|
| Status |
Public on Apr 16, 2022 |
| Title |
Expo.phase_S.aureus_sequence |
| Sample type |
SRA |
| |
|
| Source name |
Expo.phase_S.aureus
|
| Organism |
Staphylococcus aureus |
| Characteristics |
genotype: WT HG001 (AEs1)
stress: Untreated _ exponential growth _ 37C
media: BHI
|
| Treatment protocol |
The treatments used for each sample are mentioned in the characteristics field.
|
| Growth protocol |
The growth protocols used for each sample are mentioned in the characteristics field.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA from bacterial strains was extracted using the hot phenol procedure or the RiboPure™ Bacteria Kit (Invitrogen, ON, Canada, Cat. No. AM1925) following the manufacturer's recommendations. OMV-derived RNAs were isolated with RiboPure™ Bacteria Kit or RNAzol RT (Sigma, MO, USA, Cat. No. R4533) reagents following the manufacturer’s recommendations. RNAzol RT kit allowed selective isolation (enrichment) of small RNAs (<200 nt). All RNA samples were subjected to treatment with DNase-I when applicable, quantified with the NanoDrop™ 2000 Spectrophotometer (Thermo Scientific™, Cat. No. ND-2000) and saved at -80°C before downstream applications.
For each biological condition, an RNA sample was prepared by pooling equivalent amounts of total RNA isolated from 2 or 3 biological replicate samples. Total RNA was shipped on dry ice to the sequencing platform of Arraystar Inc. (Rockville, MD, USA). Total RNA of each sample was used to prepare the sRNA sequencing library which included the following steps: 1) 3’ adapter ligation with T4 RNA ligase 2 (truncated); 2) 5’ adapter ligation with T4 RNA ligase; 3) cDNA synthesis with RT primer; 4) PCR amplification; 5) extraction and purification of ~130-150 bp PCR amplified fragments from the PAGE gel. After the completed libraries were quantified with Agilent 2100 Bioanalyzer, the DNA fragments in the libraries were denatured with 0.1 M NaOH to generate single-stranded DNA molecules, captured on Illumina flow cells, amplified in situ and finally sequenced for 51 cycles on Illumina NextSeq according to the manufacturer’s instruction.
|
| |
|
| Library strategy |
ncRNA-Seq |
| Library source |
transcriptomic |
| Library selection |
size fractionation |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Description |
RNA 8-30nt library
Bacterial sRNA-Seq analysis
|
| Data processing |
Raw sequences were generated as clean reads from Illumina NextSeq by real-time base calling and quality filtering.
Subsequently, the 3’ adapter sequence was trimmed from the clean reads and the reads with lengths shorter than 8 nt were discarded. As the 5’ adapter was also used as the sequencing primer site, the 5’ adapter sequence is not present in the sequencing reads.
The trimmed reads (length ≥8 nt) were aligned to the corresponding genome database using novoalign software (http://www.novocraft.com). sRNA read counts were normalized as tag counts per million.
Sequences known to be contaminant confounders from RNA isolation procedures were discarded before analysis. Finally, a second dataset was generated for the E. coli and OMV samples by excluding the reads that are also present in the LB culture medium sample.
Assembly: See Supplementary_file_Genome_Database_GEO_PROVOST_LAB.pdf.
Supplementary files format and content: *.trimmed_tags.fa: Trimmed Reads Data contains trimmed reads (FASTA format). Defline: Unique identifier/name_Rank (from most to least abundant)_Quantitative information = copy number = Tags per million reads (TPM).
|
| |
|
| Submission date |
Apr 13, 2022 |
| Last update date |
Apr 16, 2022 |
| Contact name |
Patrick PROVOST |
| E-mail(s) |
patrick.provost@crchudequebec.ulaval.ca
|
| Organization name |
CRCHU of Québec City - Université LAVAL
|
| Department |
Centre Hospitalier de l'Université Laval (CHUL) - Microbiology, Infectious diseases, Immunology
|
| Lab |
PROVOST LAB
|
| Street address |
2705 , Boulevard Laurier, Bloc T
|
| City |
Quebec |
| State/province |
QUEBEC |
| ZIP/Postal code |
G1V4G2 |
| Country |
Canada |
| |
|
| Platform ID |
GPL24034 |
| Series (1) |
| GSE200758 |
RNA sequencing unveils very small RNAs with potential regulatory functions in bacteria |
|
| Relations |
| BioSample |
SAMN27567853 |
| SRA |
SRX14845705 |
