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Sample GSM1115751 Query DataSets for GSM1115751
Status Public on Dec 15, 2013
Title Vivo rep3
Sample type SRA
 
Source name in vivo
Organism Saccharomyces cerevisiae
Characteristics strain: BY4741
culture condition: in vivo
Treatment protocol For in vivo samples, DMS was added directly to the growing yeast culture at 2-4% for 2-3 min. 25%BME, 30% isoamyl alcholol was used to quench the DMS. Cells were lyzed with SDS and RNA was isolated with hot acid phenol. For in vitro samples, total RNA was extracted from yeast cells , polyA mRNA was selected, denatured at 95C, refolded in RNA folding buffer for 30min at 30C, and then probed with DMS at 2-5% for 3-5min.
Growth protocol WT yeast were grown to exponential phase at 30C
Extracted molecule polyA RNA
Extraction protocol Total RNA was extracted using hot acid phenol. mRNA was extracted with Dynal beads(invitrogen)
mRNA was randomly fragmented using Ambion fragmentatin reagents, followed by PNK, and ligation using T4 K227Q truncated RNA ligase. Superscript III was used for reverse trancription , and circ ligase (epicentre) for circularization. Illumina adapters were introduced by 8-10 cycles of PCR.
 
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina HiSeq 2000
 
Description Sample 3
processed data files: Feb13_VivoAllextra_1_15_PLUS.wig,
Feb13_VivoAllextra_1_15_Minus.wig,
ACviv1mset1mag_sc-rrna.align18sRNAnoMiss.wig,
r25sDMS_ACviv1mset1mag_sc-rrna.align.wigFiveCount.wig
Data processing Illumina Casava1.7 software used for basecalling.
Alignments were performed with bowtie using the first 25 nt of each read
Reads were first aligned to the ribosomal RNA and the unaligned reads were then aligned to the genome
reads were filtered for unique map to the genome and no mismatches
reads were filtered for reverse transcriptase stops at As and Cs only
Genome_build: Saccharomyces cerevisiae assembly R61 (UCSC: sacCer2).
Supplementary_files_format_and_content: The position number directly indicates the DMS modified position starting at 1. Raw reads for each replicates were combined together in wiggle format. Separate files are provided for positive and negative strand maching reads.
Supplementary_files_format_and_content: For the ribosomal RNA, processed wig files are aligned to the ribosomal RDN locus (RDN18 is 18S rRNA and RDN25 is 25S rRNA).
 
Submission date Apr 05, 2013
Last update date May 15, 2019
Contact name Silvi Rouskin
E-mail(s) srouskin@wi.mit.edu
Organization name Whitehead Institute
Street address 455 Main Street
City Cambridge
State/province MA
ZIP/Postal code 02141
Country USA
 
Platform ID GPL13821
Series (1)
GSE45803 Genome-wide probing of RNA structure reveals active unfolding of mRNA structures in vivo
Relations
SRA SRX261047
BioSample SAMN01999524

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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