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| Status |
Public on Jun 17, 2012 |
| Title |
dcp2D_xrn1D_rep2 |
| Sample type |
SRA |
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| Source name |
dcp2D::HIS3 xrn1D::kanMX
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| Organism |
Saccharomyces cerevisiae |
| Characteristics |
strain: W303-1A
genotype/variation: dcp2D::HIS3 xrn1D::kanMX
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| Growth protocol |
Cells were grown to mid-log phase (OD600 = 0.3) in YPD media at 30˚C.
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| Extracted molecule |
polyA RNA |
| Extraction protocol |
RNA was extracted from frozen cells by a hot phenol method. Polyadenylated RNA was selected using Oligo (dT) DynaBeads (Invitrogen). A 5' RNA adaptor that includes a recognition site for the MmeI restriction enzyme was selectively ligated to 5' monophosphorylated species using T4 RNA ligase (Ambion). The ligated RNA was reverse-transcribed using oligo(dT) linked to a 3' adaptor sequence, PCR-amplified for 10 cycles and treated with MmeI, which cleaves 20 or 21 nucleotides 3' of the recognition site. The digestion products were gel-purified, ligated to a 3' double-strand DNA adaptor and again gel-purified. The resultant material was PCR-amplified, gel-purified and subject to high-throughput Illumina sequencing.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
RACE |
| Instrument model |
Illumina HiSeq 2000 |
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| Data processing |
Read sequences composed of 20- or 21-nucleotide-long captured RNA sequences followed by the 3' Illumina adaptor sequence were selected using fastx_barcode_splitter.pl in FASTX-Toolkit (http://hannonlab.cshl.edu/fastx_toolkit/download.html).
The file of the barcode-matched reads was converted to a fasta file containing reads from which the adaptor sequences were removed using Perl.
Identical sequences were collapsed and counted using fastx_collapser in FASTX-Toolkit to generate a file of distinct sequences and occurrence counts.
The distinct sequences were aligned to reference sequences using Bowtie 0.12.7 with parameters -f -m 1. In this setting, two mismatches were allowed (default), and alignments were required to be unique.
The abundance of each sequence read was plotted as a function of its position in 6,603 yeast mRNA using Perl. In this process, sequence reads that were aligned to the reference with no mismatch were used. ORF annotations were downloaded from SGD (http://downloads.yeastgenome.org/chromosomal_feature/ SGD_features.tab) on May 11, 2012. UTR lengths were taken from Nagalakshmi et al. (2008) Science, 320:1344. Where UTR annotation was not available, the ORF start/end was considered to be the transcript start/end. For each sample, read counts were normalized to the total reads mapping to all mRNAs.
Genome_build: S288C_reference_genome_R64-1-1_20120203, A364A 2 micron plasmid sequence (NCBI accession number NC_001398), dsRNA virus genome (killer virus M1, virus L-A and L-BC, single stranded viruses 20S and 23S; NCBI accession numbers NC_001782, NC_003745, NC_001641, NC_004051 and NC_004050, respectively), and the sequence of the kanMX6 cassette (Longtine et al., 1998, Yeast, 14:953)
Supplementary_files_format_and_content: transcript profiles: The tab-delimited files contain read counts at all nucleotide positions in 6,603 protein-coding transcripts in Saccharomyces cerevisiae. Each line represents one transcript. The first element of each line represents the gene name. The UTR annotations were taken from Nagalakshmi et al. (2008) Science, 320:1344. When UTR annotation was not available, the ORF start/end was considered to be the transcript start/end.
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| Submission date |
May 30, 2012 |
| Last update date |
May 15, 2019 |
| Contact name |
Yuriko Harigaya |
| Organization name |
University of Colorado at Boulder
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| Department |
Department of Chemistry and Biochemistry
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| Lab |
Roy Parker Lab
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| Street address |
3415 Colorado Ave, UCB 596
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| City |
Boulder |
| State/province |
Colorado |
| ZIP/Postal code |
80303 |
| Country |
USA |
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| Platform ID |
GPL13821 |
| Series (1) |
| GSE33712 |
Global analysis of mRNA decay intermediates in Saccharomyces cerevisiae |
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| Relations |
| SRA |
SRX150816 |
| BioSample |
SAMN01001264 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM940397_dcp2D_xrn1D_profiles2.txt.gz |
360.4 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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