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| Status |
Public on Jun 26, 2013 |
| Title |
293T nt_control |
| Sample type |
SRA |
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| Source name |
293T cell line
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| Organism |
Homo sapiens |
| Characteristics |
treatment: non-targeting siRNA pool
cell line: 293T
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| Treatment protocol |
Cells were transfected with SMART-pool siRNAs (Dharmacon) using DharmaFECT1 reagent (Dharmacon), as recommended by the manufacturer. A non-targeting siRNA pool was used as a control. Cells were harvested 48 or 72 hours post transfection.
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| Growth protocol |
HeLa and 293T cell lines were maintained in Dulbecco's Modified Eagle Medium (DMEM) supplemented with 10% Fetal Bovine Serum (FBS) and antibiotics (penicillin/streptomycin). Neuro2A (N2A) cells were grown in DMEM supplemented with 10% FBS, sodium pyruvate, MEM, non-essential amino acids, and penicillin/streptomycin.
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| Extracted molecule |
polyA RNA |
| Extraction protocol |
Total RNA was extracted using Tri Reagent (Sigma) or RNeasy columns (Qiagen)
Standard Illumina library preparation according to manufacturer
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
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| Data processing |
Full genomic sequences for the analyzed species were downloaded from the Ensembl database. Full transcriptomic sequences for all species were downloaded from Ensembl. For each gene, the canonical transcript was selected for gene expression analysis based on BioMart.
For each species, we assembled all canonical transcript (as defined above) sequences. In order to calculate the effective number of unique mappable positions in each transcript (i.e. the effective length) we performed the following steps. For each read length k, we extracted the L-k+1 (L being the transcript length) k-mer sequences from each canonical transcript and then aligned the full set of k-mers against the respective genome using Bowtie, allowing for a maximum of two mismatches. k-mers with no or one unique genomic alignment were then likewise aligned back to the canonical transcriptome. For each transcript, the number of such k-mers having a unique transcriptomic alignment was determined. This corresponds to the transcript’s effective number of unique mappable positions for k-mer mRNA-Seq reads. For each sample, the corresponding mRNA-Seq data were aligned against the respective genome using Bowtie, allowing for a maximum of two mismatches. Reads with one unique genomic alignment were then aligned against the canonical transcriptome and, for each transcript, the number of reads with one unique transcriptomic alignment were counted. Gene expression levels were determined as reads per thousand mappable positions of target transcript sequence per million of reads, where the reads uniquely align to the analyzed transcriptome. This procedure for estimating gene expression levels is a corrected version of the widely used RPKM (reads per kilobase of target transcript sequence per million of total reads) metric, and is referred to as “cRPKM” [Labbe RM et al. Stem Cells (2012)].
Genome_build: hg19, mm9
Supplementary_files_format_and_content: Tab-delimited text files include cRPKM values (as defined above), as well as the total read count for each gene's canonical transcript
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| Submission date |
Mar 26, 2013 |
| Last update date |
May 15, 2019 |
| Contact name |
Manuel Irimia |
| E-mail(s) |
mirimia@gmail.com
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| Phone |
+34933160212
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| Organization name |
Centre for Genomic Regulation
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| Department |
Systems Biology Unit
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| Street address |
88 Dr. Aiguader
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| City |
Barcelona |
| State/province |
Barcelona |
| ZIP/Postal code |
08003 |
| Country |
Spain |
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| Platform ID |
GPL11154 |
| Series (2) |
| GSE45504 |
Muscleblind-like proteins regulate embryonic stem cell-specific alternative splicing and reprogramming II |
| GSE45505 |
Muscleblind-like proteins regulate embryonic stem cell-specific alternative splicing and reprogramming |
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| Relations |
| SRA |
SRX255055 |
| BioSample |
SAMN01992326 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM1105743_HsamRNA-CL_293T_cont_cRPKM.txt.gz |
240.5 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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