|
| Status |
Public on Jul 10, 2024 |
| Title |
ATCC14579, Etest |
| Sample type |
SRA |
| |
|
| Source name |
ATCC 14579
|
| Organism |
Bacillus cereus |
| Characteristics |
strain: ATCC 14579
isolate: Bacillus cereus sensu lato
group: reference strain
origin: Leibniz Insitute - DSMZ
treatment: MH agar with Etest Vancomycin
pooled biological_replicates: 4 agar plates
|
| Treatment protocol |
For vancomycin susceptibility testing, BC70 and ATCC 14579 were collected to obtain bacterial suspensions equal to McFarland 0.5. The suspensions were then evenly streaked on MH agar and Etest gradient strips with vancomycin were applied. For swarming condition, 5 µl of a bacterial suspension were spotted on MH agar with 0.7% agar. BC70 was also streaked on MH agar as additional control. All conditions were incubated at 37°C for exactly 13 h.
|
| Growth protocol |
The BC70 isolate and ATCC 14579 were initially cultured on columbia agar with 5% sheep blood at 37°C over night. Further experiments were carried out on MH agar at 37°C.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using the RNeasy Protect Bacteria Mini Kit. Approximately 1.8 OD600 of cells were used and all the RNA harvested was used to build the library.
Total RNA was further purified using the DNA-free Kit (ThermoFisher). Bacterial ribosomal RNA was depleted using the NEBNext rRNA Depletion Kit Bacteria (NEB) followed by polyadenylating with E. coli poly(A) polymerase. The direct cDNA Kit (Oxford Nanopore) was used for reverse transcription and library construction. The Native Barcoding Expansion 1-12 (Oxford Nanopore) was used for multiplexing the cDNA samples. All steps were carried out according to the manufacturers protocol. Quantification and quality control of RNA and cDNA were reduced to a minimum to obtain sufficiently high amounts of RNA or cDNA.
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| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
MinION |
| |
|
| Description |
Flow cell FLO-MIN106
BC70etest_ATCC14579etest_DGE.csv
|
| Data processing |
Sequencing data was basecalled using Guppy 6.4.6 in super-accurate mode.
Passed reads from technical replicates were analyzed for differential expression analysis using EPI2ME Desktop (v5.0.2) with the nextflow transcriptomics workflow (v0.2.0).
Reference-guided workflow was used with default parameters (gffcompare: -r -R minimap index opts: -k14 minimap2 opts: -uf minimum mapping quality: 40 stringtie opts: --conservative).
Differential expression analysis by edgeR in Epi2Me was performed with default parameters (min gene expr: 10 min feature expr: 3 min samps gene expr: 3 min samps feature expr: 1).
Assembly: The NCBI RefSeq assembly reference genome and the RefSeq annotation of Bacillus cereus (genome assembly ASM222028v1, RefSeq GCF_002220285.1, strain FORC47) were used for reference-based transcriptome assembly and reference annotation, respectively.
Supplementary files format and content: Comma-separated values (CSV) files include output data from differential expression analysis for the follwing samples/conditions: barcode01 (BC70, Etest) - barcode04 (ATCC14579, Etest), barcode01 (BC70, Etest) - barcode08(BC70, MH), barcode01 (BC70, Etest) - barcode11 (BC70, swarming). The CSV files include gene ID, log2 fold change between experimental conditions, the log-scaled counts per million measure of abundance, the p-value and the false discovery corrected p-value (FDR) for all transcripts.
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| |
|
| Submission date |
Jan 12, 2024 |
| Last update date |
Jul 10, 2024 |
| Contact name |
Paul Jakob Schmid |
| E-mail(s) |
paul.schmid@medunigraz.at
|
| Phone |
004331638573601
|
| Organization name |
Medical University of Graz
|
| Department |
Diagnostic and Research Institute of Hygiene, Microbiology and Environmental Medicine
|
| Lab |
Dr. Kittinger Lab
|
| Street address |
Neue Stiftingtalstraße 6
|
| City |
Graz |
| ZIP/Postal code |
8010 |
| Country |
Austria |
| |
|
| Platform ID |
GPL34081 |
| Series (1) |
| GSE253142 |
Pseudo-resistant Bacillus cereus uses biofilm-related mechanism to mimic vancomycin resistance during agar diffusion susceptibility testing |
|
| Relations |
| BioSample |
SAMN39423329 |
| SRA |
SRX23203829 |
