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| Status |
Public on May 26, 2020 |
| Title |
WT YCplac33 [SM042Fd] |
| Sample type |
SRA |
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| Source name |
Whole cells
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| Organism |
Saccharomyces cerevisiae |
| Characteristics |
tissue: whole cells
genotype/variation: WT BY4741
molecule subtype: Ribosome-protected RNA 54-68 nt
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| Treatment protocol |
Cells were rapidly vacuum filtered and flash frozen. Ribosome profiling samples were lysed in a freezer mill with lysis buffer. Total RNA for mRNA-Seq was extracted from whole cells.
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| Growth protocol |
Cells were generally grown to an OD of ~0.6 and harvested.
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| Extracted molecule |
total RNA |
| Extraction protocol |
Ribosome profiling lysates were clarified, treated with RNase I, and run over sucrose gradients to extract either the monosome or the disome peak. RNA was then extracted using SDS/hot phenol/chloroform and footprints of the specified size were retained by gel purification.
Briefly, RNAs were ligated to a universal DNA linker followed by reverse transcription. cDNAs were then circularized and PCR amplified. Ribosome profiling circularized cDNA was subject to ribosomal RNA subtraction using custom biotinylated oligos in some cases. In others, Ribo-Zero was used. RNA-Seq samples were prepared with the Illumina Truseq Stranded ribo-zero kit.
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| Library strategy |
OTHER |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
Illumina NovaSeq 6000 |
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| Data processing |
Library strategy: ribosome profiling
Base calling and demultiplexing was performed at the NHLBI DNA Sequencing and Genomics Core at the NIH Intramural program, Bethesda, MD.
Files were demultiplexed and then trimmed to the start of the linker sequence.
Reads were aligned to a library of non coding RNA genes and those not matching were retained.
Reads were then aligned to the genome and those not matching to a custom library of splice junctions by using Bowtie (Langmead et al., 2009). For footprints, reads that did not match in these steps were trimmed of consecutive 3' As and realigned. The aligned reads from all alignment rounds were pooled and normalized to units of reads per million (rpm) (by dividing by the total number of million mapped reads in a sample) to generate wig files.
Replicates or multiple sequencing lanes for a given sample were pooled together to create a single wig file for each condition to be used in downstream analysis.
Genome_build: R64-1-1
Supplementary_files_format_and_content: All files are fixedStep wig files (text format, each line representing one position in the genome).
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| Submission date |
Mar 30, 2020 |
| Last update date |
May 27, 2020 |
| Contact name |
Nicholas R Guydosh |
| E-mail(s) |
nicholas.guydosh@nih.gov
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| Organization name |
National Institutes of Health
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| Department |
NIDDK
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| Lab |
LBG
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| Street address |
8 Center Dr, Bldg 8, 220
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| City |
Bethesda |
| State/province |
MD |
| ZIP/Postal code |
20892 |
| Country |
USA |
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| Platform ID |
GPL27812 |
| Series (1) |
| GSE139036 |
Disome and trisome profiling reveal targets of ribosome quality control |
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| Relations |
| BioSample |
SAMN14488394 |
| SRA |
SRX8030822 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM4445993_end3SM042Fd_minus.wig.gz |
723.2 Kb |
(ftp)(http) |
WIG |
| GSM4445993_end3SM042Fd_plus.wig.gz |
745.2 Kb |
(ftp)(http) |
WIG |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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