|
| Status |
Public on Dec 03, 2016 |
| Title |
S. cerevisiae x S. kudriavzevii W27 replicate C, 12ºC fermentation against 28ºC fermentation. |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
yeast cells from wine fermentation at 28ºC
|
| Organism |
Saccharomyces cerevisiae x Saccharomyces kudriavzevii |
| Characteristics |
strain: W27
growth phase: Beginning of exponential phase
|
| Growth protocol |
Wine fermentations in Tempranillo must at 12ºC or 28ºC
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA extraction method was based on consecutive treatments with phenol-tris, phenol-chloroform (5:1) and chloroform-isoamyl alcohol (24:1) and a final precipitation with ethanol and sodium acetate (Garcia-Martinez et al., 2004).
|
| Label |
Cy3
|
| Label protocol |
2-4 ug of total RNA was linearly amplified using the Low RNA Input Fluorescent Linear Amplification kit (Agilent Technologies™, Ca, USA). 2-3 ug of amplified cRNA was used as template for cDNA synthesis. cDNA was marked indirectly with “SuperScript™ Indirect cDNA Labeling System” (Invitrogen™, SanDiego, CA). The fluorophores used were Cy3 and Cy5 mono-reactiveDye (Amersham GE Healthcare™, Amersham UK) and dye incorporation was monitored by a Nanodrop spectrophotometer.
|
| |
|
| Channel 2 |
| Source name |
yeast cells from wine fermentation at 12ºC
|
| Organism |
Saccharomyces cerevisiae x Saccharomyces kudriavzevii |
| Characteristics |
strain: W27
growth phase: Beginning of exponential phase
|
| Growth protocol |
Wine fermentations in Tempranillo must at 12ºC or 28ºC
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA extraction method was based on consecutive treatments with phenol-tris, phenol-chloroform (5:1) and chloroform-isoamyl alcohol (24:1) and a final precipitation with ethanol and sodium acetate (Garcia-Martinez et al., 2004).
|
| Label |
Cy5
|
| Label protocol |
2-4 ug of total RNA was linearly amplified using the Low RNA Input Fluorescent Linear Amplification kit (Agilent Technologies™, Ca, USA). 2-3 ug of amplified cRNA was used as template for cDNA synthesis. cDNA was marked indirectly with “SuperScript™ Indirect cDNA Labeling System” (Invitrogen™, SanDiego, CA). The fluorophores used were Cy3 and Cy5 mono-reactiveDye (Amersham GE Healthcare™, Amersham UK) and dye incorporation was monitored by a Nanodrop spectrophotometer.
|
| |
|
| |
| Hybridization protocol |
A mixture of 200 to 300 pmol of the two samples labelled was concentrated in a Concentrator Plus (Eppendorf™, Hamburg, Germany). Competitive hybridization was performed in hybridization chambers AHC (ArrayIt Corporation, CA, USA) at 42°C overnight. (Prehybridization solution contained 3X SSC, 0.1% SDS and 0.1 mg/ml BSA; hybridization solution contained 5X SSC, 0.1% SDS and 0.1 mg/ml of salmon DNA. Microarrays were washed manually with different solutions containing different SSC 20X and SDS 10% concentrations (Sol.1: 2X SSC-0.1% SDS; Sol.2: 0.1X SSC-0.1% SDS; Sol.3: 0.1 SSC; Sol4: 0.01X SSC).
|
| Scan protocol |
Signal intensities of Cy3 and Cy5 were acquired with an Axon GenePix 4100A scanner (Molecular devices, CA, USA) using GenePix Pro v.6.1 software, at a resolution of 10 μm.
|
| Description |
Comparative gene expression of S. cerevisiae x S. kudriavzevii W27 at 12ºC with respect to 28ºC. Replicate C
|
| Data processing |
Raw data with a global background subtraction were generated from GenePix pro 6.0. Analyses were done using Acuity 4.0 software (Molecular Devices, CA, USA). The individual data sets were normalized to a log2 ratio value of 1. After normalization, data were filtered to remove spots flagged as not found. Only spots with at least two replicates were considered. Finally, replicates were combined and their medians were calculated. Genes with a two-fold log2 ratio values were considered to be significantly expressed.
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| |
|
| Submission date |
Dec 02, 2016 |
| Last update date |
Dec 12, 2016 |
| Contact name |
Jordi Tronchoni |
| E-mail(s) |
[email protected]
|
| Organization name |
CSIC
|
| Street address |
Avda. Agustín Escardino, 7
|
| City |
Paterna |
| ZIP/Postal code |
46980 |
| Country |
Spain |
| |
|
| Platform ID |
GPL22734 |
| Series (1) |
| GSE90793 |
Transcriptomic landscape of S. cerevisiae x S. kudriavzevii hybrids in low temperature winemaking. |
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