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| Status |
Public on Jun 30, 2020 |
| Title |
UK3_T |
| Sample type |
SRA |
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| Source name |
UPF1 KD 03 – total RNA seq
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| Organism |
Homo sapiens |
| Characteristics |
cell line: HeLa cells
sirna: UPF1 knockdowns
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| Treatment protocol |
siRNA transfection with Lullaby (Ozscience) for 72 hours, two siRNA transfections (20 nM)
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| Extracted molecule |
total RNA |
| Extraction protocol |
ribosome-protected fragments were isolated after snap-freezing of HeLa cells and Rnase I digestion using gel filtration columns. Gel purified fragments between 26 nt and 34 nt were isolated, rRNAs were removed using the RiboZero kit (Illumina). RNA-seq libraries were prepared from cells lysed similar as the ribosome profiling samples. totalRNAs was isolated from cleared lysates by addition of TriReagent as for the ribosome-protected fragments.
Sequencing libraries from ribosome footprints were generated as previously described (Aeschimann et al., 2015). Total RNA was used for library generation with the TruSeq Stranded mRNA Library Prep Kit (Illumina).
Libraries were sequenced on an Illumina HiSeq2500 generating 100 nt single-end reads.
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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 |
Read adapter was trimmed using cutadapt (https://cutadapt.readthedocs.io/en/stable/index.html v. 2.5, option ‘--match-read-wildcards’). Reads were retained if their length was between 26 and 35 (Ribosome profiling samples) or between 21 and 101 (Total RNA samples)
Reads were filtered for rRNA and tRNA contamination mapping them to human tRNA and human rRNA databases using bowtie2 (http://bowtie-bio.sourceforge.net/bowtie2/index.shtml v. 2.3.5, options: -p 2 -L 15 -k 20)
Remaining reads were mapped to human cDNA database using bowtie2 (v 2.3.5, options: -p 2 -L 15 -k 20 --no-unal)
Expression analysis was carried out using Salmon (https://salmon.readthedocs.io/en/latest/index.html v 0.13.0) using trimmed, filtered reads
For ribosome profiling, reads counts was performed using an in-house script (https://github.com/talponer/filterBam/blob/master/bed2counts)
Genome_build: GRCh38.p10, ENSEMBL 89
Supplementary_files_format_and_content: transcript counts, TXT
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| Submission date |
Mar 12, 2020 |
| Last update date |
Jun 30, 2020 |
| Contact name |
Rene Dreos |
| E-mail(s) |
rene.dreos@unil.ch
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| Phone |
0044 (0)1603 450793
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| Organization name |
UNIL
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| Department |
CIG
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| Lab |
Gatfield Lab
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| Street address |
Ecublens
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| City |
Lausanne |
| State/province |
VD |
| ZIP/Postal code |
1015 |
| Country |
Switzerland |
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| Platform ID |
GPL16791 |
| Series (1) |
| GSE143301 |
Ribosome profiling under ABCE1 depletion to study aberrant translation termination |
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| Relations |
| BioSample |
SAMN14364335 |
| SRA |
SRX7899618 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM4407916_TR09_UK3.quant.txt.gz |
1.8 Mb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Processed data provided as supplementary file |
| Raw data are available in SRA |
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