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| Status |
Public on Feb 07, 2023 |
| Title |
RNA_pat1dhh1_Replicate_2 |
| Sample type |
SRA |
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| Source name |
Whole cells
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| Organism |
Saccharomyces cerevisiae |
| Characteristics |
genotype: W303 F2182 (MATa ura3-1 trp1-1 ade2-1 leu2-3,112 his3-11,15 delta-pat1::HIS3 delta-dhh1::KanR)
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| Growth protocol |
In additional columns of the SAMPLES section
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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 Zymo RNA Clean and Concentrator kit. 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.
In additional columns of the SAMPLES section
In additional columns of the SAMPLES section
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
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| Data processing |
For RNA-Seq: As described earlier (Martin-Marcos et al. 2017), Illumina sequencing reads were trimmed to remove the constant adapter sequence, mixed sample sequences were separated by the sample barcodes followed by removal of PCR duplicates using a custom Python (3.7) script. The sequences aligned to yeast non-coding RNAs were removed using bowtie (Langmead et al., 2009) and non-rRNA reads (unaligned reads) were then mapped to the S. cerevisiae genome (R64-1-1 S288C Saccer3 Genome Assembly) using TopHat (Trapnell et al., 2009). Only uniquely mapped reads from the final genomic alignment were used for subsequent analyses. Normalized wiggle files and reads counts are generated with RiboSeq tools (https://github.com/hzhanghenry/RiboProR). For RNA-seq, the same protocol mentioned above for ribo-seq analysis was followed.
For ChIP-Seq: Sequence data were aligned to the SacCer3 version of the genome sequence using Bowtie2 with parameters -X 1000 -very-sensitive, to map sequences up to 1 kb with maximum accuracy. PCR duplicates from ChIP-seq data were removed using the samtools rmdup package.
Assembly: sacCer3
Supplementary files format and content: Processed data (csv) files contain raw reads for each file. For RNA-Sequencing, each file contains five columns providing: systemmatic gene name (Gene_ID), total reads (trx), reads in CDS, reads in 5' UTR and reads in 3' UTR. Foe CAGE-sequencing and parallel RNA-Sequencing, each file contains two columns providing: systemmatic gene name (Gene_ID), total reads. For Rpb1 ChIP-Seq, average occupancy file contains five columns providing: systemmatic gene name (ORF), avg occupancy at -1, NDR, +1 and average of -1, NDR, +1
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| Submission date |
Dec 08, 2022 |
| Last update date |
Feb 07, 2023 |
| Contact name |
Alan G Hinnebusch |
| E-mail(s) |
alanh@mail.nih.gov
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| Organization name |
National Institutes of Health
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| Department |
Eunice Kennedy Shriver National Institute of Child Health and Human Development
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| Lab |
Section on Nutrient Control of Gene Expression
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| Street address |
6 Center Drive
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| City |
Bethesda |
| State/province |
Maryland |
| ZIP/Postal code |
20892 |
| Country |
USA |
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| Platform ID |
GPL17342 |
| Series (1) |
| GSE220578 |
Decapping factor Dcp2 controls mRNA abundance and translation to adjust metabolism and filamentation to nutrient availability II |
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| Relations |
| BioSample |
SAMN32118017 |
| SRA |
SRX18543679 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM6807048_RawCounts_RNA_pat1dhh1_Replicate_2.csv.gz |
55.2 Kb |
(ftp)(http) |
CSV |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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