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| Status |
Public on Jan 03, 2024 |
| Title |
mRNA eIF2A∆ replicate 2 |
| Sample type |
SRA |
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| Source name |
yeast cell
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| Organism |
Saccharomyces cerevisiae |
| Characteristics |
cell type: yeast cell
genotype: eIF2A{delta}
strain: F2247 (MATa his3{delta}1 leu2{delta}0 met15{delta}0 ura3{delta}0 ygr054w{delta}::kanMX4)
treatment: control
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| Treatment protocol |
No additional treatment
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| Growth protocol |
Yeast cells were generally grown to an OD600 of 0.5-0.6 in synthetic complete medium at 30°C.
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| Extracted molecule |
total RNA |
| Extraction protocol |
For RNA-seq library preparation, total RNA was extracted and purified from aliquots of the same snapped-frozen cells described above using hot phenol-chloroform extraction. Five µg of total RNA was randomly fragmented by mixing with 2× alkaline fragmentation solution (2 mM EDTA, 10 mM Na2CO3, 90 mM NaHCO3, pH ≈ 9.3). After incubation for 20 min at 95ºC, fragmentation reactions were mixed with ice-cold stop solution (300 mM NaOAc pH 5.5, 30 µg GlycoBlue (Invitrogen; AM9516)). RNA was precipitated by adding one part of isopropanol followed by the standard protocol of precipitation. Fragment size selection, library generation, and sequencing were carried out using the same protocol described above for RPF library preparation, except the Ribo-Zero Gold rRNA Removal Kit (Illumina; MRZ11124C), was employed to remove rRNA after linker-ligation.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
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| Description |
DESeq2 eIF2Adelta vs WT.xlsx and DESeq2 eIF2Adelta+SM vs eIF2Adelta.xlsx
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| Data processing |
For ribo-seq samples, Fastq records are trimmed with fastxtoolkit (version 0.0.14) to remove adapters sequences. Subsequently, rRNA sequence was removed with Bowtie2 (version 2-2.3.5.1) then mapped to yeast genome (sacCer3) with tophat (version 2.1.1). Perfect reads were extracted with samtools (version 1.9) for downstream analysis. Normalized wiggle files and reads counts are generated with RiboSeq tools (https://github.com/hzhanghenry/RiboProR). For RNA-seq samples, the same protocol mentioned above for ribo-seq analysis was followed.
Translational efficiency change analysis was performed with R DESeq2 package.
Assembly: sacCer3
Supplementary files format and content: The DESeq2 files to represent fold-change and raw reads that were used as input for the fold-change analysis
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| Submission date |
Aug 22, 2023 |
| Last update date |
Jan 03, 2024 |
| Contact name |
Swati Gaikwad |
| Organization name |
National Institutes of Health
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| Department |
Section on Nutrient Control Of Gene Expression
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| Lab |
Hinnebusch, Alan
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| Street address |
Building 6, Room 233
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| City |
Bethesda |
| State/province |
MD |
| ZIP/Postal code |
20892-2716 |
| Country |
USA |
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| Platform ID |
GPL17342 |
| Series (1) |
| GSE241473 |
Yeast eIF2A plays a minimal role in translation initiation in vivo |
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| Relations |
| BioSample |
SAMN37116712 |
| SRA |
SRX21459046 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
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