|
| Status |
Public on Jul 04, 2019 |
| Title |
2LP |
| Sample type |
SRA |
| |
|
| Source name |
Planktonic cell extract
|
| Organism |
Campylobacter jejuni |
| Characteristics |
strain: 11168-O
state: Planktonic
|
| Growth protocol |
Planktonic cultures (LP) of C. jejuni strains were grown from an overnight plate culture, which was used to inoculate 250 mL of heart infusion broth. These cultures were incubated under microaerobic conditions at 42°C and 120 x rpm for 12 hrs. Biofilm samples (BF) were grown from overnight culture of C. jejuni which was diluted to an OD600 of 0.75 in mueller hinton broth. 10 mL of the cell suspension was then inoculated into glass petri dishes and incubated aerobically at 42°C for 48 hours. Samples were then harvested in 4 M guanidine thiocyanate in a 1% sodium lauryl sarcosine solution.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Samples were carefully laid over a 3.5 mL cushion of 5.7 M cesium chloride in 0.1 M EDTA in an OptiSeal polypropylene centrifuge tube with the remaining volume of the tube being filled with RNAse water. Samples were centrifuged at 27,000 x rpm for 16 hrs at 4°C in an Optima L-100 XP ultracentrifuge using a SW32Ti rotor. Following centrifugation, the upper phase was removed and the tubes were cut before being allowed to completely drain. The RNA pellet was washed in 100 µL 70% ethanol before being resuspended in 120 µL tris EDTA (TE) / sodium dodecyl sulfate (SDS) with 80 µL of TE being used to dissolve remaining RNA. 650 µL of chilled ethanol was added to resuspended samples in addition to 10 µL sodium acetate to precipitate RNA. Samples were centrifuged at 14,000 x rpm in a Microfuge 22R centrifuge for 15 mins at 4°C. The supernatant was removed and a further 650 µL of 75% ethanol was used to wash the pellet. Samples were centrifuged for a further 5 mins at 14,000 x rpm and the supernatant removed. Pellets were allowed to air dry before resuspension in RNase free water. Quality and concentration of RNA was assessed using a Nanodrop 2000.
RNA library construction was carried out using strand- specific Illumina TruSeq technology and DNA library construction was carried out using Illumina Nextera XT technology.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Description |
C. jejuni_processed_data_files.xls
|
| Data processing |
Raw reads were trimmed of Illumina TruSeq sequencing adapters, filtered for an average Phred Q score minimum of >30 and minimum read length of 32nt after trimming. Reads were further processed by trimming 3’ ends of bases with <30 Q scores.
Processed output files were aligned to the C. jejuni NCTC 11168 genome using Bowtie2 (v2.2.5).
Output .sam files were converted to .bam format using Samtools (v0.1.19), and name-sorted prior to input into HTSeq (v0.6.1). HTSeq counting was performed in union mode.
Differential gene expression analysis was performed using edgeR and R package.
Genome_build: GCA_000009085.1
Supplementary_files_format_and_content: The .xls file contains raw counts from HTSeq showing the abundance of each genomic feature across the samples.
|
| |
|
| Submission date |
Jul 03, 2019 |
| Last update date |
Jul 07, 2019 |
| Contact name |
Greg Tram |
| E-mail(s) |
g.tram@griffith.edu.au
|
| Organization name |
Griffith University
|
| Department |
Institute for Glycomics
|
| Street address |
Parklands Drive
|
| City |
Southport |
| State/province |
Queensland |
| ZIP/Postal code |
4215 |
| Country |
Australia |
| |
|
| Platform ID |
GPL26884 |
| Series (1) |
| GSE133783 |
Assigning a role for chemosensory signal transduction in Campylobacter jejuni biofilms using a combined omics approach |
|
| Relations |
| BioSample |
SAMN12211386 |
| SRA |
SRX6402902 |
