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| Status |
Public on Jul 12, 2021 |
| Title |
set3_calu3_moi10_36hpi_RPF_rep1 |
| Sample type |
SRA |
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| Source name |
Epithelial cell in human lung adenocarcinoma
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| Organisms |
Homo sapiens; Severe acute respiratory syndrome coronavirus 2 |
| Characteristics |
cell line: Calu-3
treatment: infected, 36hpi
concentration: MOI 10
rnase: 5U
assay: RPF-seq
biorep: rep1
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| Treatment protocol |
Prior to lysis, Calu-3 and Vero cells were seeded in 100mm dishes, and were infected with SARS-CoV-2 (MOI = 10 or 0.1) for 1 hour then washed with PBS. At 0, 1, 2, 4, 12, 16, 24, 48 hpi (Calu-3) and 0, 24 hpi (Vero), both mock and infected dishes for RPF, QTI, and mRNA-seq were treated with either 100 μg/ml cycloheximide (CHX) or 2 µg/ml harringtonine (Harr) for 10 min.
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| Growth protocol |
Calu-3 and Vero cells were cultured in DMEM with 10% FBS and 1% penicillin/streptomycin
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| Extracted molecule |
total RNA |
| Extraction protocol |
For sRNA-seq, RNA was extracted using Trizol. For rest, cells were washed with PBS then lysed with lysis buffer (10 mM Tris-HCl (pH 7.4), 5 mM MgCl2, 100 mM KCl, 1% Triton X-100, 100 μg/ml CHX, 1 mM dithiothreitol, 0.2 U/μl RiboLock RNase inhibitor, and 1 × EDTA-free protease inhibitor cocktail). For RPF-seq and QTI-seq, lysates were treated with RNase I and ribosome protected fragments were isolated using illustra MicroSpin S-400 HR Columns (GE Healthcare).
All of the libraries (mRNA-seq, RPF-seq, QTI-seq, and sRNA-seq) were constructed following the Illumina Truseq small RNA library preparation protocol (TruSeq Small RNA Library Prep Guide, Part # 15004197 Rev. G)
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| Library strategy |
OTHER |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
Illumina HiSeq 2500 |
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| Description |
RPF
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| Data processing |
1. Base-calling by RTA(Real Time Analaysis) software within HiSeq 2500 (mRNA-Seq) or HiSeq 2000 (sRNA-Seq)
2. Quality trimming by the highest scoring path algorithm (hsptrim-1.2.6)
3. Adapter trimming by cutadapt-1.2.1
4. Remove sequencing artifacts by fastx_artifacts_filter of fastx_toolkit
5. Remove ncRNA-derived reads by Bowtie2 alignment
6. Remove mycoplasma-derived reads by Bowtie2 alignment
7. Alignment of filtered reads to SARS-CoV-2 and host genome by STAR (mRNA-seq, RPF-seq, QTI-seq) or Bowtie2 (sRNA-seq)
8. Calculation of log2(expression fold changes) and differential expression for human genes by DESeq2
Genome_build: NC_045512.2 (SARS-CoV-2)
Genome_build: hg38 (GRCh38)
Genome_build: chlSab2
Supplementary_files_format_and_content: bigwig (coverage depth), tab-delimited text files, containing gene expression levels (average RPKM within replicates), log2(expression fold changes), and statistical significances calculated by DESeq2.
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| Submission date |
Sep 21, 2020 |
| Last update date |
Sep 30, 2021 |
| Contact name |
Daehyun Baek |
| E-mail(s) |
baek@snu.ac.kr
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| Organization name |
Seoul National University
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| Department |
School of Biological Sciences
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| Lab |
The Baek Research Group of Computational Biology
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| Street address |
Rm 402, Bldg 504, School of Biological Sciences, Seoul National University, 1 Gwanak-ro, Gwanak-gu, Seoul
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| City |
Seoul |
| ZIP/Postal code |
08826 |
| Country |
South Korea |
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| Platform ID |
GPL29105 |
| Series (1) |
| GSE157490 |
A high-resolution temporal atlas of the SARS-CoV-2 translatome and transcriptome |
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| Relations |
| BioSample |
SAMN16230160 |
| SRA |
SRX9162256 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM4796139_RF0042_004.virus.minus.bw |
68.0 Kb |
(ftp)(http) |
BW |
| GSM4796139_RF0042_004.virus.plus.bw |
181.7 Kb |
(ftp)(http) |
BW |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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