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Sample GSM1253089 Query DataSets for GSM1253089
Status Public on Oct 31, 2013
Title HM1.2
Sample type SRA
 
Source name yeast culture
Organism Saccharomyces cerevisiae x Saccharomyces paradoxus
Characteristics developmental stage: M1
replicate: 2
index 1: TGGCATA
Treatment protocol We sampled, washed with distilled water and snap-froze 10 ml cultures prior to RNA extraction.
Growth protocol We inoculated 100 ml of YPD (1% BactoTM yeast extract, 2% BactoTM peptone and 2% dextrose) in 250 ml Erlenmeyer flasks, and incubated the culture at 30ºC and 340 rpm for 15 hours. To sporulate cells we washed cells with water and resuspended the cells in 250 ml of SPO (1% potassium acetate, 0.1% BactoTM yeast extract, 0.05% dextrose) for a final concentration of 107 cells/ml. We incubated the cultures in 1 L baffled flasks at 30ºC at 340 rpm for 24 hours. We used distilled water for all media.
Extracted molecule total RNA
Extraction protocol We extracted total RNA from samples using Ambion RiboPure-Yeast Kit. We purified mRNA from total RNA using Ambion Micro PolyPurist Kit, and reversed transcribed mRNA into cDNA using polyA primers.
We used T4 DNA Polymerase (NEB), T4 Polynucleotide Kinase (NEB), and Taq Polymerase (NEB) for end repair of the sheered cDNA. We used T4 ligase (NEB) to ligate index adapters to the cDNA . We pooled each sample to an equal concentration and gel extracted 250 – 350 bp of ligated cDNA and used PCR to amplify the selected fragment to a final concentration of 1 nM. We submitted our final cDNA library to GTAC at Washington University for sequencing on two Illumina HiSeq lanes. At a run concentration of 6 pM
 
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina HiSeq 2000
 
Description hybrid S. MATa S. MATalpha, developmental stage M1, replicate 2
Data processing original base-calling,quality control and trimming of indexes was performed by GTAC at Washington University
aligned sequencing reads using Bowtie v 0.12.7 with parameters -v 1 -m 0
used lab perl script to map alignments to ORFs that are shared between S. cerevisiae and S. paradoxus
used lab perl script to filter out ORFs in which all samples had 0 reads
For allele specific variance stabiliting, use DESeq v1.8.3 with command estimateDispersions(cds, method="pooled-CR", modelFormula=count~stage*allele*species)
For allele non-specific variance stabiliting, use DESeq v1.8.3 with command estimateDispersions(cds, method="pooled-CR", modelFormula=count~stage*species)
Genome_build: parental and hybrid indexes created from S. cerevisiae S288c (SGD) and S. paradoxus N17 (SGRP) genomes
Supplementary_files_format_and_content: 2013.02.06.1.mm.0.MA.all.species.high.counts.allele.non: read counts for all samples, hybrid counts are allele-nonspecific, this is count data for DESeq
Supplementary_files_format_and_content: 2013.02.06.1.mm.0.MA.all.species.high.counts.allele.sp: read counts for all samples, hybrid counts are allele-specific in which "c" designates S. cerevisiae allele and "p" designates S. paradoxus allele, this is count data for DESeq
Supplementary_files_format_and_content: 2013.02.06.allele.non.vsd: variance stabilized data for all samples derived from DESeq, hybrid data is allele-nonspecific
Supplementary_files_format_and_content: 2013.02.07.allele.specific.hybrid.high.counts.vsd: variance stabilized data for all samples derived from DESeq, hybrid counts are allele-specific in which "c" designates S. cerevisiae allele and "p" designates S. paradoxus allele
 
Submission date Oct 28, 2013
Last update date May 15, 2019
Contact name Devjanee Swain Lenz
E-mail(s) ds394@duke.edu
Organization name Duke University
Street address 130 Bioscience Drive
City Durham
State/province NC
ZIP/Postal code 27708
Country USA
 
Platform ID GPL17601
Series (1)
GSE51809 Heterochronic meiotic misexpression in an interspecific yeast hybrid
Relations
BioSample SAMN02385574
SRA SRX369183

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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