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Sample GSM1216867 Query DataSets for GSM1216867
Status Public on Mar 18, 2014
Title dbr1.Δ _2
Sample type SRA
Source name S. pombe cells, dbr1.Δ
Organism Schizosaccharomyces pombe
Characteristics strain:
genotype: h+ ade6-M216 ura4-D18 leu1-32
Growth protocol Two biological replicates of wild-type (ED668) and dbr1.Δ cultures were grown in YES (yeast extracts plus supplements) media at 32°C till they reached OD600 0.4.
Extracted molecule total RNA
Extraction protocol Cells were harvested and total RNA was isolated by hot-phenol extraction, and RNA quality was assessed on a Bioanalyzer instrument (Agilent). Total RNA was treated with DNase (Turbo DNA-free by ambion), and thereafter 4 μg was treated with a beta version of Ribo-Zero™ Magnetic Gold Kit (Yeast) to deplete rRNAs.
RNA-seq libraries were prepared from rRNA-free RNA using a strand-specific library preparation protocol based on an early version of the Illumina TruSeq Small RNA Sample Prep Kit. In brief, rRNA-depleted RNA was fragmented to an average size of ~200nt. Fragmented RNA was 3’-de-phosphorylated with Antartic phosphatase and 5’-phosphorylated with polynucleotide kinase; this treatment prepares RNA fragments for subsequent ligation of Illumina RNA adaptors to their 5’ and 3’ ends using a 3’-RNA ligase and a T4 RNA ligase, respectively. First-strand cDNA was produced using a primer specific for the Illumina 3’-adaptor. The library was amplified with 15 PCR cycles using primers specific for the Illumina adaptors and purified using SPRI-beads (Agencourt, Beckman Coulter). Library size distributions and concentrations were determined on a Bioanalyzer (Agilent). RNA-seq libraries were sequenced on an Illumina HiSeq 2000 instrument at the Core Facility of the Huntsman Cancer Institute (University of Utah).
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina HiSeq 2000
Description dbr1_2 (Name in ExpressionTable.txt)
total RNA (rRNA depleted)
Strand specific RNA-Seq
Data processing Sequence reads of 49 base length originating from each sample were aligned, using Bowtie 0.12.725, to the S. pombe genome sequence (Ensembl S. pombe, Build EF1, version 13) and to the corresponding exon-exon junctions database. Up to 3 base-pair mismatches were allowed. Reads that matched multiple loci were removed from further analysis, and the resultant alignment files were processed to generate ‘pile-ups’ against each chromosome. Unmapped reads were used for lariat mapping,.
Searches were performed against the genome sequence combined with a dataset of known exon-exon junctions as defined by Ensembl S. pombe, release-13. To ensure that a 49-base read mapped to a splice junction, only the last 43 bases of the first exon and the first 43 bases of the second exon were considered (if the exon exceeded length 43). In this way, reads that overlapped a junction by less than 6 nucleotides were excluded. Reads that matched to more than one junction or elsewhere in the genome were also discarded.
Genome_build: Ensembl S. pombe, Build EF1, version 13
Supplementary_files_format_and_content: Expression table and RPKM summary for all samples in tab-delimited file ExpressionTable.txt (linked as supplementary file on Series record)
Submission date Aug 27, 2013
Last update date May 15, 2019
Contact name Danny Asher Bitton
Phone +44(0)203-108-1604
Organization name University College London
Department Department of Genetics, Evolution & Environment
Lab Genome Regulation/Bähler Lab
Street address Darwin Building, Gower Street
City London
ZIP/Postal code WC1 6BT
Country United Kingdom
Platform ID GPL13988
Series (1)
GSE50246 LaSSO, a strategy for genome-wide mapping of intronic lariats and branch-points using RNA-seq
BioSample SAMN02335224
SRA SRX340112

Supplementary data files not provided
SRA Run SelectorHelp
Raw data are available in SRA
Processed data are available on Series record

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