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| Status |
Public on Mar 25, 2015 |
| Title |
Rot1mut_spb2011_1 |
| Sample type |
SRA |
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| Source name |
Rot1mut_spb2011_1
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| Organism |
Schizosaccharomyces pombe |
| Characteristics |
strain: spb2011
genotype/variation: Rot1-Q472Stop, ade6-hp+
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| Growth protocol |
Strains grown at 30 degress in YES medium to OD600=0.5
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| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated from exponentially growing cells using the hot phenol method (Leeds, 1991). The RNA was fractionated using RNeasy Midi columns (Qiagen) following the „RNA cleanup protocol“ provided by the manufacturer. The flow-through fraction was precipitated („small RNA“ fraction). 25 μg of the "small RNA" fraction were separated on a 17.5% PAGE and the 18 – 28 nt population was purified.
The isolated small RNA populations were prepared for sequencing using the Illumina® TruSeq small RNA preparation protocol (Cat.# RS-930-1012).
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| Library strategy |
miRNA-Seq |
| Library source |
transcriptomic |
| Library selection |
size fractionation |
| Instrument model |
Illumina HiSeq 2000 |
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| Data processing |
All samples were sequenced (together with other libraries) on one lane of an HiSeq 2000 instrument. Individual reads were assigned to their sample based on the TruSeq barcode. Illumina software Casava v 1.8 was used. The 3' adapter was removed by aligning it to the read allowing one or two mismatches in prefix alignments of at least 7 or 10 bases respectively. Low complexity reads were filtered out based on their dinucleotide entropy (removing <1% of the reads). All the reads that were shorter than 14 nucleotides were removed.
Alignments to the S. pombe genome (May 8, 2009, http://www.sanger.ac.uk/Projects/S_pombe/) were performed for the minus strand by the software bowtie (version 0.9.9.1) ( Langmead et al., 2009) with parameters -v 2 -a -m 100, tracking up to 100 best alignment positions per query and allowing at most two mismatches. Alignments were normalized for the size of the each library, transformed into log2 scale and corrected with a pseudocount of 3. Annotations were used as described in Woolcock et al., Genes Devc 2012, PMID: 22431512
Genomic read coverage plots were made created separately for the plus (wiggle files with _p_) and minus (wiggle files with _m_) strand, transformed into log2 scale and normalized for the size of each library based on weights of the alignments
Genome_build: May 8, 2009, http://www.sanger.ac.uk/Projects/S_pombe/
Supplementary_files_format_and_content: wiggle files containing log2 scaled and normalized coverage values
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| Submission date |
Jul 07, 2014 |
| Last update date |
May 15, 2019 |
| Contact name |
Tim Roloff |
| E-mail(s) |
tim.roloff@fmi.ch
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| Organization name |
FMI
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| Department |
Functional Genomics
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| Street address |
Maulbeerstrasse 66
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| City |
Basel |
| ZIP/Postal code |
4058 |
| Country |
Switzerland |
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| Platform ID |
GPL13988 |
| Series (2) |
| GSE59170 |
RNA expression profiling for Paf1 complex mutant strains [miRNA] |
| GSE59171 |
RNA expression profiling for Paf1 complex mutant strains |
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| Relations |
| BioSample |
SAMN02904626 |
| SRA |
SRX647472 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM1429723_spb2011_m_log_libnorm.wig.gz |
849.1 Kb |
(ftp)(http) |
WIG |
| GSM1429723_spb2011_p_log_libnorm.wig.gz |
823.1 Kb |
(ftp)(http) |
WIG |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
| Processed data are available on Series record |
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