification was performed using Subread’s featureCounts functionality. Raw, quantified read counts loaded into R were normalized using edgeR’s Trimmed Mean of M values (TMM) algorithm. Subsequent values were then converted to counts per million (CPM).
Assembly: Pseudomonas aeruginosa PAO1 (RefSeq: GCF_000006765.1)
Supplementary files format and content: tab-delimited text file contains counts per million (CPM) for all samples as a single matrix table
|
| |
|
| Submission date |
Jul 03, 2024 |
| Last update date |
Jul 03, 2024 |
| Contact name |
Ashwin Ramachandran |
| Organization name |
Princeton University
|
| Street address |
Department of Molecular Biology, Department of Mechanical and Aerospace Engineering
|
| City |
Princeton |
| State/province |
NJ |
| ZIP/Postal code |
08542 |
| Country |
USA |
| |
|
| Platform ID |
GPL34676 |
| Series (1) |
| GSE271426 |
RNA-Seq of Planktonic P. aeruginosa in flow |
|
| Relations |
| BioSample |
SAMN38710958 |
| SRA |
SRX25181864 |
