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| Status |
Public on Nov 28, 2017 |
| Title |
Strand-specific RNA-seq in siCtrl-treated proliferative WI38 hTERT RAF1-ER cells- Replicate #2 |
| Sample type |
SRA |
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| Source name |
siCtrl-treated proliferative WI38 hTERT RAF1-ER cells
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| Organism |
Homo sapiens |
| Characteristics |
cell line: WI38 hTERT RAF1-ER
state: proliferative
treatment: transfected with control siRNA for 3 days
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| Treatment protocol |
Transfection with siRNA during three days.
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| Growth protocol |
Proliferative WI38 hTERT RAF1-ER cells
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| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA were prepared using the MasterPure RNA Purification Kit (Epicentre Biotechnologies) supplemented with Baseline-ZERO DNAse (Epicentre). BGI or EMBL-GeneCore treated the RNA by Ribozero kit to remove ribosomal RNA.
Strand-specific RNA-seq, including the library construction, which is based on UTP incorporation in the second strand cDNA, was performed at BGI (HONG KONG) for the first replicates (lncRNA-seq) and at EMBL-GeneCore (Heidelberg, Germany) for the second replicates (stranded rRNA-minus RNA-seq).
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
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| Description |
total RNA (ribosomal RNA depleted)
Strand-specific RNA-sequencing in proliferative cells treated with Control siRNA was performed paired-end (HMKMGBGXY_siCtrl_16s005987-1-1_Nicolas_lane116s005987_1_sequence.txt and HMKMGBGXY_siCtrl_16s005987-1-1_Nicolas_lane116s005987_2_sequence.txt) by Illumina’s NextSeq 500 technology at EMBL-GeneCore (stranded rRNA-minus RNA-seq) with ~100 millions paired reads per library. After alignment on human genome hg38 using Spliced Transcripts Alignment to a Reference (STAR) version 2.5.2a_modified, depending on the strand of the genome, 2 files were generated (HMKMGBGXY_siCtrl_16s005987-1-1_Nicolas_lane116s005987_minus_normalized_hg38.bw and HMKMGBGXY_siCtrl_16s005987-1-1_Nicolas_lane116s005987_plus_normalized_hg38.bw).
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| Data processing |
Paired-end reads of the strand-specific RNA-Seq (siCtrl or siH2A.Z) data were aligned using Spliced Transcripts Alignment to a Reference (STAR) version 2.5.2a_modified. For these alignments all parameters were kept as default.
After the alignment, we applied on all aligned datasets several steps using samtools software: we converted aligned file from Sequence Alignments/Map (sam) format into the Binary Alignment/Map (bam) format which stores the same data in a compressed, indexed, binary form and a cutoff was applied to keep only reads with MAPQ > 25. We sorted data by position on the genome, and created an index of each bam file (bai format). We then converted these cleaned files in wiggle files using R via the rtracklayer bioconductor package. On each dataset, we applied for each base position of the genome a normalization factor (100,000,000 / total numbers of aligned reads).
Note that, for the 1st replicates of siCtrl and siH2A.Z samples, although strand-specific RNA-seq has been performed, we found that the data did not show strand-specificity. We thus only analyzed for this dataset intervals, which do not show evidence of transcription on the 2 strands of DNA, such as the intergenic regions of the convergent gene pairs to analyze START RNA expression.
Genome_build: hg38
Supplementary_files_format_and_content: BigWig
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| Submission date |
Sep 13, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Estelle Nicolas |
| E-mail(s) |
estelle.nicolas@univ-tlse3.fr
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| Phone |
+33 5 61 55 81 11
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| Organization name |
INSERM
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| Lab |
UMR5077, MCD, CBI
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| Street address |
118 route de Narbonne
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| City |
Toulouse |
| ZIP/Postal code |
31062 |
| Country |
France |
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| Platform ID |
GPL18573 |
| Series (2) |
| GSE85083 |
Control of gene expression in senescence through transcriptional read-through of convergent protein-coding genes [RNA-Seq siRNA H2A.Z] |
| GSE85085 |
Control of gene expression in senescence through transcriptional read-through of convergent protein-coding genes |
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| Relations |
| BioSample |
SAMN07638085 |
| SRA |
SRX3182088 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM2781957_HMKMGBGXY_siCtrl_16s005987-1-1_Nicolas_lane116s005987_minus_normalized_hg38.bw |
313.1 Mb |
(ftp)(http) |
BW |
| GSM2781957_HMKMGBGXY_siCtrl_16s005987-1-1_Nicolas_lane116s005987_plus_normalized_hg38.bw |
324.4 Mb |
(ftp)(http) |
BW |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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