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| Status |
Public on May 25, 2024 |
| Title |
Wild-type_24_3 |
| Sample type |
SRA |
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| Source name |
mycelium
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| Organism |
Aspergillus fumigatus |
| Characteristics |
tissue: mycelium
genotype: CEA17 delta_akuB::KU80
growth condition: Aspergillus minimal media
genotype: wild type
time: 24 h
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| Growth protocol |
A. fumigatus was grown on Aspergillus minimal medium (AMM) agar plates (70 mM NaNO3, 11.2 mM KH2PO4, 7 mM KCl, 2 mM MgSO4, 1% (w/v) glucose and 1 μl/ml trace element solution (pH 6.5)). The trace element solution was composed of 1 g FeSO4 • 7 H2O, 8.8 g ZnSO4 • 7 H2O, 0.4 g CuSO4 • 5 H2O, 0.15 g MnSO4 • H2O, 0.1 g NaB4O7 • 10 H2O, 0.05 g (NH4)6Mo7O24 • 4 H2O, and ultra-filtrated water to 1000 ml. Spores were harvested after 5 days with 10 ml of sterile distilled water using a T-shaped inoculation spreader and spore suspensions were filtered through a 30-µm cell strainer (MACS, Miltenyi Biotec GmbH, Germany) to exclude mycelium.
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| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated from three biological replicates of CEA17ΔakuBKU80 from conidia, 24-h-, and 48-h-old mycelium. For A. fumigatus conidia, the spores were first homogenized in a bead beater (0.5 mm beads; FastPrep-24, MP Biomedicals) in Trizol. Fungal mycelium was collected from liquid culture using Miracloth (Millipore) and disrupted in liquid nitrogen using a precooled mortar and pestle. Roughly 0.5 g of homogenized mycelium was transferred into a 2-ml Eppendorf tube. 800 µl TRIzol was added to the ground mycelia and vortexed vigorously. Tubes were frozen briefly for 5 sec in liquid nitrogen and allowed to thaw on ice. To all samples, chloroform was added to the fungal samples in TRIzol, vortexed, and centrifugated for 5 min at 4°C at full speed. The aqueous upper phase was transferred to a fresh 2-ml tube without disturbing the interphase. RNA extraction from aqueous phase was done with 1 volume of phenol/chloroform/isoamyl alcohol (25:24:1, v/v). Brief vortexing preceded centrifugation for 5 min at 4°C. RNA was precipitated using 400 µl isopropanol for 20 min, followed by pelleting by centrifugation for 20 min at 4°C. The pellet was washed with 700 µl 70% ethanol and air dried at 37°C for 5 min prior to resuspension in RNase free water. The RNA isolation was followed by a DNase treatment using 2 units of TURBO DNase (Thermo Fisher) per 10 µg RNA for 30 min at 37°C in 100 µl total volume. Total RNA was then collected using the RNA Clean and Concentrator-25 kit (Zymo Research) according to the manufacturer’s instructions.
tRF&tiRNA-seq (CD Genomics)
The following treatments were performed before library preparation for total RNA samples: 3’-aminoacyl (charged) deacylation to 3’-OH for 3’ adaptor ligation, 3’-cP (2’,3’-cyclic phosphate) removal to 3’-OH for 3’ adaptor ligation, 5’-OH (hydroxyl group) phosphorylation to 5’-P for 5’-adaptor ligation, m1A and m3C demethylation for efficient reverse transcription. Sequencing libraries were size-selectedfor the RNA biotypes to be sequenced using an automated gel cutter.The libraries were qualified and absolutely quantified using Agilent BioAnalyzer2100.
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| Library strategy |
OTHER |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
Illumina NextSeq 500 |
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| Description |
50-bp single read sequencing at 10M reads
CEA17_24h_R3
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| Data processing |
*library strategy: tRF&tiRNA-seq
The sequencing results were analyzed with Unitas (https://doi.org/10.1186/s12864-017-4031-9) using default parameters.
A. fumigatus nuclear tRNAs were referenced from GtRNAdb, mitochondrial tRNAs were scanned from the mitochondrial genome using the 27 tRNAs predicted by tRNAscan-SE. All analyses were performed using “R” version 4.3.1 (2023-06-16) and Rstudio (2022-02-02), an integrated development environment (IDE) for R.
Assembly: Aspergillus fumigatus A1163 ASM15014v1
Supplementary files format and content: A single csv file (tiRNA_processed) contains the abundance of each target from each sample replicate.
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| Submission date |
May 14, 2024 |
| Last update date |
May 25, 2024 |
| Contact name |
Matthew George Blango |
| Organization name |
Leibniz Institute for Natural Product Research and Infection Biology
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| Department |
Junior Resaerch Group RNA Biology of Fungal Infections
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| Street address |
Adolf-Reichwein-Str. 23
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| City |
Jena |
| ZIP/Postal code |
07745 |
| Country |
Germany |
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| Platform ID |
GPL23268 |
| Series (1) |
| GSE267453 |
tRF & tiRNA sequencing of Aspergillus fumigatus |
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| Relations |
| BioSample |
SAMN41396874 |
| SRA |
SRX24550034 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
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