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Sample GSM2148032 Query DataSets for GSM2148032
Status Public on May 11, 2016
Title J49Lib41
Sample type SRA
 
Source name yeast strain, 30-rep2-12
Organism Saccharomyces cerevisiae
Characteristics strain: 12
time point: 30
single-mixed original culture: Single culture
Treatment protocol Samples were rapidly centrifuged to eliminate the must fractions and RNAlater (Ambion, Monza) was added and immediately frozen in liquid nitrogen. Samples were kept at -80ーC until flow cytometry separation. Cells were sorted with a MoFlo (Dako cytomation, Glostrup, Denmark) cytometer, on the basis of labels according to fluorescence intensities of the populations. The cytometer was equipped with two lasers: 488nm (for GFP exitation) and 345 nm (for Sapphire exitation). For GFP and Sapphire FL7 570/20 and FL1 530/20 channels were respectively used. Separation was performed with a sorting rate up to 20000 events per second for the time necessary to reach between 500000 and 1000000 cells. Cells were collected in tubes containing PBS buffer and directly put in dry ice to flash freeze them at the exit of the cytometer. Frozen samples were kept at -80ーC and thawed in ice for the subsequent RNA extraction
Growth protocol Single and mixed fermentations, including strain-dye swap, were performed in triplicate. Dominant and non-dominant strains were inoculated reaching the same cell density (0.1 OD) in 10 ml of Nebbiolo natural must (Fig. 1A). Fermentations were carried out at 25°C with continuous shaking. Samples of 1 ml were taken at 3 points of the fermentation. The first sample was taken at the beginning of the exponential growth phase, the second in the middle of the exponential growth phase and the last at the beginning of the stationary phase.
Extracted molecule total RNA
Extraction protocol The RNA Mini Kit (Qiagen, Madrid) was used following manufacturer’s instructions. RNA quality was checked using the Bioanalyser (Agilent Technologies Spain, Madrid) and samples with good quality RNA, showing 5 as minimum of RNA Integrity Number (RIN) were sequenced
Illumina RNA amplification kit (Thermo-Fisher) was used in order to amplify the low amount of RNA extracted from cells separated by cytometry. Sequencing was performed with the 5500 SOLiD system (Life Technologies, Madrid). Libraries were performed using standard SOLID protocol.
 
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model AB 5500 Genetic Analyzer
 
Data processing Sequencing was performed with the 5500 SOLiD system (Life Technologies, Madrid). Seventy-eight million reads per strain were obtained and short sequences (75bp) were assembled and aligned using a reference S. cerevisiae strain (Saccer2). Values of FPKM (Fragments Per Kilobase of exon per Million fragments mapped) were transformed for the statistical test as log2 (FPKM1/FPKM2) and uploaded on Cuffdiff (Cufflinks V1.3.0, http://cufflinks.cbcb.umd.edu/) where FPKM1 represents data of 12 strain and FPKM2 represent data of 11 strain. The FPKM parameter was used for pair end RNA-Seq experiments, where fragments were sequenced from both ends, providing two reads for each fragment. The Q value, representing significance value for statistical analysis performed through the use of cufflink platform was calculated and significant transcriptome differences were identified
 
Submission date May 10, 2016
Last update date May 15, 2019
Contact name Roberto Pérez
E-mail(s) rober@iata.csic.es
Phone 0034963900022
Organization name Centro Superior de Investigaciones Científicas
Street address Av Agustín Escardino 7
City Paterna
State/province Valencia
ZIP/Postal code 46980
Country Spain
 
Platform ID GPL16234
Series (1)
GSE81270 Transcriptome of two S. cerevisiae strain in competition in natural must
Relations
BioSample SAMN04966328
SRA SRX1753815

Supplementary file Size Download File type/resource
GSM2148032_J49Lib41.coverage.wig.tar.gz 13.9 Mb (ftp)(http) TAR
GSM2148032_J49Lib41.exon.counts.gtf.gz 130.6 Kb (ftp)(http) GTF
GSM2148032_J49Lib41.gene.counts.tab.gz 129.0 Kb (ftp)(http) TAB
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file
Processed data are available on Series record

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