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GEO help: Mouse over screen elements for information. |
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| Status |
Public on Sep 25, 2014 |
| Title |
Murine activated T cells (Replicate 1) |
| Sample type |
SRA |
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| Source name |
T cells
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| Organism |
Mus musculus |
| Characteristics |
strain: C57BL/6
cell type: T cells
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| Treatment protocol |
For mouse T cells, spleen and lymph nodes were dissected from 8-10 week old female C57BL/6 mice and total, untouched T cells were isolated by MACS purification (Pan T cell isolation kit; Miltenyi) according to the manufacturer's protocol. T cells were activated for 72 hrs with mouse T-Activator CD3/CD28 Dynabeads (Gibco/Life Technologies) and 30 Units of recombinant IL-2 (Peprotech). Corresponding naive cells were from the same T cell preparations. Human T cells from single donor human blood samples were isolated with the Pan T cell isolation kit from Miltenyi and either left untreated or were stimulated with human T-Activator CD3/CD28 Dynabeads (Life Technologies) and 30 Units of recombinant IL-2 from Peprotech.
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| Extracted molecule |
total RNA |
| Extraction protocol |
Selection for poly(A)+ RNA (for details see A-seq protocol)
A-seq protocol (see Martin G, Gruber AR, Keller W, Zavolan M (2012) Genome-wide analysis of pre-mRNA 3’ end processing reveals a decisive role of human cleavage factor I in the regulation of 3' UTR length. Cell Rep 1: 753–763.)
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
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| Description |
3' end sequencing of murine activated Tcells
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| Data processing |
For the murine A-seq libraries, reads were pre-processed to remove the six diagnostic adenosines derived from the poly(A) tail as well as the 3' adaptor sequence and then were mapped to the mouse genome (mm9) and annotated using the CLIPZ server. For the human A-seq libraries, only reads that contained three T residues at positions 5-7 (indicating the beginning of the poly(A) tail) were selected for further use. From these, randomized nucleotides at positions 1-4 at the 5’ end of the reads (needed for cluster coordination in Illumina sequencing) were trimmed together with the three Ts, thus removing seven nucleotides. The reverse complement of the remaining sequences, presumably representing mRNA 3’ ends, were then mapped to the human genome (hg19). For the construction of the set of 3' end cleavage sites, we only used reads that had the adaptor removed, mapped uniquely to genomic regions. Based on the precise mapping of the 3' ends of these reads we computed putative cleavage sites with their associated abundance. To minimize the frequency of internal priming sites in our data set, we discarded those that in the eight-nucleotide-region immediately downstream of the putative cleavage site had at least seven A nucleotides. The raw reads were processed on the CLIPZ server (http://http://www.clipz.unibas.ch; Khorshid et al., Nucleic Acids Research 2011, 39:D245).
Genome_build: hg19 and mm9
Supplementary_files_format_and_content: gzipped BED files with inferred 3' ends of Pol II transcripts; column 5 represents the number of reads
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| Submission date |
Feb 12, 2014 |
| Last update date |
May 15, 2019 |
| Contact name |
Andreas R Gruber |
| E-mail(s) |
agruber@tbi.univie.ac.at
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| Organization name |
University of Basel
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| Department |
Biozentrum
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| Lab |
Zavolan
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| Street address |
Klingelbergstrasse 50-70
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| City |
Basel |
| ZIP/Postal code |
4056 |
| Country |
Switzerland |
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| Platform ID |
GPL13112 |
| Series (1) |
| GSE54950 |
The systematic shortening of 3’ UTRs has limited influence on the relative change in protein abundance in proliferating cells |
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| Relations |
| BioSample |
SAMN02641043 |
| SRA |
SRX470073 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM1327167_mm9.sample1.stim.sites.bed.gz |
9.2 Mb |
(ftp)(http) |
BED |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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